title: Signaling between mammalian adiponectin and a mosquito adiponectin receptor reduces Plasmodium transmission

city: New Haven

first: Yu-Min

last: Chuang

state: CT

address: 1 Gilbert St TAC S410

department: Infectious disease

institute: Yale University

summary: In this study we determine the effects of silencing adiponectin receptor in A. gambiae mosquitoes after taking Plasmodium gambiae infected blood meals. dsRNA (dsAdpR and dsGluc) microinjected A. gambiae mosquitoes were fed on P. berghei-infected Swiss Webster mice. Two days after a blood meal, the mosquitoes were collected for midgut dissection.

overall_design: Examination of the transcriptomic change in midguts after silencing adiponectin receptor

first: Erol

last: Fikrig


title: FEZ1 participates in human embryonic brain development by modulating neuronal progenitor subpopulation specification and migration

status: Public on Nov 10 2023

city: Singapore

first: John Chua

last: Jia En

address: 28 Medical Dr

department: Centre for Life Sciences

country: Singapore

institute: National University of Singapore

type: Expression profiling by high throughput sequencing

summary: Using bulk RNAseq and scRNAseq, we identifed gene expression changes between wildtype and FEZ1 knockout brain organoids.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for wildtype and FEZ1 knockout brain organoids derived from H1 human embryonic stem cells

first: Yinghua

last: Qu

first: Omer

last: An

first: Henry

last: Yang

first: Yi-Chin

last: Toh

first: John

middle: J

last: Chua


title: Expression profiles of oviductal mRNAs and lncRNAs in the follicular phase and luteal phase of sheep (Ovis aries) with two fecundity gene (FecB) genotypes

status: Public on Nov 08 2023

city: Yangzhou

first: Weihao

last: Chen

address: 88 Daxuenan Rd.

country: China

institute: Yangzhou University

summary: FecB (also known as BMPR1B) is a crucial gene in sheep reproduction, which has a mutation (A746G) that was found to increase the ovulation rate and litter size. The FecB mutation is associated with reproductive endocrinology, such mutation can control external estrous characteristics and affect follicle-stimulating hormone during the estrous cycle. Previous researches showed that the FecB mutation can regulate the transcriptomic profiles in the reproductive-related tissues including hypothalamus, pituitary, and ovary during the estrous cycle of Small Tailed Han sheep (STH). However, little research has been reported on the correlation between FecB mutation and the estrous cycle in STH sheep oviduct. To investigate the coding and non-coding transcriptomic profiles involved in the estrous cycle and FecB in the sheep oviduct, RNA sequencing was performed to analyze the transcriptomic profiles of mRNAs and long non-coding RNAs (lncRNAs) in the oviduct during the estrous cycle of STH sheep with mutant (FecBBB) and wild-type (FecB++) genotypes. In total, 21,863 lncRNAs and 43,674 mRNAs were screened.Together, our results can provide novel insights into the oviductal transcriptomic function against a FecB mutation background in sheep reproduction.

overall_design: All experimental sheep and were kept in a sheltered outdoor paddock and were provided with alfalfa hay and concentrate, with clear water available ad libitum, the TaqMan probe was firstly applied to genotype the sheep population (n = 890). Then, six FecB ++ sheep and six FecB BB with no significant differences in age, weight, and body size (2.5±0.5 years old; weight 70±5 kg) were randomly selected from  STH sheep with the FecB ++ genotype (n=142, wild type, w) and STH sheep with the FecB BB genotype (n=78, mutant type, M), respectively. All selected sheep were processed by estrous synchronization with Controlled Internal Drug Releasing Device (CIDR; progesterone 300 mg; Zoetis Australia Pty. Ltd., NSW, Australia) for 12 days. The six sheep, comprising three FecB ++ sheep and three FecB BB sheep, were euthanized within 45–48 h after CIDR removal (follicular phase) by administering a pentobarbital sodium overdose (1.5 mg/kg), the remaining six sheep were euthanized on day 9 after CIDR removal (luteal phase). Finally, the selected sheep were divided into four groups, including FecB ++ sheep in the follicular phase (wF), FecB ++ sheep in the luteal phase (wL), FecB BB sheep in the follicular phase (MF), and FecB BB sheep in the luteal phase (ML), based on their genotype record and estrous cycle. The oviduct isthmus tissues were collected for RNA-seq.

first: Zhifeng

last: Li


title: Stiffness-induced cancer-associated fibroblasts are responsible for immunosuppression in a PDGF ligand-dependent manner

city: Bron

first: Hector

last: Hernandez-Vargas

state: Rhone

address: 36 bis Rue de la Batterie

country: France

institute: Genomics Consulting

summary: Pancreatic ductal adenocarcinoma (PDAC) is associated with a vast stromal reaction that arises mainly from cancer-associated fibroblasts (CAFs) and promotes both immune escape and tumor growth. Here, we used a mouse model of KrasG12D mutation (KC) without or with deletion of the activin A receptor ALK4 (4KC), which strongly drives collagen deposition that leads to tissue stiffness. To understand this process, single cell transcriptomes were obtained from purified cancer associated fibroblasts (CAFs) and ductal tumor cells from KC and 4KC tissues. These analyses led to the identification of increased number of CAFs/Ducts receptor:ligand interactions under stiffness conditions and a population of stiffness-induced CAFs with a role in immunosuppression.

overall_design: Single cell RNA sequencing was performed in FACS purified ductal tumor cells from a pool of five KC or 4KC mice (LSL-KrasG12D/+;Ptf1a-Cre mice).

first: Pia

last: Gamradt

first: Ana

last: Hennino


title: Distinct baseline immune characteristics associated with responses to conjugated and unconjugated pneumococcal polysaccharide vaccines in older adults [bulk RNA-seq]

city: Farmington

first: Duygu

last: Ucar

address: 10 Discovery Dr

country: USA

institute: The Jackson laboratory for Genomic Medicine

summary: Pneumococcal infections cause serious illness and death among older adults. A capsular polysaccharide vaccine PPSV23 (Pneumovax®) and a conjugated polysaccharide vaccine PCV13 (Prevnar®) are used to prevent these infections, yet underlying immunological responses, and baseline predictors remain unknown. We recruited and vaccinated 39 older adults (>60 years) with PPSV23 or PCV13. Both vaccines induced strong antibody responses at day 28 and similar plasmablast transcriptional signatures at day 10, however, their baseline predictors were distinct. Analyses of baseline flow cytometry and RNA-seq data (bulk and single cell) revealed a novel baseline phenotype that is specifically associated with weaker PCV13 responses, characterized by i) increased expression of cytotoxicity-associated genes and increased CD16+ NK frequency; ii) increased Th17 and decreased Th1 cell frequency. Men were more likely to display this cytotoxic phenotype and mounted weaker responses to PCV13 than women. Baseline expression levels of a distinct gene set was predictive of PPSV23 responses. This first precision vaccinology study for pneumococcal vaccine responses of older adults uncovered novel and distinct baseline predictors that might transform vaccination strategies and initiate novel interventions.

overall_design:
In this study, we performed bulk RNA-sequencing of peripheral blood mononuclear cells (PBMCs) obtained from 30 healthy adults aged 60 years and older at four distinct time points. Out of the 30 participants, 14 individuals received the Prevnar 13 vaccine (Pfizer), a conjugated 13-valent vaccine, while the remaining 16 participants received the Pneumovax 23 vaccine (Merck), a non-conjugated 23-valent vaccine. The four time points considered in this study included: Day -7 (baseline), Day 1, Day 10, and Day 60 (± 5 days) post-vaccination.
***Submitter declares that the raw data will be available through dbGAP (phs002361) due to patient privacy concerns ***

first: Sathyabaarathi

last: Ravichandran


title: Genome-wide profiling of transcription factor activity in primary liver cancer using single cell ATAC sequencing

city: Bethesda

first: Amanda

last: Craig

state: Maryland

address: 37 Convent Dr. Bld.37/Rm.3044

institute: National Institutes of Health

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Genome wide chromatin profiling of 16 primary liver cancer patients was performed using scATAC-seq. The aim of this study was to assess transcription factor activity in malignant and microenviornment component cells of human primary liver cancers. scATAC-seq profiles reveal similar microenvironment composition, yet malignant cells have high levels of heterogeneity in TF activation within and between patients.  Further analysis of transcription factor activity determined nuclear receptors as crucial for maintaining hepatocytic features in HCC, while ETS factors are crucial for maintaining cholangiocytic features in iCCA.

first: Lichun

last: Ma

first: Mahler

last: Revsine

first: Limin

last: Wang

first: Sophia

last: Heinrich

first: Zachary

last: Rae

first: Allison

last: Ruchinskas

first: Kimia

last: Dadkhah

first: Whitney

last: Do

first: Shay

last: Behrens

first: Farid

middle: R

last: Mehrabadi

first: Maruhen

middle: A

last: Datsch Silveira

first: Dana

last: Dominguez

first: Marshonna

last: Forgues

first: Jonathan

middle: M

last: Hernandez

first: Jeremy

middle: L

last: Davis

first: Bao

last: Tran

first: Jens

middle: U

last: Marquardt

first: Michael

last: Kelly

first: Tim

middle: F

last: Greten

first: Xin

middle: W

first: Anuradha

last: Budhu


title: RNA profile of mouse intestinal organoids  upon deletion of aPKCs

status: Public on Nov 07 2023

city: New York

first: Jorge

last: Moscat

state: NY

address: 1188 York Avenue

department: Pathology and Laboratory Medicine

institute: WEILL CORNELL MEDICINE

summary: sgPrkci and Prkcz MTOs  were profiled in three biological replicates under normal conditions.

first: Yu

last: Muta

first: Anxo

last: Martinez

first: Angeles

last: Duran

first: Yuki

last: Nakanishi

first: Preeti

last: Arora

first: Tania

last: Cid

first: Marc

last: Ruiz

first: Hiroto

last: Kinoshita

first: Thekla

last: Cordes

first: Christian

last: Metallo

first: Timothy

last: Osborne

first: Masakazu

last: Yashiro

first: Kiyoshi

last: Maeda

first: Maria

middle: T

last: Diaz-Meco


title: Single-cell analysis predicts that mesenchymal ovarian cancer cells promote CD8+ T cell exhaustion through the LGALS3 - LAG3 axis

city: Ottawa

first: Edward

last: Yakubovich

state: Ontario

address: 451 Smyth Rd

country: Canada

institute: Ottawa Hospital Research Institute

summary: Cancer cells often metastasize by undergoing an epithelial-mesenchymal transition (EMT). Although abundance of CD8+ T-cells in the tumor microenvironment correlates with improved survival, mesenchymal cancer cells acquire greater resistance to antitumor immunity in some cancers. We hypothesized the EMT modulates the immune response to ovarian cancer. Here we show that cancer cells from infiltrated/inflamed tumors possess more mesenchymal cells, than excluded and desert tumors. We also noted high expression of LGALS3 is associated with EMT in vivo, a finding validated with in vitro EMT models. Dissecting the cellular communications among populations in the tumor revealed that mesenchymal cancer cells in infiltrated tumors communicate through LGALS3 to LAG3 receptor expressed by CD8+ T cells. We found CD8+ T cells express high levels of LAG3, a marker of T cell exhaustion. The results indicate that EMT in ovarian cancer cells promotes interactions between cancer cells and T cells through the LGALS3 - LAG3 axis, which could increase T cell exhaustion in infiltrated tumors, dampening antitumor immunity.

overall_design: OVCA420 cells (10,000/well) were plated into in 6-well plates. Cells were treated with 10ng/mL TGF-β1 (R&D Systems, #240-B-010), with treatment timed in such a way that all time-points were synchronized at the time of collection. Cells were passaged as needed to avoid confluence, and fresh TGF-β1 was added every two days with refreshed media. Cells were not passaged in the 2 days prior to final collection to avoid artifacts during sequencing. Single-cell suspensions were processed using the 10x Genomics Single Cell 3’ RNA-seq kit v3. Final libraries were sequenced on an Illumina sequencer after gene expression libraries were prepared according to the manufacturer’s protocol.

first: David

middle: P

last: Cook

first: Galaxia

last: Rodriguez

first: Barbara

middle: C

last: Vanderhyden


title: Loss of Cohesin regulator PDS5A reveals repressive role of Polycomb loops

city: Los Angeles

first: Oliver

last: Bell

state: California

address: 1450 Biggy St, NRT6510

institute: USC

summary: Polycomb Repressive Complexes 1 and 2 (PRC1, PRC2) are conserved epigenetic regulators that promote transcriptional silencing. PRC1 and PRC2 converge on shared targets, catalyzing repressive histone modifications. In addition, a subset of PRC1/PRC2 targets engage in long-range interactions whose functions in gene silencing are poorly understood. Using a CRISPR screen in mouse embryonic stem cells, we discovered that the cohesin regulator PDS5A links transcriptional silencing by Polycomb and 3D genome organization. PDS5A deletion impairs cohesin unloading and results in derepression of subset of endogenous PRC1/PRC2 target genes. Importantly, derepression is not associated with loss of repressive Polycomb chromatin modifications. Instead, loss of PDS5A leads to aberrant cohesin activity, ectopic insulation sites and specific reduction of ultra-long Polycomb loops. We infer that these loops are important for robust silencing at a subset of Polycomb target genes and that maintenance of cohesin-dependent genome architecture is critical for Polycomb regulation.

overall_design: 105 samples in total:  34 ChIP-Seq (17x2 replicates each), 12 RNA-Seq (6x2 replicates each), 6 ATACseq (2x3 replicates each), 43 HiC, 10 ChIPCapture (5x2 replicates each)

first: Daniel

last: Bsteh


title: Epithelial aPKC deficiency leads to stem cell loss preceding metaplasia in colorectal cancer initiation [ATAC-Seq]

first: Jorge

last: Moscat

summary: The early mechanisms of spontaneous tumor initiation that precede the accumulation of cancer mutations are largely unknown. We show that reduced aPKC levels correlate with stem cell loss and the induction of revival and metaplastic programs in serrated- and conventional-initiated pre-neoplastic lesions, which is perpetuated in colorectal cancers (CRC). Acute inactivation of PKCl/i in vivo and in organoids is sufficient to stimulate JNK in non-transformed intestinal epithelial cells (IECs), which promotes cell death and the rapid loss of the intestinal stem cells (ISC), including LGR5+ cells. This is followed by the accumulation of revival stem cells (RSC) at the bottom of the crypt and fetal metaplastic cells (FMC) at the top, creating two spatiotemporally distinct cell populations that depend on JNK-induced AP-1 and YAP. These cell lineage changes are maintained during cancer initiation and progression and determine the aggressive phenotype of human CRC irrespective of their serrated or conventional origin

overall_design: ATACseq samples from small intestinal epithelial cells from intestinal epithelial cell-specific PKCz and PKCz/PKCi DKO mice

first: Anxo

last: Martinez-Ordoñez

last: Cid-Diaz

first: Qixiu

last: Han

first: Angeles

last: Duran

first: Yu

last: Muta

first: Yuki

last: Nakanishi

first: Hiroaki

last: Kasashima

first: Ana

last: Albaladejo-Gonzalez

last: Torres-Moreno

first: José

last: García-Solano

first: Pablo

last: Conesa-Zamora


title: The Genetic and Epigenetic Features of Bilateral Wilms Tumor Predisposition

status: Public on Nov 09 2023

city: Birmingham

first: Changde

last: Cheng

state: AL

address: 1824 6th Ave S, Wallace Tumor Institute 520A

department: Medicine

institute: University of Alabama at Birmingham

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

overall_design: Refer to individual Series


title: Expression data from human pancreatic cancer cell lines and mouse orthotopic tumors generated using human PDAC cell lines replete and depleted for RBFOX2

city: Tampa

first: Karen

last: Mann

state: FL

address: 12902 Magnolia Drive

institute: Moffitt Cancer Center

summary:
RBFOX2 is an RNA binding protein that directs alternative splicing. In this study, we characterized RBFOX2-mediated alternative splicing in pancreatic cancer (PDAC)
In this dataset, we assayed gene-level and exon-level expression differences in pancreatic cancer cell lines replete and depleted for RBFOX2 expression and in 8 pairs of orthotopic pancreas tumors and related liver metastases generated from human 4039 or Panc1 cell lines replete or depleted for RBFOX2.

sample_organism:
Homo sapiens
Mus musculus

overall_design: 3 human PDAC cell line pairs replete (NTC) and depleted (KD) for RBFOX2 with no biological replicates were analyzed for differential gene expression and exon usage on a population level, comparing KD vs.NTC. Gene-level and exon-level data for human genes expressed in the population of orthotopic pancreas tumors generated using RBFOX2 depleted human 4039 or Panc1 cell lines was compared with gene-level and exon-level data for human genes expressed in the population of RBFOX2 replete orthotopic pancreas tumors generated using 4039 or Panc1 cells (KD vs. NTC). Array data from matched liver metastases and from two unmodified cell lines with low RBFOX2 expression were included in the normalization.


title: Distinct baseline immune characteristics associated with responses to conjugated and unconjugated pneumococcal polysaccharide vaccines in older adults [scRNA-seq]

first: Duygu

last: Ucar

overall_design:
Our study involved the single-cell RNA sequencing (scRNA-seq) profiling of peripheral blood mononuclear cells (PBMCs) obtained from 11 donors before PCV13 vaccination. Among these donors, six are strong responders (SR) to PCV13, and five are weak responders (WR).
***Submitter declares that the raw data will be available through dbGAP (phs002361) due to patient privacy concerns***

first: Sathyabaarathi

last: Ravichandran


title: Covalent PARylation of DNA base excision repair proteins regulates DNA demethylation

city: Basel

first: DBM Bioinformatics

last: Core Facility

state: BS

address: Hebelstrasse 20

department: Departement of Biomedicine

country: Switzerland

institute: University of Basel

summary: Poly(ADP-ribose) polymerases (PARP1 and PARP2) contribute to DNA base excision repair (BER) and DNA demethylation and has been implicated in epigenetic programming in early mammalian development. Recently, proteomic analyses identified BER proteins to be covalently poly-ADP-ribosylated by PARPs. The role of this posttranslational modification in the BER process is unknown. Here, we show that PARP1 senses AP-sites and SSBs generated during TET-TDG mediated active DNA demethylation and covalently attaches PAR to each BER protein engaged. Covalent PARylation dissociates BER proteins from DNA, which accelerates completion of the repair process. Consistently, inhibition of PARylation in mESC resulted both in reduced locus specific TET-TDG-targeted DNA demethylation, and in reduced general repair of random DNA damage. Our findings establish a critical function of covalent protein PARylation in coordinating molecular processes associated with dynamic DNA methylation.

overall_design: Single-strand break sequencing of 2i-cultivated murine embryonic stem cells (+/- TDG), treated with PARP-inhibitor Talazoparib

first: Jianming

last: Xu

first: Simon

middle: D

last: Schwarz

first: Primo

last: Schär


title: Epithelial aPKC deficiency leads to stem cell loss preceding metaplasia in colorectal cancer initiation [scRNA-Seq]

first: Jorge

last: Moscat

overall_design: WT (Prkcif/fPrkcif/f, Prkczf/f, and  Prkcif/f mice),   LKO  (Prkcif/f;Villin-Cre),  ZKO (Prkczf/f;Villin-Cre) and DKO (Prkcif/fPrkcif/f;Villin-Cre) mice small intestines were profiled for scRNAseq

first: Hiroto

last: Kinoshita

first: Anxo

last: Martinez-Ordoñez

first: Tania

last: Cid-Diaz

last: Han

first: Masakazu

last: Yashiro

first: Kiyoshi

last: Maeda

first: Daniel

first: Maria

last: Diaz-Meco


title: Cross-species comparative transcriptomics examining the role of Type-2 immunity in acute colitis

status: Public on Nov 06 2023

city: Palo Alto

first: Avnesh

last: Thakor

address: 3155 Porter Dr

department: Radiology

institute: Stanford Medicine

summary: Most preclinical models of IBD were developed in mice which share 85% genomic homology with humans and are genetically tractable. However, rats have historically been the model of choice for immunologic, cardiovascular, and neurologic diseases because they more closely recapitulate human physiology6. Recent work on the role of microbial dysbiosis in IBD has also shown that rat microbiota is more closely related to those of humans than mice7. Taken together, this suggests that a single preclinical model may not be optimal for delineating the molecular pathways involved in IBD pathogenesis. Previous research in rat models of IBD have focused primarily on characterization of clinical score and histopathological findings8, with a lack of systematic investigation into potential molecular pathways affected, thereby making it difficult to integrate this model with other preclinical and clinical research. In the current study, we have undertaken a systematic transcriptomic analysis of a rat model of experimental colitis along with a three-way comparison between an established mouse preclinical model and available human clinical datasets in an effort to identify conserved immunological pathways in IBD.

overall_design:
IBD was induced using chemical DSS at a concentration of 5.5 % w/v in drinking water for rats for 6 days. At the end of day 6 rats were sacrified and whole colonic tissue was collected and homogenized in trizol (Thermofisher Scientific, USA). RNA extraction was then done using RNA mini kit (Qiagen, Germany) according to the manufacturer’s guidelines with the quality of RNA evaluated using an Agilent bioanlyzer 2100 system. Samples with RNA integrity number (RIN) ≥ 6.8 were considered for library preparation.
IL: distal ileum
RC: Right (proximal) colon
LC: Left (distal) colon

first: Abantika

last: Ganguly

first: Steven

last: Levitte

first: Shobha

last: Regmi

first: Benjamin

last: William Dulken

first: Alix

last: Andrea Guevara Tique

first: Jing

last: Wang

middle: S


title: Collective signalling drives rapid jumping between cell states

city: London

first: Vlatka

last: Antolovic

address: Gower Street

department: MRC Laboratory for Molecular Cell Biology

country: United Kingdom

institute: University College London

summary: A continuous single cell transcriptomic time course of Dictyostelium discoideum early development.

overall_design: To capture development in a continuous manner, we collected cells from plaques of cells feeding on their bacterial food source. We collected a continuous streak of cells, from the bacterial zone through to the mounds, then generated single cell transcriptomes for 5329 cells.

last: Antolović

first: Tchern

last: Lenn

first: Elizabeth

last: Westbrook

first: Jonathan

last: Chubb


title: Activated CD4 T cells in tuberculosis express OX40, a target for host-directed immunotherapy [GFP_HI_LO_scRNASeq]

city: Minneapolis

first: Tyler

last: Bold

state: MN

address: 2101 6th St SE

institute: University of Minnesota

summary: Define genes differentially expressed by Nur77-GFP HI and LO CD4 T cells FACS sorted from the lungs of Mycobacterium tuberculosis-infected mice

overall_design: M. tuberculosis infected Nur77-GFP mice at day 28 of infection were injected with an oligonucleotide (ADT/HTO/FBT) labeled anti-CD45.2 antibody immediately prior to euthanasia to identify cells in the lung vasculature. Lung cell homogenates were stained with fluorescent antibodies to identify CD44hi, CD4+ T cells, then run on a Sony MA900 cell sorter, with gating on either highest or lowest 1/3 of population-wide GFP expression. FACS-sorted GFP HI or LO cells underwent scRNA-seq, with CITE-Seq to identify oligonucleotide (HTO/FBT) labeled IV+/vasculature cells and with TCR library corresponding to mouse replicate 1.

first: Bold

last: Tyler


title: Activated CD4 T cells in tuberculosis express OX40, a target for host-directed immunotherapy [Human_CSF_T_cell_scRNASeq]

summary: Define genes expressed by CD4 and CD8 T cells FACS sorted from a CSF specimen of a human subject with HIV-associated tuberculous meningitis

overall_design: Cells from the CSF of an individual treated for HIV-associated TB meningitis was obtained from a subject enrolled in the HARVEST (ISRCTN15668391) treatment study. Cells were obtained on either the day of study enrollment, initially stored at 4˚C, then cells were isolated by centrifugation at 300xg for 10 minutes within 6 hours of collection. The CSF cell pellet used for scRNA-Seq was resuspended in freezing medium (FCS with 10% DMSO) prior to cryopreservation at -80˚C using a Mr. Frosty and final storage at -150˚C. Within 6 months of cryopreservation, cells were transported at -50˚C, cryo-recovered, washed with complete RPMI medium, and labeled with a fluorescent viability dye and antibodies for sorting on a MA900 Multi-Application Cell Sorter (Sony Biotechnology) into DMEM with 40% FBS.


title: Effect of depletion of FOXL2 on gene expression in KGN cells

city: PARIS

first: Anne Laure

last: Todeschini

address: 15 rue hélène brion

institute: CNRS/Université Paris Cité

summary: To investigate the  targets of FOXL2 in human granulosa cells, we performed Knock-Down experiment in established homozygous cell lines for FOXL2 WT (KGN).

overall_design: We  performed knock-down of FOXL2 with siRNA followed by RNAseq .

first: Anne

first: Reiner

last: Veitia


title: An Engineered Niche Delineates Metastatic Potential of Breast Cancer

city: Ann Arbor

first: Sophia

last: Orbach

state: MI

address: 1600 Huron Parkway

institute: University of Michigan

summary:
We aimed to identify metastatic disease prior to the formation of an overt secondary tumor in triple-negative breast cancer using sister cell lines 4T1 (metastatic), 4T07 (invasive, non-metastatic), and 67NR (non-metastatic).
We used a porous, polycaprolactone scaffold, that serves as an engineered metastatic niche, to identify metastatic disease through the changing microenvironment.

overall_design: Comparative gene expression profiling analysis of RNA-seq data of scaffolds, lungs, spleens, and primary tumors of mice inoculated with the sister cell lines or a healthy control

first: Lonnie

last: Shea


title: Activated CD4 T cells in tuberculosis express OX40, a target for host-directed immunotherapy [GFP_HI_LO_BULKRNASeq]

summary: Define genes differentially expressed by Nur77-GFP HI and LO CD4 T cells FACS sorted from the lungs of Mycobacterium tuberculosis-infected mice with bulk RNA seq

overall_design: M. tuberculosis infected Nur77-GFP mice at day 28 of infection. Lung cell homogenates were stained with fluorescent antibodies to identify CD44hi, CD4+ T cells, then run on a Sony MA900 cell sorter, with gating on either highest or lowest 1/3 of population-wide GFP expression. FACS-sorted GFP HI or LO cells underwent bulk RNA-Seq.


title: Multi-Omic and Functional Analysis for Classification and Treatment of Sarcomas with FUS-TFCP2 or EWSR1-TFCP2 Fusions

city: Heidelberg

first: Claudia

last: Scholl

address: Im Neuenheimer Feld 581

country: Germany

institute: German Cancer Research Center (DKFZ)

summary: Linking clinical multi-omics analyses with mechanistic studies may improve the understanding of rare cancers. We leverage two precision oncology programs to investigate rhabdomyosarcoma with FUS/EWSR1-TFCP2 fusions, an orphan malignancy without effective therapies. All tumors exhibit outlier ALK expression, partly accompanied by intragenic deletions and aberrant splicing resulting in truncated ALK variants that are oncogenic and sensitive to ALK inhibitors. Additionally, recurrent CKDN2A/MTAP co-deletions provide a rationale for CDK4/6- and PRMT5-targeted therapies. Functional studies show that FUS-TFCP2 blocks myogenic differentiation, induces transcription of ALK and truncated TERT, and inhibits DNA repair. Unlike other fusion-driven sarcomas, TFCP2-rearranged tumors exhibit genomic instability and signs of defective homologous recombination. DNA methylation profiling indicate a close relationship with undifferentiated sarcomas. Finally, in two patients, sarcoma was preceded by benign lesions carrying FUS-TFCP2, providing insight into stepwise sarcomagenesis. This study illustrates the potential of linking precision oncology with preclinical research to gain insight into the classification, pathogenesis, and therapeutic vulnerabilities of rare cancers.

overall_design:
The RNA-seq and ACT-seq data from cell lines generated in this study have been deposited under this accession code.
RNA-sequencing: MCF10A (1x10*6) or SCP-1 (5x10*6) cells stably transduced in triplicate (MCF10A) or three to nine replicates (SCP-1) with EV, FUS-TFCP2, EWSR1-TFCP2, FUS, EWSR1, or TFCP2 were seeded in 10- or 15-cm dishes, and RNA was isolated the next day using the RNeasy Mini Plus Kit (Qiagen). Library preparation and 125-nt paired-end read sequencing on an Illumina HiSeq 4000 system were performed at the DKFZ Genomics and Proteomics Core Facility.
ACT-seq: MCF10A cells were transduced with two different lentiviral vectors to obtain four biological replicates expressing HA-FUS-TFCP2 or EV (R1–R4) and three biological replicates expressing HA-TFCP2 (R2–R4). For each replicate, ACT-seq was performed with HA-Tag (C29F4) rabbit mAb (Cell Signaling, #3724), HA-Tag (F-7) mouse mAb (Santa Cruz, sc-7392), anti-histone H3 (acetyl K27) antibody – ChIP grade (Abcam, ab4729), and rabbit IgG polyclonal antibody (Merck, #PP64) as negative control.
Further details can be found in the Supplementary Information of the related manuscript.


title: Activated CD4 T cells in tuberculosis express OX40, a target for host-directed immunotherapy

platform_organism:
Homo sapiens
Mus musculus


title: Genomic, transcriptomic, functional, and mechanistic characterization of rhabdomyosarcoma with FUS-TFCP2 or EWSR1-TFCP2 fusions (RNA-Seq)

summary: Many rare cancers are not well understood, and pathogenesis-directed therapies are often lacking, resulting in poor patient outcomes. Leveraging two (inter)national precision oncology trials that enroll large numbers of rare cancers, we investigated the clinical, histopathologic, molecular, and functional characteristics of rhabdomyosarcoma (RMS) with fusions of FUS or EWSR1 to the TFCP2 transcription factor, a recently discovered ultra-rare entity whose classification, pathogenesis, and optimal treatment are unclear. Unusually for fusion-driven sarcomas, most cases had highly rearranged genomes, including chromothripsis, and signs of defective homologous recombination DNA repair. All tumors showed extreme expression of a truncated TERT variant and the ALK receptor tyrosine kinase, which was additionally affected by intragenic deletions and aberrant splicing, resulting in the expression of shortened variants in 58% of cases. Three ALK variants were oncogenic in immortalized cells, and patient-derived tumor cells expressing two variants responded to certain ALK inhibitors. Other recurrent alterations included CDKN2A/MTAP co-deletions and mutations in PAPPA2, encoding an IGFBP5-specific proteinase, in 67% and 25% of cases, respectively. DNA methylation analysis, along with 19 other soft-tissue sarcoma classes, revealed a close relationship with undifferentiated sarcoma but not other RMS subtypes, suggesting that TFCP2-rearranged RMS is a separate entity, possibly arising from a distinct cell of origin. Expression of TFCP2 fusions in immortalized human cells blocked late myogenic differentiation and significantly induced genes that were also highly expressed in patient tumors, including ALK, TERT, and two known regulators of skeletal muscle cells, IGFBP5 and PTH1R. Genome-wide chromatin profiling demonstrated direct binding of FUS-TFCP2 to the ALK and TERT loci outside their regular promoters, which correlated with the expression of alternative transcript variants. Finally, FUS-TFCP2 inhibited the repair of DNA double-strand breaks in immortalized cells, rendering them sensitive to treatment with cisplatin. This study provides insight into the pathogenesis and therapeutic vulnerabilities of a new RMS subtype and illustrates the value of linking comprehensive molecular diagnostics and functional annotation within multicenter consortia to improve the understanding and clinical management of rare cancers.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for MCF10A and SCP-1 cells transduced with FUS-/EWSR1-TFCP2, wild-type TFCP2/FUS/EWSR1, and empty vector

first: Julia

last: Schoepf

first: Sebastian

last: Uhrig

first: Christoph

middle: E

last: Heilig

first: Kwang-Seok

last: Lee

first: Tatjana

last: Walther

first: Alexander

last: Carazzato

first: Anna

last: Dobberkau

first: Dieter

last: Weichenhan

last: Plass

first: Mark

last: Hartmann

first: Gaurav

last: Diwan

first: Zunamys

last: Carrero

last: Ball

first: Tobias

last: Hohl

first: Thomas

last: Kindler

first: Patricia

last: Rudolph-Hähnel

last: Nilsson

first: Ingrid

last: Øra

first: Roland

last: Imle

first: Ana

last: Banito

first: Robert

last: Russell

first: Barbara

last: Jones

first: Daniel

middle: B

last: Lipka

first: Hanno

last: Glimm

last: Hübschmann

first: Wolfgang

first: Stefan

last: Fröhling


title: Inguinal white adipose tissue expression profiles of WT mice and Slc35d3 adipose tissue  knock-in mice

city: Beijing

first: Yibo

last: Wang

state: Beijin

address: 167 Beilishi Road, Xicheng District, Beijing

institute: Fuwai Hospital, National Center for Cardiovascular Diseases, Chinese Academy of Medical Sciences & Peking Union Medical College

summary: Slc35d3 adipose-specific knockin (SAKI) mice were generated by crossbreeding mice carrying the Slc35d3 transgene locus controlled by loxp-stop-loxp sequence (LSL-Slc35d3) with adiponectin-derived Cre mice, which is a mouse model that expresses the cre enzyme specifically in adipose tissues. Male 6-week-old Slc35d3 adipose-specific knockin mice (SAKI group) and their control mice (wild-type mice, WT group) were fed a normal standard chow diet (NCD, 10 kal% fat, D09100304, Research Diets, Inc) for 10 weeks. All mice reached the experimental endpoint at 16 weeks of age. Mice in WT group were used as controls. Upon reaching the experimental endpoint, inguinal white adipose tissue (IngWAT) from three SAKI mice and three WT mice were removed for RNA sequencing.

overall_design: Slc35d3 adipose-specific knockin (SAKI) mice were generated by crossbreeding mice carrying the Slc35d3 transgene locus controlled by loxp-stop-loxp sequence (LSL-Slc35d3) with adiponectin-derived Cre mice, which is a mouse model that expresses the cre enzyme specifically in adipose tissues. To investigate the differences in adipose tissue expression profiles between Slc35d3 adipose-specific knock-in (SAKI) mice and wild-type (WT) mice, 6-week-old male mice were fed a normal standard chow diet (NCD, 10 kal% fat, D09100304, Research Diets, Inc) for 10 weeks. WT mice were used as controls. All mice reached the experimental endpoint at 16 weeks of age.

first: Hongrui

first: Liang

last: Yu


title: Upregulated Hepatic Lipogenesis from Dietary Sugars Supplies Palmitic Acid to the Developing Brain of Mice fed Low Palmitic Acid from Birth

city: Toronto

first: Mackenzie

last: Smith

address: 1 King's College Circle

department: Nutritional Sciences

institute: University of Toronto

summary: Background: Palmitic acid (PAM) can be provided in the diet or synthesized via de novo lipogenesis (DNL) primarily from glucose. Preclinical work on the origin of brain PAM during development is scarce and contrasts results on the origin of adult brain PAM. Here, we utilize naturally occurring carbon isotope ratios (13C/12C; δ13C) and RNA sequencing to uncover the origin of brain PAM, and pathways involved in maintaining brain PAM, respectively, during development. maintaining brain PAM, respectively, during development.    Methods: Dams were equilibrated onto diets low (<2%), medium (47%) or high (>95%) in PAM prior to breeding. Dietary PAM was depleted in δ13C, while dietary sugars were enriched. Offspring stayed on the respective dam diet and were euthanized at postnatal day 0, 10, 21, and day 35. Pup liver and brain fatty acids were quantified, after which, tissue δ13C-PAM was measured by compound specific isotope analysis. Day 35 tissue RNA was sequenced on a NovaSeq S4 Flowcell.    Results: Although PAM levels in the liver reflected levels of dietary PAM, PAM was maintained in total and individual brain phospholipid fractions across diet groups at all timepoints. Tissue δ13C-PAM was enriched overall and augmented in mice fed low PAM, compared to medium and high PAM suggesting that DNL from dietary sugars maintained the majority of the brain PAM pool. Furthermore, DNL pathways were upregulated in mice fed low compared to high PAM in the liver, but not the brain at day 35.   Conclusions: Lipogenesis from dietary sugars maintains the majority of brain PAM during development and is augmented in mice fed low PAM from birth. Importantly, hepatic lipogenesis from dietary sugars determines PAM availability to the developing brain when low in the diet – a compensatory mechanism identified to maintain total brain PAM pools compared to periphery which ultimately suggests an importance of brain PAM regulation during development.

overall_design: Pairwise comparisons of RNAseq data of brain and liver tissue from 35-day old BALB/c mice fed custom diets containing low (<2%), medium (~47%) and high (>95%) PAM

first: Chuck

last: Chen

first: Chiraag

last: Gohel

first: Giulia

last: Cisbani

middle: K

first: Kimia

last: Rezaei

first: Richard

last: Bazinet


title: Selective oxytocin receptor activation prevents prefrontal circuit dysfunction and social behavioral alterations in response to chronic prefrontal cortex activation in rats

status: Public on Nov 05 2023

first: Roland

last: Schmucki

address: Grenzacherstrase

institute: F. Hoffmann - La Roche AG

summary: Social behavioral changes are a hallmark of several neurodevelopmental and neuropsychiatric conditions, nevertheless the underlying neural substrates of such dysfunction remain poorly understood. Building evidence points to the prefrontal cortex (PFC) as one of the key brain regions that orchestrates social behavior. We used this concept with the aim to develop a translational rat model of social-circuit dysfunction, the chronic PFC activation model (CPA). Chemogenetic designer receptor hM3Dq was used to induce chronic activation of the PFC over 10 days, and the behavioral and electrophysiological signatures of prolonged PFC hyperactivity were evaluated. To test the sensitivity of this model to pharmacological interventions on longer timescales, and validate its translational potential, the rats were treated with our novel highly selective oxytocin receptor (OXTR) agonist RO6958375, which is not activating the related vasopressin V1a receptor. CPA rats showed reduced sociability in the three-chamber sociability test, and a concomitant decrease in neuronal excitability and synaptic transmission within the PFC as measured by electrophysiological recordings in acute slice preparation. Sub-chronic treatment with a low dose of the novel OXTR agonist following CPA interferes with the emergence of PFC circuit dysfunction, abnormal social behavior and specific transcriptomic changes. These results demonstrate that sustained PFC hyperactivity modifies circuit characteristics and social behaviors in ways that can be modulated by selective OXTR activation and that this model may be used to understand the circuit recruitment of prosocial therapies in drug discovery.

overall_design: Three groups of rats (N=4 for each group) with or without repeated chemogenetic activation of the PFC over 10 days and subsequent repeated treatment with an oxytocin receptor agonist or vehicle for 7 days

first: Philipp

last: Janz

first: Martin

last: Ebeling

first: Christophe

last: Grundschober

first: Madhurima

last: Benekareddy

first: Megana

last: Prasad

first: Fabian

last: Koechl


title: Purine and carbohydrate availability drive Enterococcus faecalis fitness in wound infections.

city: Singapore

first: Casandra

last: Tan

address: 60 Nanyang Drive

institute: SCELSE

overall_design: Refer to individual Series


title: Purine  and carbohydrate availability drive Enterococcus faecalis fitness in wound infections [dataset 1]

summary: Enterococcus faecalis is commonly isolated from different wound types. However, despite its prevalence, the pathogenic mechanisms of E. faecalis during wound infections remain poorly understood. We adopted an in vivo E. faecalis transposon sequencing and RNA sequencing approach to identify fitness determinants that are crucial for replication and persistence of E. faecalis during wound infections in a mouse model. We demonstrated that E. faecalis purine biosynthesis genes are important for replication as purine metabolites are low in wounds during the early phase of wound infections. Furthermore, we found that the E. faecalis MptABCD phosphotransferase system (PTS) involved in the uptake of galactose and mannose, is crucial for E. faecalis persistence in wounds, and that carbohydrate availability changes as the infection progresses. To understand how MptABCD PTS contributes to the reduced fitness observed during E. faecalis wound persistence, we performed an in vitro transcriptomic analysis using the galactose/mannose PTS gene deletion mutant (∆mptD). When mannose was the sole carbohydrate source, shikimate and purine biosynthesis genes in the ∆mptD mutant were downregulated compared to the isogenic wild-type strain, indicating that mannose transport is interconnected with shikimate and purine biosynthesis. Together, our results suggest that dynamic and temporal microenvironment changes at the wound site affects pathogenic requirements and mechanisms of E. faecalis during infections and raise the possibility of inhibiting purine biosynthesis and/or MptABCD PTS to control wound infections.

overall_design: We aim to identify E. faecalis genes that are differentially expressed between E. faecalis inoculum used for mouse wound infection and E. faecalis harvested from 8h post-infection mouse wound samples.

last: TAN

first: Kimberly

last: KLINE


title: RNA sequence in alveolar macrophage of mice treated with HDM or Combined with PR8

city: Shanghai

first: Haibo

last: Zhang

address: Dongchuan road 800

institute: Shanghai Jiaotong University

summary: To research the different expressive genes in alveolar macrophage during asthma excerbation, we build asthma excerbation mice use HDM combined with PR8. we sepreated and collected the alveolar macrophages in BALF of mice then test the RNA expression. in this data, LysMCre Cul5fl/fl mice also challenaged with HDM combined with PR8 to compared the different expressive genes in alveolar macrophage.

overall_design: LysMCre Cul5fl/fl mice and Cul5fl/fl mice were treated HDM or HDM+PR8 to build asthma or asthma excerbation mice models, AMs were collected and RNA-seq were performed and analysised. these data have four groups. AMs from Cul5fl/fl mice treated by PBS named as WT-C. AMs from Cul5fl/fl mice treated by HDM named as WT-H. AMs from Cul5fl/fl mice treated by HDM combined with PR8 named as WT-H-P. AMs from LysMCre Cul5fl/fl mice treated by HDM combined with PR8 named as KO-H-P.


title: Activated CD4 T cells in tuberculosis express OX40, a target for host-directed immunotherapy [Tetramer_scRNASeq]

first: Tyler

last: Bold

summary: Define genes expressed by CD4 T cells in the lungs of Mycobacterium tuberculosis-infected mice

overall_design: Lung cell homogenate from M. tuberculosis infected mice at day 28 of infection were stained with fluorophore-labeled tetramers for Ag85B and ESAT-6 peptide, then magnetically enriched for fluorphore labeled events. Tetramer-enriched cells underwent scRNA-seq.

first: Bold

last: Tyler


title: Genomic, transcriptomic, functional, and mechanistic characterization of rhabdomyosarcoma with FUS-TFCP2 or EWSR1-TFCP2 fusions (ACT-Seq)

first: Claudia

last: Scholl

overall_design: Antibody-guided chromatin tagmentation sequencing (ACT-seq) for FUS/EWSR1-TFCP2 and wild-type TFCP2/FUS/EWSR1 binding sites as well as H3K27ac histone marks

first: Julia

last: Schoepf

first: Sebastian

last: Uhrig

first: Christoph

last: Heilig

first: Kwang-Seok

last: Lee

first: Tatjana

last: Walther

first: Alexander

last: Carazzato

first: Anna

last: Dobberkau

first: Dieter

last: Weichenhan

last: Plass

first: Mark

last: Hartmann

first: Gaurav

last: Diwan

first: Zunamys

last: Carrero

last: Ball

first: Tobias

last: Hohl

first: Thomas

last: Kindler

first: Patricia

last: Rudolph-Hähnel

last: Nilsson

first: Ingrid

last: Øra

last: Imle

first: Ana

last: Banito

first: Robert

last: Russell

first: Barbara

last: Jones

first: Daniel

last: Lipka

first: Hanno

last: Glimm

last: Hübschmann

first: Wolfgang

first: Stefan

last: Fröhling


title: iPSC-derived hypoimmunogenic T cells enriched with tissue resident memory T cells powerfully attack cervical cancer

city: Stanford

first: Kyomi

last: Igarashi

state: CA

address: 265 Campus Drive

department: Genetics

institute: Stanford University

summary: We generated HLA-edited functionally rejuvenated human papilloma virus-specific CTLs (HPV-rejTs). These rejTs successfully suppressed recipient immune rejection, while retaining more robust cytotoxicity than that of original CTLs. To clarify this, single-cell RNA sequencing was performed.

overall_design: Original HPV-CTL clone and HLA-editied HPV-rejTs were generated then analyzed using scRNA-seq

first: Miki

last: Ando

first: Yoshiki

last: Furukawa


title: Purine  and carbohydrate availability drive Enterococcus faecalis fitness in wound infections [dataset 2]

first: Casandra

last: Tan

overall_design: We aim to identify E. faecalis genes that are differentially expressed between wild-type OG1RF and ∆mptD mutant when grown in Tryptic Soy Broth without dextrose (TSBd) supplemented with 1% (w/v) mannose.

last: TAN

first: Kimberly

last: KLINE


title: single nuclei RNA-seq at RA in juvenile zebra finch and owl finch F1 hybrid birds

status: Public on Nov 04 2023

city: Sapporo

first: Noriyuki

last: Toji

state: Hokkaido

address: North10-West8, Kita-ku,

department: Department of Biological Sciences

country: Japan

institute: Hokkaido university

summary: To investigate the cellular basis of parental species bias at birdsong, we performed single nuclei RNA-seq for six zebra finch and owl finch F1 hybrid juvenile birds.

sample_organism: Taeniopygia guttata x Taeniopygia bichenovii

overall_design: Comparative gene expression profiling analysis of single nuclei RNA-seq data among individual birds.

first: Hongdi

last: Wang

first: Azusa

last: Sawai

first: Yasuhiro

last: Go

first: Kazuhiro

last: Wada


title: Unraveling the diversity and functions of tissue-resident plasma cells (RNA-seq of mouse antibody-secreting cells)

status: Public on Nov 12 2023

city: Parkville

first: Gordon

last: Smyth

state: Vic

address: 1G Royal Pde

department: Bioinformatics

country: Australia

institute: Walter and Eliza Hall Institute of Medical Research

summary: Antibody-secreting plasma cells (PCs) are generated in secondary lymphoid organs, but are reported to reside in an emerging range of anatomical sites. Analysis of the transcriptome of different tissue-resident (Tr)PC populations revealed that they each have their own transcriptional signature indicative of functional adaptation to the host tissue environment. In contrast to expectation, all TrPCs were extremely long-lived, regardless of their organ of residence, with the longevity influenced by intrinsic factors like the immunoglobulin isotype. This study reveals that extreme longevity is an intrinsic property of TrPCs whose transcriptome is imprinted by signals received both at the site of induction and within the tissue of residence.

overall_design: RNA-seq gene expression profiling analysis was performed on tissue-resident antibody-secreting cells (plasma cells and plasmablasts. Ex vivo cells were sorted by Blimp1-gfp+ CD138+ or Blimp1-gfp+ CD98+. Plasma cells were isolated from bone marrow (1 biological replicate), colon (3 replicates), small intestine (3 replicates), mammary gland (3 replicates), Peyer’s patches (2 replicates) and spleen (2 replicates). Plasmablasts were isolated from Peyer's patches (2 replicates) and spleen (2 replicates). Bone marrow plasma cells were further sorted into IgA and non-IgA isotypes (3 biological replicates).

first: Ilariya

last: Tarasova

first: Julie

last: Tellier

first: Stephen

last: Nutt


title: single nuclei RNA-seq at Area X in juvenile zebra finch and owl finch F1 hybrid birds

platform_organism: Taeniopygia guttata x Taeniopygia bichenovii


title: Profiling of RNA-binding protein binding sites by in-situ reverse transcription-based sequencing

city: Chicago

first: Yanming

last: Chen

address: 929 E 57th Street

institute: The University of Chicago

summary: RNA-binding proteins (RBPs) regulate diverse cellular processes by dynamically interacting with RNA targets. However, effective methods to capture both stable and transient interactions between RBPs and their RNA substrates are still lacking, especially when the interaction is dynamic or materials are limited. Here we present an assay of reverse transcription-based RBP binding sites sequencing (ARTR-seq), which relies on in-situ reverse transcription of RBP-bound RNAs guided by antibodies to identify RBP binding sites. ARTR-seq avoids ultraviolet cross-linking and immunoprecipitation, allowing for efficient and specific identification of RBP binding sites from as few as 20 cells or a tissue section. Taking advantage of rapid formaldehyde fixation, ARTR-seq enables capturing the dynamic binding of RBPs over a short period of time, as demonstrated by the discovery of dynamic RNA binding of G3BP1 during stress granule assembly on a timescale as short as 10 min.

overall_design: We first developed an assay of reverse transcription-based RBP binding sites sequencing (ARTR-seq) to identify PTBP1 binding sites in HepG2 cells. To validate the PTBP1 binding sites identified by ARTR-seq, we performed ARTR-seq for PTBP1 knocked down HepG2 cells. To determine the efficiency of ARTR-seq for small cell number inputs, we constructed ARTR-seq libraries using different numbers of HepG2. We next constructed ARTR-seq libraries to identify the binding sites for several RBPs, including RBFOX2 and HNRNPC in HepG2 cells, and YTHDF1, YTHDF2 and YTHDC1 in HeLa cells. We then generated ARTR-seq libraries in a section of OCT-embedded E11 mouse embryos to identify the RBFOX2 binding sites. To monitor the dynamic binding of G3BP1 during the stress granules assembly, we performed ARTR-seq for the HeLa cells treated with sodium arsenite for various time intervals.

first: Yan-Ming

first: Yu

last: Xiao

first: Chuan

last: He


title: Bulk RNA-sequencing of ILC2s from IPF patients

city: Suita-shi

first: Kazuyo

last: Moro

state: Osaka

address: 2-2 Yamadaoka

department: Department of Microbiology and Immunology

institute: Osaka University Graduate School of Medicine

summary: To investigate the comprehensive mRNA expression profile of ILC2s from IPF patients, we performed bulk RNA-sequencing analysis of ILC2s sorted from IPF patients and healthy controls.

overall_design: Blood ILC2s of 12 healthy controls and 19 IPF patients were sorted and library construction and sequencing were performed. One sample of IPF patients was omitted before sequencing because of the low quality of RNA.

first: Natsuko

last: Otaki

first: Yasutaka

last: Motomura


title: Genome-resolved metatranscriptomics reveals conserved root colonization determinants in a synthetic microbiota

status: Public on Nov 03 2023

city: Cologne

first: Fantin

last: Mesny

address: Carl-von-Linné-Weg 10

department: Plant-Microbes interaction

institute: Max Planck Institute for Plant Breeding Research

summary: The identification of processes activated by specific microbes during microbiota colonization of plant roots has been hampered by technical constraints in metatranscriptomics. These include lack of reference genomes, high representation of host or microbial rRNA sequences in datasets, or difficulty to experimentally validate gene functions. Here, we recolonized germ-free Arabidopsis thaliana with a synthetic, yet representative root microbiota comprising 106 genome-sequenced bacterial and fungal isolates. We used multi-kingdom rRNA depletion, deep RNA-sequencing and read mapping against reference microbial genomes to analyse the in-planta metatranscriptome of abundant colonizers. We identified over 3,000 microbial genes that were differentially regulated at the soil-root interface. Translation and energy production processes were consistently activated in planta, and their induction correlated with bacterial strains’ abundance in roots. Finally, we used targeted mutagenesis to show that several genes consistently induced by multiple bacteria are required for root colonization in one of the abundant bacterial strains (a genetically tractable Rhodanobacter). Our results indicate that microbiota members activate strain-specific processes but also common gene sets to colonize plant roots.

sample_organism:
Arabidopsis thaliana
unidentified soil bacteria

overall_design: Comparative gene expression profiling analysis of RNA-seq data from soil and root samples inoculated with Synthetic microbial consortia

first: Nathan

last: Vannier

first: Felix

last: Getzke

first: Guillaume

last: Chesneau

first: Laura

last: Dethier

first: Jana

last: Ordon

first: Thorsten

last: Thiergart

first: Stéphane

last: Hacquard


title: hUC-MSCs-derived MFGE8 ameliorates locomotor dysfunction via inhibition of ITGB3/NF-κB signaling in an NMO mouse model

city: Guangzhou, Guangdong

first: Longhui

last: Deng

address: No. 1 Helix 4th Road, International Biological Island

institute: Guangzhou SALIAI Stemcell Science and Technology Co., Ltd.

summary: Neuromyelitis optica (NMO) is a severe autoimmune inflammatory disease of the central nervous system that affects motor function and causes relapsing disability. Human umbilical cord-derived mesenchymal stem cells (hUC-MSCs) have been used extensively in the treatment of various inflammatory diseases, due to their potent regulatory roles that can mitigate inflammation and repair damaged tissues. However, their use in NMO is currently limited, and the mechanism underlying the beneficial effects of hUC-MSCs on motor function in NMO remains unclear. In this study, we investigate the effects of hUC-MSCs on the recovery of motor function in an NMO systemic model. Our findings demonstrate that milk fat globule epidermal growth 8 (MFGE8), a key functional factor secreted by hUC-MSCs, plays a critical role in ameliorating motor impairments. We also elucidate that the MFGE8/Integrin αvβ3/NF-κB signaling pathway is partially responsible for structural and functional recovery, in addition to motor functional enhancements induced by hUC-MSC exposure. Taken together, these findings strongly support the involvement of MFGE8 in mediating hUC-MSCs-induced improvements in motor functional recovery in an NMO mouse model. In addition, this provides new insight on the therapeutic potential of hUC-MSCs and the mechanisms underlying their beneficial effects in NMO.

overall_design: Scanning gene expression profiling analysis of cytokines for hUC-MSC cells.

first: Huiming

last: Xu

first: Wei

last: Jiang

first: Xuejia

last: Li

first: Jiaohua

first: Haijia

last: Qiu

first: Changyong

last: Tang


title: Single cell RNA-sequencing of lungs of Ifngr1-/-Rag-2-/- mice

first: Kazuyo

last: Moro

summary: We discovered Ifngr1-/-Rag-2-/- mice spontaneously develop severe pulmonary fibrosis (PF), and divided the progression process of PF in this strain into 3 phases; the pre-disease phase (until 12 weeks old; intact), the onset phase (from 12 to 18 weeks old; characterized by inflammations), and the chronic phase (after 18 weeks old; characterized by collagen deposition and obvious clinical symptoms). For the purpose of understanding the transcriptome dynamics of each phase, we performed single-cell RNA-Seq analysis on the whole lung cells from Rag-2-/- mice and Ifngr1-/-Rag-2-/- mice of different ages, including the pre-disease (7 weeks old), the onset (12 and 16 weeks old), and chronic phases (19 and 24weeks old).

overall_design: The whole lungs of Ifngr1-/-Rag-2-/- mice (7, 12, 16, 19, and 24 weeks old), and Rag-2-/- mice (18 weeks old; control) were isolated and dissociated, and then single-cell libraries of the whole lung cells were generated using the Chromium Single Cell 3’ v2 Reagents kit (10x Genomics).

first: Natsuko


title: Stage-specific dynamic reorganization of genome topology shapes transcriptional neighborhoods in developing human retinal organoids

city: Bethesda

first: Zachary

last: Batz

state: MD

address: 6 Center Dr Bldg 6, Rm 303

department: NEI

institute: NIH

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

We have generated high-resolution Hi-C map of developing human retinal organoids to elucidate spatiotemporal dynamics of genomic architecture and its relationship with gene expression patterns. We demonstrate progressive and developmental stage-specific alterations in DNA topology and correlate these changes with transcription of cell type-restricted gene markers during retinal differentiation. Temporal Hi-C reveals a shift towards A compartment for protein-coding genes and B compartment for non-coding RNAs, displaying high and low expression respectively. Notably, retina-enriched genes are clustered near lost boundaries of topologically associated domains (TADs), and higher-order assemblages of TADs (i.e., TAD cliques) tend to localize in the active chromatin regions with binding sites for eye-field transcription factors. These genes show a gain of chromatin contacts at their transcription start site as organoid differentiation proceeds. Our study provides a global view of chromatin architecture dynamics associated with diversification of cell types during retinal development and serves as a foundational resource for in-depth functional investigations of retinal developmental traits.


title: Multimodal profiling reveals site-specific adaptation and tissue residency hallmarks of γδ T cells across organs in mice

city: Freiburg

first: Sagar

address: Hugstetter Straße 55

department: Department of Internal Medicine II

institute: University Medical Center Freiburg

summary: γδ T cells perform heterogeneous functions in homeostasis and disease across tissues. However, it is unclear whether these roles correspond to distinct γδ subsets or to a homogeneous population of cells exerting context-dependent functions. Here, by cross-organ multimodal single-cell profiling, we reveal that various mouse tissues harbor unique site-adapted γδ subsets. Epidermal and intestinal intraepithelial γδ T cells are transcriptionally homogeneous and exhibit epigenetic hallmarks of functional diversity. Through parabiosis experiments, we uncovered cellular states associated with cytotoxicity, innate-like rapid IFN-γ production and tissue repair functions displaying tissue residency hallmarks. Notably, our observations nuance the link between IL-17-producing γδ T cells and tissue residency. Moreover, transcriptional programs associated with tissue-resident γδ T cells are analogous to those of CD8+ tissue-resident memory T cells. Altogether, this study provides the first multimodal landscape of tissue-adapted γδ T cells, revealing heterogeneity, lineage relationships and their tissue residency program.

overall_design: To identify and compare various subsets of γδ T cells present across different tissues, we took a single-cell multimodal approach and sorted gd T cells from lymph nodes, spleen, liver, lung, skin, small and large intestine using flow cytometry. Sorted cells were subjected to single-cell sequencing using two different approaches. In the first approach, γδ T cells from different organs were barcoded with distinct oligonucleotide-conjugated antibodies as well as 26 other antibodies. The transcriptomes and epitopes were simultaneously profiled using single-cell RNA sequencing (scRNA-seq) and TotalSeq, respectively. In the second approach, simultaneous single-nucleus RNA sequencing (snRNA-seq) and assay for transposase-accessible chromatin using sequencing (scATAC-seq) were performed on pooled γδ T cells from different organs without sample barcoding. Furthermore, simultaneous profiling of RNA and gamma delta T cell receptor (TCR) profiling was also performed at a single-cell resolution.

last: Sagar


title: A modular dCas9-based recruitment platform for combinatorial epigenome editing

status: Public on Nov 02 2023

city: Perth

first: Ryan

last: Lister

state: WA

address: 35 Stirling Highway

institute: The University of Western Australia

type: Methylation profiling by high throughput sequencing

summary: Targeted epigenome editing tools allow precise manipulation and investigation of genome modifications, however they often display high context dependency and variable efficacy between target genes and cell types. While systems that simultaneously recruit multiple distinct ‘effector’ chromatin regulators can improve efficacy, they generally lack control over effector composition and spatial organisation. To overcome this we have created a modular combinatorial epigenome editing platform, called SSSavi. This system is an interchangeable and reconfigurable docking platform fused to dCas9 that enables simultaneous recruitment of up to four different effectors, allowing precise control of effector composition and spatial ordering. We demonstrate the activity and specificity of the SSSavi system and compare it to existing multi-effector targeting systems, demonstrating its efficacy. Furthermore, we demonstrate the importance of the effector recruitment spatial ordering for effective transcriptional regulation. Together, the SSSavi system enables exploration of combinatorial effector co-recruitment to enhance manipulation of chromatin contexts previously resistant to targeted editing.

overall_design:
Chromatin immunoprecipitation sequencing (ChIP-seq) for histone modifications H3K4me3, H3K9me3 and H3K27me3 in HEK293T cells stably expressing dCas9-SSSavi and 6x sgRNA, and transfected with either DNMT3A+KRAB+EZH2 (D3A-K-E) or aGCN4-mCherry (non-catalytic control) for 72h, followed by flow cytometric enrichment of 200,000 cells positive for GFP (containing SSSavi components).
WGBS was performed on HSB6G cells transfected with either D3A-K-E or 𝛼GCN4-mCherry for 72h, followed by flow cytometric enrichment of 100,000 cells positive for GFP (containing SSSavi components). Genomic DNA was extracted and processed as detailed in the protocols section.

first: Tessa

last: Swain

first: Christian

last: Pflueger

first: Saskia

last: Freytag

first: Daniel

last: Poppe

first: Jahnvi

first: Trung

last: Nguyen

first: Ji


summary: We have generated high-resolution Hi-C map of developing human retinal organoids to elucidate spatiotemporal dynamics of genomic architecture and its relationship with gene expression patterns. We demonstrate progressive and developmental stage-specific alterations in DNA topology and correlate these changes with transcription of cell type-restricted gene markers during retinal differentiation. Temporal Hi-C reveals a shift towards A compartment for protein-coding genes and B compartment for non-coding RNAs, displaying high and low expression respectively. Notably, retina-enriched genes are clustered near lost boundaries of topologically associated domains (TADs), and higher-order assemblages of TADs (i.e., TAD cliques) tend to localize in the active chromatin regions with binding sites for eye-field transcription factors. These genes show a gain of chromatin contacts at their transcription start site as organoid differentiation proceeds. Our study provides a global view of chromatin architecture dynamics associated with diversification of cell types during retinal development and serves as a foundational resource for in-depth functional investigations of retinal developmental traits.

overall_design: RNA-seq at five time points in retinal organoid development (D60, D70, D90, D120, D200).

first: Zepeng

last: Qu

first: Nivedita

last: Singh

first: Claire

last: Marchal

first: Anand

last: Swaroop


title: Synthetically mannosylated antigens induce antigen-specific humoral tolerance and prevent anti-drug antibody responses to immunogenic biologics

first: Jeffrey

last: Hubbell

state: Illinois

address: 5640 S. Ellis Ave

institute: University of Chicago

summary: The purpose of this study were to test the feasibility of using pMan-antigen therapy to induce humoral tolerance. We used OVA as a model protein along with TCR transgenic OT-I and OT-II to study antigen-specific CD4+ T cell phenotypes and transcriptomic responses using RNA-seq readouts.

overall_design: Mice were adoptively transferred with 750,000 OT-I and OT-II cells at day 0. At day 1, mice received i.v. saline, 5 µg pMan-OVA, or equimolar OVA. Four days later, mice were sacrificed and spleens and liver and peripheral lymph nodes were pooled.Each replicate was generated from fluorescent activated cell sorted OTI and OTII cells pooled from 3 mice.

first: Rachel

last: Wallace

first: Jennifer

last: Antane

first: Kirsten

last: Refvik

first: Kym

last: Brünggel

first: Andrew

last: Tremain

first: Michal

last: Raczy

first: Aaron

last: Alpar

first: Mindy

first: Ani

last: Solanki

first: Elyse

last: Watkins

first: Anna

last: Slezak

first: Abigail

last: Lauterbach

first: D

last: Wilson


title: Epigenetic fingerprint of the SARS-CoV-2 infection in the lung of lethal COVID-19.

city: Badalona

first: Manel

last: Esteller

state: Barcelona

address: Ctra. de Can Ruti, Cami de les Escoles s/n

department: Cancer and Leukemia Epigenetics and Biology Program (PEBCL)

country: Spain

institute: Josep Carreras Leukaemia Research Institute (IJC)

type: Methylation profiling by genome tiling array

summary: Interrogation of the DNA methylome from lung autopsies of 36 patients who died of SARS-CoV-2 vs 18 control non-COVID-19 individuals.

overall_design: Bisulphite converted DNA from 54 samples (36 infected of SARS-CoV-2 vs 18 control non-COVID-19 individuals) were hybridized to the Illumina Infinium HumanMethylationEPIC version 1.0 microarray

first: A

last: Noguera-Castells

first: J

last: Parra

first: V

last: Davalos

first: C

last: García-Prieto

first: Y

last: Veselinova

first: T

last: Caniego-Casas

last: Palacios

first: X

last: Saenz-Sardà

first: E

last: Englund

last: Musulen

first: M


title: Neuronal TIMP2 regulates hippocampus-dependent plasticity and extracellular matrix complexity

city: New York

first: Joseph

last: Castellano

state: NY

address: 1425 Madison Avenue

department: Neuroscience

institute: Icahn School of Medicine at Mount Sinai

summary: The functional output of the hippocampus, a brain region subserving memory processes, depends on highly orchestrated cellular and molecular processes that regulate synaptic plasticity throughout life. The structural requirements of such plasticity and molecular processes involved in this regulation are poorly understood. Specific molecules, including tissue inhibitor of metalloproteinases-2 (TIMP2) have been implicated in serving a pro-plasticity role in the hippocampus, a role that decreases with brain aging. Here, we report that TIMP2 is highly expressed by neurons within the hippocampus and its loss drives changes in cellular programs related to adult neurogenesis and dendritic spine turnover with corresponding impairments in hippocampus-dependent memory. We find that TIMP2 regulates accumulation of extracellular matrix (ECM) around synapses in the hippocampus with concomitant hindrance in migration of newborn neurons through a denser ECM network. A conditional TIMP2 KO mouse reveals that neuronal TIMP2 regulates adult neurogenesis, accumulation of ECM, and ultimately hippocampus-dependent memory. Our results define a mechanism whereby hippocampus-dependent function is regulated by TIMP2 and its interactions with the ECM to regulate diverse processes associated with synaptic plasticity.

overall_design: Bulk RNAseq from hippocampus dissected from WT and TIMP2 KO mice.

first: Ana

last: Ferreira

first: Sarah

last: Philippi


title: Tumor and immune cell types interact to produce heterogeneous phenotypes of pediatric high grade glioma

city: Aurora

first: John

last: DeSisto

state: CO

department: Pediatrics

institute: University of Colorado

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

overall_design: Refer to individual Series


title: Active natural compounds perturb the melanoma risk-gene network

status: Public on Nov 01 2023

city: London

first: Claudia

last: Manzoni

address: 29-39 Brunswick Square

institute: UCL

summary: Cutaneous melanoma is an aggressive type of skin cancer with a complex genetic landscape caused by the malignant transformation of melanocytes. The study aimed at providing an in-silico network model based on the systematic profiling of the melanoma-associated genes considering germline mutations, somatic mutations, and genome-wide association studies (GWAS) signals (collectively melanoma risk genes). A protein-protein interaction network (melanoma risk network) was constructed using the melanoma risk genes to describe the functional landscape in which the melanoma genes operate within the cellular milieu. A significant portion of the melanoma risk network showed differential expression when SK-MEL-28 human melanoma cells were exposed to the phytochemicals harmine and berberine chloride reinforcing the hypothesis that network modelling may represent an alternative screening approach to prioritize potentially active compounds.

overall_design:
SK-MEL-28 human melanoma cells were treated with harmine and berberine chloride and differential gene expression following RNAseq was calculated in comparison with DMSO treated controls. All treatments were analysed using experimental triplicates.
The genes with significant alteration in expression following drug treatment, were mapped onto an in-silico model of melanoma obtained using protein network analysis of melanoma risk genes. This led to the identification of the portions of the model affected/modulated by the treatment.

first: Luying

last: Shao

first: Yibo

last: Zhao

first: Michael

last: Heinrich

first: Jose

last: Prieto


title: Transcriptional changes during Isoproterenol induced cardiac fibrosis in mice

city: Hyderabad

first: Kumarswamy

last: Regalla

state: Telangana

address: Centre for Cellular and Molecular Biology (CSIR-CCMB), Habsiguda

country: India

institute: CCMB

summary: Introduction: β-adrenergic stimulation using β-agonists such as isoproterenol has been routinely used to induce cardiac fibrosis in experimental in animal models. While transcriptome changes in surgical models of cardiac fibrosis such as Transverse aortic constriction (TAC), and coronary artery ligation (CAL) are well-studied, transcriptional changes during isoproterenol induced cardiac fibrosis is not well explored. Methods: Cardiac fibrosis was induced in male C57BL6 mice by administration of isoproterenol for 4, 8 or 11 days at 50mg/kg/day dose. Temporal changes in gene expression were studied by RNA sequencing.  Results:  We observed a significant alteration in the transcriptome profile across the different experimental groups compared to the saline group. Isoproterenol treatment caused upregulation of genes associated with ECM organization, cell-cell contact, three-dimensional structure, and cell growth, while genes associated with fatty acid oxidation, sarcoplasmic reticulum calcium ion transport, and cardiac muscle contraction are downregulated. A number of known long non-coding RNAs (lncRNAs) and putative novel lncRNAs exhibited differential regulation.  Conclusion: In conclusion, our study shows that isoproterenol administration leads to the deregulation of genes relevant to ECM deposition and cardiac contraction and serves as an excellent alternate model to the surgical models of heart failure.

overall_design:
To investigate the cardiac transcriptional changes upon isoproterenol administration in C57BL6 male mice( 12 weeks age).
We took 12 weeks old male C57BL6 mice which were subjected to subcutaneous administration of Isoproterenol at 50mg/kg/day for a period of 4,8 and 11 days.
Hearts were collected at respective time points for RNA isolation and RNA sequencing. Comparative gene expression profiling was done with respect to saline.

first: Disha

last: Nanda

first: Priyanka

last: Pant

first: Pratheusa

last: Machha

first: Divya

last: Sowpati

first: Regalla

last: Kumarswamy


title: Analysis of Differentially Expressed Genes in Torn Rotator Cuff Tendon Tissues in Diabetic Patients through RNA-sequencing

city: Beijing

first: Ziyang

last: Yuan

address: Yongan Road No.95

institute: Beijing Friendship Hospital

summary:
Background Rotator cuff tears (RCT) is a common musculoskeletal disorder in the shoulder which cause pain and functional disability. Diabetes mellitus (DM) is characterized by impaired ability of producing or responding to insulin and has been reported to act as a risk factor of the progression of rotator cuff tendinopathy and tear. Long non-coding RNAs (lncRNAs) are involved in the development of various diseases, but little is known about their potential roles involved in RCT of diabetic patients.
Methods RNA-Sequencing (RNA-Seq) was used in this study to profile differentially expressed lncRNAs and mRNAs in RCT samples between 3 diabetic and 3 nondiabetic patients. Gene ontology (GO) and Kyoto encyclopedia of genes and genomes (KEGG) pathway analysis were performed to annotate the function of the differentially expressed genes (DEGs). LncRNA-mRNA co-expression network and competing endogenous RNA (ceRNA) network were constructed to elucidate the potential molecular mechanisms of DM affecting RCT.
Results In total, 505 lncRNAs and 388 mRNAs were detected to be differentially expressed in RCT samples between diabetic and nondiabetic patients. GO functional analysis indicated that related lncRNAs and mRNAs were involved in metabolic process, immune system process and others. KEGG pathway analysis indicated that related mRNAs were involved in ferroptosis, PI3K-Akt signaling pathway, Wnt signaling pathway, JAK-STAT signaling pathway and IL-17 signaling pathway and others. LncRNA-mRNA co-expression network was constructed, and ceRNA network showed the interaction of differentially expressed RNAs, comprising 5 lncRNAs, 2 mRNAs, and 142 miRNAs. TF regulation analysis revealed that STAT affected the progression of RCT by regulating the apoptosis pathway in diabetic patients.
Conclusions We preliminarily dissected the differential expression profile of lncRNAs and mRNAs in torn rotator cuff tendon between diabetic and nondiabetic patients. And the bioinformatic analysis suggested some important RNAs and signaling pathways regarding inflammation and apoptosis were involved in diabetic RCT. Our findings offer a new perspective on the association between DM and progression of RCT.

overall_design: To elucidate the  potential roles of diabetes affecting rotator cuff tears, we harvested the torn rotator cuff tendon between diabetic and nondiabetic patients  and analyzed using RNA-Seq.


title: Synergistic effects of intermittent ethanol exposure and increased GIRK2 expression in NGN2 neurons

first: Paul

last: Slesinger

address: 1425 Madison Ave

summary: Increased expression of G-coupled Inwardly Rectifying Potassium Channel Subunit 2 (GIRK2) has been linked to an electrophysiological endophenotype for alcohol use disorder. We have increased GIRK2 expression in human glutamatergic (NGN2-induced) neurons using two distinct strategies (CRISPRa targeting of the endogenous gene and ectopic expression with a lentiviral vector) to investigate the interplay between GIRK2 expression levels and neuronal response to alcohol (ethanol). We used six healthy donor cell lines (2 CRISPRa and 4 lenti) to draw  comparisons within the same genetic background.  Findings from functional data (calcium imaging, electrophysiology, and mitochondrial stress tests) indicate that increased GIRK2 expression, combined with prolonged (21-day) intermittent ethanol exposure (IEE), counteracts IEE-induced changes in glutamatergic activity and cellular respiration. To understand the transcriptomic underpinnings of these functional data, we conducted bulk RNAseq in the same donor cohort. Differential gene expression analysis indicates that both CRISPRa and lenti-increased GIRK2 interacted with IEE to produce unique differential regulation of synaptic genes, including glutamate receptors and voltage-gated ion channels, distinct from the effects of IEE or increased GIRK2 alone.  Increased expression of G-coupled Inwardly Rectifying Potassium Channel Subunit 2 (GIRK2) has been linked to an electrophysiological endophenotype for alcohol use disorder. We have increased GIRK2 expression in human glutamatergic (NGN2-induced) neurons using two distinct strategies (CRISPRa targeting of the endogenous gene and ectopic expression with a lentiviral vector) to investigate the interplay between GIRK2 expression levels and neuronal response to alcohol (ethanol). We used six healthy donor cell lines (2 CRISPRa and 4 lenti) to draw  comparisons within the same genetic background.  Findings from functional data (calcium imaging, electrophysiology, and mitochondrial stress tests) indicate that increased GIRK2 expression, combined with prolonged (21-day) intermittent ethanol exposure (IEE), counteracts IEE-induced changes in glutamatergic activity and cellular respiration. To understand the transcriptomic underpinnings of these functional data, we conducted bulk RNAseq in the same donor cohort. Differential gene expression analysis indicates that both CRISPRa and lenti-increased GIRK2 interacted with IEE to produce unique differential regulation of synaptic genes, including glutamate receptors and voltage-gated ion channels, distinct from the effects of IEE or increased GIRK2 alone.

overall_design: Two donor cohorts: CRISPRa (2 donors) and Lenti (4) donors were used to generate NGN2 neurons with endogenous and increased levels of GIRK2 expression. These neurons were then subject to 21 days of intermittent exposure to 17 mM ethanol. Two batches of neurons and three technical replicates/donor/batch were generated for RNA harvest and bulk RNAseq. The two cohorts were then analysed separately from one another, to separate donor effects from GIRK2 expression manipulation method. Additional analyses were performed on the pooled cohort, correcting for systematic noise and donor effects.

first: Iya

last: Prytkova

first: Yiyuan

last: Liu

first: Alison

last: Goate


title: Brn3bactsviaTac2 to regulatedefensive responses to visual threat

city: New Haven

first: In-Jung

last: Kim

state: CT

address: 300 George Street

department: Ophthalmology

institute: Yale University

summary: Defensive responses to visually threatening stimuli represent an essential fear-related survival instinct, widely detected across species. The neural circuitry mediating visually triggered defensive responses has been delineated in the midbrain. However, the molecular mechanisms regulating the development and function of these circuits remain unresolved. Here we show that midbrain-specific deletion of the transcription factor Brn3b causes a loss of neurons projecting to the lateral posterior nucleus of the thalamus. Brn3b deletion also downregulates the expression of the neuropeptide tachykinin 2 (Tac2). Furthermore, Brn3b mutant mice display impaired defensive freezing responses to visual threat precipitated by social isolation. This behavioral phenotype could be ameliorated by overexpressing Tac2, suggesting that Tac2 acts downstream of Brn3b in regulating defensive responses to threat. Together, our experiments identify specific genetic components critical for the functional organization of midbrain fear-related visual circuits. Similar mechanisms may contribute to the development and function of additional long-range brain circuits underlying fear-associated behavior.

overall_design: To investigate the role of Brn3b on the development of mouse midbrain, we utilized the conditional knockout strategy to delete Brn3b in the midbrain only. We then conducted RNA sequencing and compared gene expression between heterozygotes and homozygotes of Brn3b at neonatal stage (P2-P4).

first: Hyoseo

last: Lee

first: Hannah

last: Weinberg-Wolf

first: Hae-Lim

first: Taeji

first: Joseph

last: Conte

first: Carlos

last: Godoy-Parejo

first: Jonathan

last: Demb

first: Andrii

last: Rudenko


title: PIWI-interacting RNA-YBX1 inhibits proliferation and metastasis by the MAPK signaling pathway via YBX1 in triple-negative breast cancer

city: Guangzhou

first: Huang

middle: Shan

last: Shan

address: 651 Dongfeng Road East

institute: Sun Yat-sen University Cancer Center

summary: Breast cancer is the second leading cause of death in women worldwide, with triple-negative breast cancer (TNBC) having the worst prognosis. Although there are numerous studies on TNBC, there is no effective treatment for it, and it is still a major problem today. Studies on PIWI-interacting RNAs (piRNAs) are increasing and investigating the mechanism of piRNAs in the proliferation and metastasis of TNBC may lead to new potential treatment targets. Here, we identified a novel piRNA, piR-YBX1, which was downregulated in TNBC compared to matched normal breast tissue. Overexpression of piR-YBX1 significantly inhibited the proliferation, migration, invasion ability of TNBC cells both in vivo and in vitro. Mechanistically, piR-YBX1 could bind directly to mRNA of YBX1 and overexpression of piR-YBX1 downregulated YBX1 in both mRNA and protein levels, while the function of piR-YBX1 could be partly rescued by overexpression of YBX1. In addition, YBX1 could bind to RAF1 which is the key molecule in the MAPK signaling pathway, and overexpression of piR-YBX1 inhibited the p-MEK and p-ERK1/2 which can be reverted by YBX1. In conclusion, our findings discovered that the piR-YBX1/YBX1/MAPK axis suppresses the proliferation and metastasis of TNBC and therefore piR-YBX1 has the potential to be an effective therapeutic agent for breast cancer.

overall_design: 3 pairs of TNBC tissue and matched adjacent normal breast tissue were collected to obtain profiles of differentially expressed piRNAs using piRNA microarray

first: Linyu

last: Wu

first: Shanshan

last: Huang

first: Wenwen

last: Tian

first: Peng

first: Yi

last: Xie

first: Yu

last: Qiu

first: Xing

last: Li

first: Yuhui

last: Tang

first: Shaoquan

last: Zheng

first: Yuying

last: Sun

first: HaiLin

first: Wei

last: Du

first: Weige

last: Tan

first: Xinhua


title: Disrupted RNA editing in beta cells mimics early-stage type 1 diabetes

city: Jerusalem

first: agnes

last: klochendler

address: Hadassah Ein Kerem

country: Israel

institute: Hebrew University of Jerusalem

summary: A major hypothesis for the etiology of type 1 diabetes (T1D) postulates initiation by viral infection, leading to double-stranded RNA (dsRNA)-mediated interferon response and inflammation; however, a causal virus has not been identified. Here we use a mouse model, corroborated with human islet data, to demonstrate that endogenous dsRNA in beta cells can lead to a diabetogenic immune response, thus identifying a virus-independent mechanism for T1D initiation. We found that disruption of the RNA editing enzyme ADAR in beta cells triggers a massive interferon response, islet inflammation and beta cell failure and destruction, with features bearing striking similarity to early-stage human T1D. Glycolysis via calcium enhances the interferon response, suggesting an actionable vicious cycle of inflammation and increased beta cell workload.

overall_design:
Inactivation of Adar in pancreatic beta-cells from young adult mice (MipCreER; Adar floxed/floxed) causes insulitis and diabetes. To understand the molecular basis for islet inflammation, we have performed RNA-Sequencing on FACS-sorted beta-cells isolated from beta-cell specific Adar -Knockout (betaAdarKO) and control beta-cell specific Adar-heterozygotes (betaAdarHET) (n=4 for each genotype).
We then profiled and compared gene expression from RNA-Seq data of AdarKO and AdarHET beta-cells

first: Udi

last: Knebel

first: Shani

last: Peleg

first: Roni

last: Cohen-Fultheim

first: Erez

middle: Y

last: Levanon

first: Agnes

last: Klochendler

first: Yuval

last: Dor


title: Design, Construction, and Functional Characterization of a tRNA Neochromosome in Yeast

first: Tao

last: Pan

address: 929 E. 57th Street

department: Biochemistry and Molecular Biology

type: Non-coding RNA profiling by high throughput sequencing

summary: Here we report the design, construction and characterization of a tRNA neochromosome, a designer chromosome that functions as an additional, de novo counterpart to the native complement of Saccharomyces cerevisiae. Intending to address one of the central design principles of the Sc2.0 project, the ~190 kb tRNA neochromosome houses all 275 relocated nuclear tRNA genes. To maximize stability, the design incorporated orthogonal genetic elements from non-S. cerevisiae yeast species. Furthermore, the presence of 283 rox recombination sites enable an orthogonal SCRaMbLE system capable of adjusting tRNA abundance. Following construction, we obtained evidence of a potent selective force once the neochromosome was introduced into yeast cells, manifesting as a spontaneous doubling in cell ploidy. Furthermore, tRNA sequencing, transcriptomics, proteomics, nucleosome mapping, replication profiling, FISH and Hi-C were undertaken to investigate questions of tRNA neochromosome behavior and function. Its construction demonstrates the remarkable tractability of the yeast model and opens up new opportunities to directly test hypotheses surrounding these essential non-coding RNAs

overall_design: To investigate tRNA expression from native chromosomes and neochrosome in Saccharomyces cerevisiae (DSy063, DSy162, DSy177, DSy199), we carried out multiplex small RNA sequencing (MSR-seq) of total RNA isolated from these yeast strains.  We then performed sequencing data analysis for mature tRNA and for precursor tRNAs containing the upstream (up) or downstream (down) sequences of these 4 strains of 3 biological replicates each.  We compared the expression levels between the mature tRNAs of these 4 strains, and the expression levels of the precursor tRNAs derived from the native chromosomes versus those derived from the neochromosome.

first: Daniel

last: Schindler

first: Roy

last: Walker

first: Shuangying

last: Jiang

first: Aaron

middle: N

last: Brooks

first: Yun

last: Wang

first: Carolin

last: Müller

first: Charlotte

last: Cockram

first: Yisha

last: Luo

first: Alicia

last: García

last: Schraivogel

first: Julien

last: Mozziconacci

first: Benjamin

last: Blount

first: Jitong

last: Cai

first: Lois

last: Ogunlana

last: Liu

first: Katarina

last: Jönsson

first: Dariusz

last: Abramczyk

first: Eva

last: Garcia-Ruiz

first: Tomasz

last: Turowski

first: Reem

last: Swidah

first: Tom

last: Ellis

first: Francisco

last: Antequera

first: Yue

last: Shen

first: Conrad

last: Nieduszynski

first: Romain

last: Koszul

first: Junbiao

last: Dai

first: Lars

last: Steinmetz

first: Jef

last: Boeke

first: Yizhi

first: Mahdi

last: Assari

first: Noah

last: Pena


title: BRD9 determines the cell fate of hematopoietic stem cells by regulating chromatin state [Mouse_Brd9_CTCF ChIP-seq]

city: Kobe

first: Weijia

last: Zang

address: Hyogo, Kobe, Chuo Ward, Minatojima Minamimachi, 6 Chome−３−7

department: Department of Hematology-Oncology

institute: Foundation for Biomedical Research and Innovation at Kobe

summary: Switch/sucrose nonfermentable (SWI/SNF) complexes are ATP-dependent chromatin remodeler complexes that play critical roles in timely and appropriate gene regulation by modulating chromatin architecture and DNA accessibility. SWI/SNF complexes can be grouped into three subcomplexes of differing sizes, canonical BAF (cBAF), polybromo BAF (PBAF), and newly identified noncanonical BAF (ncBAF). The most recently characterized ncBAF lacks the core BAF subunits ARID, BAF47, and BAF57, but includes unique subunits GLTSCR1/1L and BRD9, one of the bromodomain-containing proteins.We recently demonstrated the novel mechanism for the ncBAF disruption caused by mutations in a spiceosomal protein, SF3B1, especially in 65–83% for myelodysplastic syndromes (MDS) with ring sideroblasts as well as in >20% of mucosal/uveal melanomas, suggesting that the disturbed ncBAF may have some roles in the malignant hematopoiesis.Mechanistically, SF3B1 mutant recognizes an aberrant, deep intronic branchpoint within BRD9 and thereby induces the inclusion of an aberrant exon and subsequently profound degradation of BRD9 mRNA by triggering nonsense-mediated RNA decay (NMD). However, compared to the roles of cBAF, the functions of BRD9/ncBAF in normal and malignant hematopoiesis in vivo have been totally uncharacterized.

overall_design: 2.0 x 10E6 total BM cells of Mx1-Cre control, Mx1-Cre Brd9fl/fl mice transplanted via tail vein injection into lethally irradiated (two times 450 cGy) CD45.1+ recipient mice. Four weeks after BMT, the recipient mice received 20 mg/kg pIpC injection every other day for a total of 3 doses. Five months after pIpC injection, the recipient mice were sacrificed for sample preparation.

first: Daichi

last: Inoue

first: Koutarou

last: Nishimura

first: Muran

last: Xiao


title: Orphan quality control by an SCF ubiquitin ligase directed to pervasive C-degrons

city: Mainz

first: Frank

last: Rühle

address: Ackermannweg 4

department: Bioinformatics Core Facility

institute: Institute of Molecular Biology (IMB)

summary: Selective protein degradation typically involves substrate recognition via short linear motifs known as degrons. Various degrons can be found at protein termini from bacteria to mammals. While N-degrons have been extensively studied, our understanding of C-degrons is still limited. Towards a comprehensive understanding of eukaryotic C-degron pathways, we performed an unbiased survey of C-degrons in budding yeast. We identified over 5000 potential C-degrons by stability profiling of random peptide libraries and of the yeast C-terminome. Combining machine learning, high-throughput mutagenesis and genetic screens revealed that the SCF ubiquitin ligase targets ~40% of degrons using a single F-box substrate receptor Das1. Although sequence-specific, Das1 is highly promiscuous, recognizing a variety of C-degron motifs. By screening for full-length substrates, we implicate SCFDas1 in degradation of orphan protein complex subunits. Altogether, this work highlights the variety of C-degron pathways in eukaryotes and uncovers how an SCF/C-degron pathway of broad specificity contributes to proteostasis.

overall_design: Multiplexed protein stability (MPS) profiling using DNA amplicon-seq data. For this, a library of peptide variants (based on inserted random or pre-determined DNA sequences) fused to a tandem fluorescent protein timer (tFT) is expressed from yeast plasmids and sorted into stability bins using the tFT readout. The contained variable regions are sequenced and the occurrences of all observed DNA sequence variants are counted per stability bin. The distributions of read counts are summarized in the form of a protein stability index (PSI) for each peptide.

first: Ka-Yiu

last: Kong

first: Susmitha

last: Shankar

first: Anton

last: Khmelinskii


title: Integration of miRNA in exosomes and single-cell RNA-seq profiles in endemic osteoarthritis, Kashin-Beck disease

city: Xi'an

first: Xi

last: Wang

address: No.76 Yanta west road

institute: Xi'an Jiaotong University

summary: Kashin-Beck disease (KBD) is an endemic, chronic degenerative joint disease in China. Exosomes miRNAs, as signaling molecules in intercellular communication, can transfer specific biological martials into target cell to regulate the their function and might participate in the pathogenesis of KBD. We isolated serum and chondrocytes-derived exosomes, miRNA sequencing revealed exosomes miRNA profiles and differentially expressed miRNAs (DE-miRNAs) were identified. The target genes were predicted of known and novel DE-miRNAs with TargetScan 5.0 and miRanda 3.3a database. Single-cell RNA sequencing (scRNA-seq) was performed to identify chondrocyte clusters and their gene signatures in KBD. And we performed comparative analysis between the serum and chondrocytes-derived exosomes DE-miRNA target genes and differentially expressed genes of each cell clusters.A total of 20 DE-miRNAs were identified in serum-derived exosomes. In the miRNA expression of chondrocytes-derived exosomes, 53 DE-miRNAs were identified. 16063 predicted targets were identified as the target genes in the serum-derived exosomes, 57316 predicted targets were identified as the target genes in the chondrocytes-derived exosomes. Seven clusters were labelled by cell type according to the expression of previously described markers. 315 common genes were found among serum/chondrocytes-derived exosomes DE-miRNA target genes and DEGs identified by scRNA-seq analysis. We firstly integratly analyzed the serum and chondrocytes exosomes miRNA with single-cell RNA sequencing (scRNA-seq) data of KBD chondrocyte, the results showed that DE-miRNAs in exosomes might play a potential role in regulating genes expression in different KBD chondrocytes clusters by exosomes mediating cell-cell communications functions, which could improve the new diagnosis and treatment methods for KBD.

overall_design: observation study

first: Yu

last: Zhang

first: Yifan

last: Wu

first: Yujie

last: Ning


title: BRD9 determines the cell fate of hematopoietic stem cells by regulating chromatin state [MOLM13 DMSO vs dBRD9 RNA-seq]

overall_design: The MOLM13 cell lines were cultured in RPMI with 10% FCS and 1% P/S to the logarithmic growth phase,  treated MOLM13 cells with DMSO and 50nM dBRD9 for 4 days and abstract RNA for RNA-seq.


title: The Triptryium wilfordii derivative celastrol has anti-fibrotic effects in systemic sclerosis [In vivo bulk RNA-seq]

city: Saskatoon

first: Andrew

last: Leask

state: Saskatchewan

address: 105 Wiggins

department: College of Dentistry

institute: University of Saskatchewan

summary: Objectives: Scleroderma (systemic sclerosis, SSc), as a prototypic inflammation-driven fibrotic disease, possesses connective tissue lesions populated with persistently activated myofibroblasts maintained by an mechanotranductive/pro-adhesive signaling loop. Drugs targeting this pathway are therefore of likely therapeutic benefit. The mechanosensitive transcriptional co-activator, yes activated protein-1 (YAP1), is activated in SSc fibroblasts. The terpenoid celastrol has recently been identified as a YAP1 inhibitor: however, if celastrol can alleviate SSc fibrosis is unknown. Methods: Human dermal fibroblasts from healthy individuals and patients with diffuse cutaneous SSc were treated with or without transforming growth factor b1 (TGFb1) in the presence or absence of celastrol. C57BL6J mice were subjected to the inflammatory-driven bleomycin-induced model of skin SSc, in the presence or absence of celastrol. RNA expression was assessed using RNAseq, real-time polymerase chain reaction and spatial transcriptomic analyses. Protein expression was determined using Western blot and enzyme-linked immunosorbent assay. Fibrosis was monitored by hematoxylin and eosin and trichrome staining, and indirect immunofluorescence analysis. Results: In dermal fibroblasts, celastrol impaired the ability of TGFβ1 to induce an SSc-like pattern of gene expression, including the induction of cellular communication network factor 2 (CCN2), collagen I and TGFβ1 protein. Celastrol alleviated the persistent fibrotic phenotype of dermal fibroblasts cultured from lesions of SSc patients. In the bleomycin-induced model of SSc dermatopathology, celastrol inhibited fibrosis and blocked nuclear localization of YAP in myofibroblasts. Conclusion: Our data are consistent with the hypothesis that compounds, such as celastrol, that antagonize the YAP pathway may be potential treatments for SSc skin fibrosis.

overall_design: To investigate the gene expression of the PBS, Bleomycin and Celastrol treated mouse skin tissue samples

first: Pratyusha

last: Chitturi

first: Xu

last: Shi-wen

first: Bahja

last: Ahmed Abdi

first: John

last: Nguyen

first: David

last: Carter

first: Richard

middle: J

last: Stratton


title: The Triptryium wilfordii derivative celastrol has anti-fibrotic effects in systemic sclerosis [in vitro bulk RNA-seq]

overall_design: To investigate the effect of Celastrol on TGFB1 mediated gene expression in human skin fibroblasts


title: The Triptryium wilfordii derivative celastrol has anti-fibrotic effects in systemic sclerosis