title: Poly (ADP-Ribose) Polymerase 1 Regulates Cajal-Retzius Cell Development and Neural Precursor Cell Adhesion

city: Ann Arbor

first: GABRIEL

last: CORFAS

state: MI

address: U of M Kresge Hearing Research Institute 1150 West Medical Center Drive Medical Sciences I Building, Rm. 5428

institute: The University of Michigan, Kresge Hearing Research Institute

summary: Poly (ADP-Ribose) Polymerase 1 (PARP1) is a ubiquitously expressed enzyme that regulates DNA damage repair, cell death, inflammation, and transcription. PARP1 functions by adding ADP-ribose polymers (PAR) to proteins including itself, using NAD+ as a donor. This post-translational modification known as PARylation results in changes in the activity of PARP1 and its substrate proteins and has been linked to the pathogenesis of various neurological diseases. PARP1 KO mice display schizophrenia-like behaviors, have impaired memory formation, and have defects in neuronal proliferation and survival, while mutations in genes that affect PARylation have been associated with intellectual disability, psychosis, neurodegeneration, and stroke in humans. Yet, the roles of PARP1 in brain development have not been extensively studied. We now find that loss of PARP1 leads to defects in brain development and increased neuronal density at birth. We further demonstrate that PARP1 loss increases the expression levels of genes associated with neuronal migration and adhesion in the E15.5 cerebral cortex, including Reln. This correlates with an increased number of Cajal-Retzius (CR) cells in vivo and in cultures of embryonic neural progenitor cells (NPCs) derived from the PARP1 KO cortex. Furthermore, PARP1 loss leads to increased NPC adhesion to N-cadherin, like that induced by experimental exposure to Reelin. Taken together, these results uncover a novel role for PARP1 in brain development, i.e., regulation of CR cells, neuronal density, and cell adhesion.

overall_design: mRNA profiles of wild-type (WT) and PARP1 KO E15.5 cortex (n = 4 of each genotype)

first: Megan

last: Nelson

first: J

last: Hoff

first: Mya

last: Zeese

first: Gabriel

last: Corfas


title: Chromatin-accessibility estimation from single-cell ATAC data with scOpen

status: Public on Sep 17 2021

city: Aachen

first: Zhijian

last: Li

state: North Rhine-Westphal

address: Pauwelsstr. 19

country: Germany

institute: RWTH Aachen University

type: Genome binding/occupancy profiling by high throughput sequencing

type: Expression profiling by high throughput sequencing

summary: We propose a computational method for quantifying the open chromatin status of regulatory regions from single cell ATAC-seq experiments. scOpen, which is based on positive-unlabeled learning of matrices, is able to estimate the probability that a region is open at a given cell mitigating the sparsity of scATAC-seq matrices. We demonstrate that scOpen improves all down-stream analysis steps of scATAC-seq data as clustering, visualization, detection of transcription factors and chromatin conformation in several scATAC-seq data.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design:
We used scATAC product from 10x Genomics to profile UUO mouse kidney at Day 0, 2 and 10.
We generated Runx1 overexpressing RNA-seq data from human kidney fibroblasts.

first: Christoph

last: Kuppe

first: Mingbo

last: Cheng

first: Sylvia

last: Menzel

first: Martin

last: Zenke

first: Rafael

last: Kramann

first: Ivan

middle: G

last: Costa


title: Concurrently expressed tumor neoantigens drive phenotypically distinct CD8 T cell responses

status: Public on Sep 16 2021

city: Cambridge

first: Amanda

middle: Margarita

last: Cruz

state: MA

address: 500 Main Street, 76-431

department: Biology

country: USA

institute: Massachusetts Institute of Technology

summary: CD8 T cell responses against different tumor neoantigens occur simultaneously, yet little is known about the interplay of these responses or its impact on tumor control. Here, we characterized the CD8 T cell response against two co-expressed tumor neoantigens, SIINFEKL (SIIN) and SIYRYYGL (SIY), in a genetically engineered mouse model of lung adenocarcinoma. In this model, tumors are initiated in KrasLSLG12D;p53fl/fl (KP) mice by intratracheal installation of lentiviruses expressing Cre recombinase and the antigens as a fusion to luciferase (LucOS). In these KP LucOS mice, the CD8 T cell response peaks around 5 weeks post-tumor initiation and is dominated by expansion of cells specific to SIIN. SIY drives a much smaller T cell response, but it persists better over time. To further investigate phenotypic differences in the T cell responses against SIIN versus SIY, we subjected antigen-specific CD8 T cells from 5 week mice to paired single cell RNA and TCR sequencing using the 10X Genomics platform.

overall_design: SIIN- and SIY-specific CD8 T cells were sorted from the lungs of KP LucOS mice 5 weeks post-tumor initiation and subjected to paired RNA and TCR single cell RNA sequencing using the 10X Genomics platform.

middle: L

last: Burger

middle: M

first: Arjun

last: Bhutkar

first: Sarah

last: Blatt

first: Tyler

last: Jacks


title: Sequencing of RNA and footprints of wildtype and RpS5b mutant Drosophila ovaries [RIBO-seq]

city: Dallas

first: Michael

last: Buszczak

state: Texas

address: 5323 Harry Hines Boulevard

department: Molecular Biology

institute: University of Texas Southwestern Medical Center

summary: Emerging evidence suggests that ribosome heterogeneity may have important functional consequences in the translation of specific mRNAs within different cell types and under various conditions. Ribosome heterogeneity comes in many forms including post-translational modification of ribosome proteins (RPs), lack of specific RPs, and different RP paralogs incorporation. The Drosophila genome encodes for two RPS5 paralogs, RpS5a and RpS5b. While RpS5a is expressed ubiquitously, RpS5b exhibits enriched expression in the reproductive system. Deletion of RpS5b results in female sterility marked by developmental arrest of egg chambers at stages 7-8 and posterior follicle cell (PFC) hyperplasia. Both phenotypes are caused by defects in the germline and over-expression of RpS5a can compensate it. RpS5b mutant display increased rRNA transcription and RP production, accompanied by increased protein synthesis. Loss of RpS5b leads to microtubule-based defects and mislocalization of several factors, leading failure of Notch pathway activation in PFCs. Together, our results indicate that germ cell specific expression of RpS5b promotes proper egg chamber development by regulating both ribosome homeostasis and protein trafficking during mid-oogenesis.

overall_design: RNA-seq and footprint-seq experiment to compare gene expression as well as ribosome distribution between wildtype and the mutant ovaries.

first: Seoyeon

last: Jang


title: RNA-seq expression profiling of murine breast tumour cell lines and bone metastasis-associated endothelial cells

city: Parkville

first: Gordon

middle: K

last: Smyth

state: Vic

address: 1G Royal Pde

department: Bioinformatics

country: Australia

institute: Walter and Eliza Hall Institute of Medical Research

summary:
Breast tumor cells were found to remodel the bone marrow vascular microenviornment to support metastatic expansion. To identify tumor-derived factors that stimulate marrow endothelium, we studied the transcriptomes of four isogenic murine mammary tumor cell lines, 4T1.2, 4T1, 66cl4 and 67NR. To gain insight into the host vasculature in response to tumor colonization, we analysed the transcriptional signatures of EMCNhi/CD31hi endothelial cells extracted from heavily infiltrated bone marrow stroma (Metastatic) and from adjacent tumour-free bone marrow stroma (Non-metastatic), as well as from healthy bones (Control).

This SuperSeries is composed of the SubSeries listed below.

overall_design: Refer to individual Series

first: Yunshun

last: Chen

first: Raymond

last: Yip

first: Jane

middle: E

last: Visvader


title: RNA-seq expression profiling of metastasis-associated endothelial cells from murine bone marrow

summary:
Breast tumour cells were found to remodel the murine bone marrow vascular microenvironment to support metastatic expansion. To gain insight into the host vasculature in response to tumor colonization, cells from the isogenic murine mammary tumour cell line 4T1.2 were intracardially injected into immunocompetent BALB/c mice. We analysed the transcriptional profiles of endothelial cells extracted from heavily infiltrated (Metastatic) bone marrow stroma and the adjacent tumor-free bone marrow stroma (Non-metastatic), as well as from healthy bones (Control).
Methods: Immunocompetent BALB/c mice were intracardically injected with 4T1.2 tumor cells. The bones of these tumor-burdened mice were examined under fluorescence stereomicroscope to microdissect heavily-infiltrated (Metastatic) and the adjacent tumour-free (Non-metastatic) areas. Bones from healthy mice were used as controls. The dissected bone material from both burdened and control mice was processed to isolate EMCNhi/CD31hi endothelial cells using  flow cytometry. RNA was extracted for 75bp paired-end RNA-seq profiling using an Illumina NextSeq 500 sequencer.

overall_design: Two biological independent mouse pools were formed of healthy control mice and four pools of tumor-burdened mice. Each pool comprised between 1 and 4 mice in order to achieve sufficient material for RNA sequencing. Two control libraries, six non-metastatic libraries and three metastatic libraries were prepared for sequencing.


title: Mesenchymal stromal cell apoptosis is required for their therapeutic function

city: Clayton

first: David

middle: Richard

last: Powell

state: VIC

address: Wellington Road

department: Bioinformatics Platform

institute: Monash University

summary: The demonstration that mesenchymal stromal cells (MSCs) can ameliorate a wide range of diseases in preclinical models has led to many clinical trials investigating their use as a treatment modality. Their therapeutic effects are often attributed to their secretome, a proposition predicated on their long-term survival. However, the majority of MSCs do not survive in vivo administration. We sought to reconcile the immunosuppressive effects of MSC therapy with their short lifespan. We showed that almost all MSCs underwent apoptosis associated with caspase-3 activation within 1 hour of intravenous administration and were efferocytosed by lung phagocytes. The potent immunosuppressive effects of MSCs did not require them to remain viable, but instead involved efferocytosis-induced transcriptional changes in metabolic and inflammatory pathways in alveolar macrophages. We identified the mitochondrial pathway as the predominant mechanism involved in MSC apoptosis, as deletion of the effectors BAK/BAX prevented their death. Importantly, BAK/BAX deficiency in MSCs weakened their immunosuppressive effects in 2 different disease models used to define MSC potency. Our data showed that apoptosis of MSCs and their subsequent efferocytosis and macrophage immunometabolic reprogramming are key steps underlying their mode of action. These findings have significant implications for the development of MSC potency assays and biomanufacturing for clinical translation.

overall_design: The experiment was conducted on alveolar macrophages from OVA-sensitised mice that were treated with human mesenchymal stromal cells (MSCs). The macrophages were isolated at 2 different timepoints after treatment with MSCs. For each sample, macrophages were pooled from 5 mice. The experiment was performed twice to obtain 2 biological replicates for each condition. 4 conditions, 2 timepoints, 2 replicates. 16 samples in total.

first: Tracy

middle: S

last: Heng

first: Adele

last: Barugahare


title: RNA-seq expression profiling of murine breast tumor cell lines

summary: Breast tumour cells were found to remodel the bone marrow vascular microenviornment to support metastatic expansion. To identify tumour-derived factors that stimulate marrow endothelium, we studied the transcriptomes of four isogenic murine mammary tumour cell lines, 4T1.2, 4T1, 66cl4 and 67NR.

overall_design: Two biological repicates were prepared from each cell line (4T1, 4T1.2, 67NR, 66cl4).


title: Transcriptional profiling of distinct fibroblast populations in the pancreatic tumor microenvironment

city: Portland

first: Mara

last: Sherman

state: OR

address: 2730 SW Moody Avenue CL6C

institute: Oregon Health & Science University

summary: Pancreatic stellate cells are thought to be the predominant source of cancer-associated fibroblasts (CAFs) in pancreatic cancer. We developed a mouse model which allows us to track and analyze stellate cells and stellate cell-derived CAFs in vivo during pancreatic tumorigenesis for the first time. We find that stellate cells in fact give rise to a minority of all CAFs. Here, we have used lineage reporters to isolate stellate cell-derived and non-stellate cell-derived CAFs and compared them by RNA-seq.

overall_design: Rosa-mTmG; Fabp4-Cre mice at 8 weeks of age were injected with FC1199 murine pancreatic cancer cells orthotopically. After 4 weeks, tumors were harvested, single cell suspensions generated, and FACS performed to isolate GFP+ (stellate cell-derived) and Tomato+ (non-stellate cell-derived) CAFs. RNA was isolated and RNA-seq performed on both cell populations.

first: Wes

last: Horton

first: Erin

last: Helms


title: Integrative RNA-omics discovers GNAS alternative splicing as a phenotypic driver of splicing factor mutant neoplasms

city: La Jolla

first: Gene

last: Yeo

state: CA

address: 2880 Torrey Pines Scenic Dr. Room 3805/Yeo Lab

institute: UCSD

summary: Mutations in splicing factors (SFs) are the predominant class of mutations in myelodysplastic syndrome (MDS), but convergent downstream disease drivers remain elusive. To identify common direct targets of mis-splicing by mutant U2AF1 and SRSF2, we performed RNA-Seq and eCLIP in human hematopoietic stem/progenitor cells (HSPCs) derived from isogenic induced pluripotent stem cell (iPSC) models. Integrative analyses of alternative splicing and differential binding converged on a long isoform of GNAS (GNAS-L), promoted by both mutant factors. MDS population genetics, functional and biochemical analyses support that GNAS-L is a driver of MDS and encodes a hyperactive long form of the stimulatory G protein alpha subunit, Gas-L, that activates ERK/MAPK signaling. SF-mutant MDS cells have activated ERK signaling and consequently are sensitive to MEK inhibitors. Our findings highlight an unexpected and unifying mechanism by which SF mutations drive oncogenesis with potential therapeutic implications for MDS and other SF-mutant neoplasms.

overall_design: Isogenic iPSCs derived with CRISPR genome editing to insert heterozygous point mutations in SRSF2 and U2AF1 and insertion of a FLAG tag a the endogeneous were differentiated into heatopoietic stem and progenitor cells (HSPCs). eCLIP-Seq of each splicing factor and RNA-Seq were performed on the HSPCs.

first: Emily

middle: C

last: Wheeler

first: Shailee

last: Vora

first: Daniel

last: Mayer

first: Andriana

last: Kotini

first: Malgorzata

last: Olszewska

first: Samuel

last: Park

first: Ernesto

last: Guccione

first: Julie

last: Teruya-Feldstein

first: Lewis

last: Silverman

first: Roger

last: Sunahara

first: Eugene

middle: W

first: Eirini

middle: P

last: Papapetrou


title: Targeting miR-126 in inv(16) acute myeloid leukemia inhibits leukemia development and leukemia stem cell maintenance

status: Public on Sep 15 2021

city: Duarte

first: wei

last: chen

address: 1500 E Duarte Rd

department: Cancer Biology-BRI

institute: City of Hope National Medical Center

summary: Acute myeloid leukemia (AML) harboring inv(16)(p13q22) expresses high levels of miR-126. Here we show that the CBFB-MYH11 (CM) fusion gene upregulates miR-126 expression through aberrant miR-126 transcription and perturbed miR-126 biogenesis via the HDAC8/RAN-XPO5-RCC1 axis. Aberrant miR-126 upregulation promotes survival of leukemia-initiating progenitors and is critical for initiating and maintaining CM-driven AML. We show that miR-126 enhances MYC activity through the SPRED1/PLK2-ERK-MYC axis. Notably, genetic deletion of miR-126 significantly reduced AML rate and extended survival in CM knock-in mice. Therapeutic depletion of miR-126 with an anti-miR-126 (miRisten) inhibited AML cell survival, reduced leukemia burden and leukemia stem cell (LSC) activity in inv(16) AML murine and xenograft models. Combination of miRisten with chemotherapy further enhanced the anti-leukemia and anti-LSC activity. Overall, this study provides molecular insights for the mechanism and impact of miR-126 dysregulation in leukemogenesis and highlights the potential of miR-126 depletion as a new therapeutic approach for inv(16) AML.

overall_design: Total 10 LSK samples were analyzed in this study. 5 samples as CM, 5 samples as CM/miR-126Δ/Δ

first: Lianjun

last: Zhang

first: Dijiong

last: Wu

first: Man

last: Li

first: Shu

first: Wei

last: Chen


title: Role of activated PI3K-delta signaling in humoral immunity

city: Seattle

first: David

middle: J

last: Rawlings

address: 1900 9th Ave

institute: Seattle Children's Research Institute

summary: We report the high-throughput profiling of murine naive B cells, germinal center (dark zone and light zone) B cells, and plasma cells transcriptome. By obtaining over 5 million bases of sequence, we generated genome-wide expression maps of cell subsets from WT mice and aPIK3CD mice. We find that activated PIK3CD signaling lead to significant alteration in gene expression of plasma cells.

overall_design: Transcriptome profiling of naïve B cells, germinal centers (dark zone and light zone) B cells, and plasma cells to determine the effect of activated PIK3CD signaling

first: Fahd

middle: A

last: Qureshah

first: Chris

last: Thouvenel

first: Richard

last: James


title: Liver Homeostasis, Hepatocyte Ploidy, and Circadian Rhythm-controlled Gene Expression are Perturbed in Liver-Specific METTL3 Knockout Mice

city: Columbus

first: Kalpana

last: Ghoshal

state: OH

address: 420 W 12th Ave

institute: The Ohio State University

summary: METTL3 is essential for ESC differentiation and embryonic development, and its dysregulation is linked to various diseases including metabolic disorders and cancer. However, the consequence of m6A perturbation in the mammalian liver is not understood. Here, we provide insight into this by generating mice M3LKO (Mettl3fl/fl; Alb-Cre) mice carrying a hepatocyte-specific deletion of Mettl3.

overall_design: Liver mRNA profiles of 5-week old wild type and Mettl3 liver specific knockout mice

first: Juan

last: Barajas

first: Hui-Lung

last: Sun

first: Cho-Hao

last: Lin


title: IL-1 Mediates Microbiome-Induced Inflamm-Aging of Hematopoietic Stem Cells

city: Madrid

first: Santiago

middle: García

last: Martín

state: Madrid

address: Calle de Melchor Fernández Almagro, 3

department: Bioinformatics Unit

country: Spain

institute: Spanish National Cancer Research Centre

summary: Mature blood cells are maintained throughout life by hematopoietic stem cells (HSC). With aging, both HSC self-renewal as well as multi-lineage differentiation fidelity decline, a process determined by cell-intrinsic and –extrinsic factors. We here studied which aging-associated bone marrow (BM) alterations contribute to this process. Aged specific pathogen free (SPF) WT mice have increased systemic levels of microbial compounds compared to their young counterparts. This associates with increased steady-state IL-1a/b production by multiple BM cell populations. Aging-associated inflammatory transcriptional signatures and functional myeloid-differentiation bias in HSCs were mitigated in aged IL-1R1KO SPF and WT germ-free mice. Moreover, myeloid cells from aged mice produce more IL-1b and aged mice show higher and more durable IL-1a/b increase to lipopolysaccharide (LPS) challenges. Reducing inflammatory burden in aged mice by inhibiting IL-1 signaling or by antibiotic treatment rejuvenate HSCs functions. Collectively we define the microbiome-IL-1-IL1R1 axis as a key driver of HSC aging.

overall_design: RNA-sequencing of 30 mice bone marrow haematopoietic stem cells. Three genotypes: wildtype (WTF), bowel germ-free (GF) and IL1R1 KO(IL1RIKO). Three timepoints: 2 month, 1 year and 2 year for WT and KO mice and 2 month and 1 year for GF mice.

first: Larisa

middle: V

last: Kovtonyuk

first: Francisco

last: Caiado

first: Patrick

last: Helbling

first: Cesar

middle: N

last: Arrieta

first: Catharine

last: Aquino

first: Timothy

last: Sykes

first: Lennart

last: Opitz

last: Garcia-Martin

first: Fátima

last: Al-Shahrour


title: Human blastoids model blastocyst development and implantation [Bulk RNA-seq]

city: Vienna

first: Giovanni

last: Sestini

address: Dr.-Bohr-Gasse 3, 1030 Wien

country: Austria

institute: IMBA - Institute of Molecular Biotechnology

summary: Shortly after fertilization, human embryos implant into the uterus. This requires the formation of a blastocyst consisting of a sphere encircling a cavity lodging the embryo proper. Appropriate stem cells can form a blastocyst model, which we termed blastoid. Here we show that naive human pluripotent stem cells (hPSCs) triply inhibited for the Hippo, TGF-β, and ERK pathways consistently and efficiently (>70%) form blastoids that generate transcriptional pre-implantation analogs of the three founding lineages (trophoblast, epiblast, primitive endoderm; >97%) according to the  sequence and pace of blastocyst development. Blastoids spontaneously form an axis marked by the maturation of the polar region, which acquires the potential to specifically attach to hormonally stimulated endometrial cells, as during in utero implantation. Such human blastoids are scalable, versatile, and ethical models to explore human implantation and development.

overall_design: 61 samples


title: Predicting master transcription factors from pan-cancer expression data

last: Young

address: 9 Cambridge Center

institute: Whitehead Institute for Biomedical Research

summary: Critical developmental “master transcription factors” (MTFs) can be subverted during tumorigenesis to control expression of oncogenic transcriptional programs. Current approaches to identify MTFs rely on chromatin immunoprecipitation-sequencing data, which is currently unavailable for many cancer types. We developed the CaCTS (Cancer Core Transcription factor Specificity) algorithm to prioritize candidate MTFs using pan-cancer RNA-sequencing data. CaCTS identified candidate MTFs across 34 tumor types and 140 subtypes, including known MTFs. We also made novel predictions, including for cancer types/subtypes for which MTFs are unknown. This included PAX8, SOX17, and MECOM as candidate MTFs in ovarian cancer (OV). In OV cells, these factors are required for viability, lie proximal to super-enhancers, co-occupy regulatory elements globally and co-bind at critical gene loci encoding OV biomarkers. Identification of tumor MTFs, especially for tumor types with limited understanding of transcriptional drivers, paves the way to therapeutic targeting of MTFs in a broad spectrum of cancers.

overall_design: ChIP-Seq of H3K27ac and master transcription factors in ovarian cancer patient samples and cell lines

first: Jessica

last: Reddy

first: Marcos

last: Fonsseca

first: Rosario

middle: I

last: Corona

first: Robbin

last: Nameki

first: Felipe

last: Dezem

first: Isaac

last: Klein

first: Heidi

last: Chang

first: Daniele

last: Chaves-Moreira

first: Lena

last: Afeyan

first: Tathiane

last: Malta

first: Xianzhi

first: Forough

last: Abbasi

first: Alba

last: Font-Tello

first: Thais

last: Sabedot

first: Paloma

last: Cejas

first: Norma

last: Rodiguez-Malave

first: Ji-Huei

last: Seo

first: De-Chen

first: Ursula

last: Matulonis

first: Beth

middle: Y

last: Karlan

first: Simon

last: Gayther

first: Alexander

last: Gusev

first: Houtan

last: Noushmehr

first: Henry

last: Long

first: Matthew

last: Freedman

first: Ronny

last: Drapkin

first: Brian

last: Abraham

first: Kate

last: Lawrenson


title: A cell-based platform for the investigation of immunoproteasome subunit β5i expression and biology of β5i-containing proteasomes

status: Public on Sep 14 2021

city: Moscow

first: Sergei

last: Funikov

state: Moscow

address: Vavilov str. 32

country: Russia

institute: Engelhardt Insitute of Molecular Biology

summary: The degradation of intracellular proteins is a dynamic and tightly regulated process. Most of such substrates is hydrolyzed by proteasomes. Different forms of proteasomes are known. The role of immunoproteasomes (iPs) and intermediate proteasomes (intPs) is emerging. Here, using a CRISPR-Cas9 nickase technology a panel of four cell lines was obtained. Cells of histiocytic lymphoma, colorectal adenocarcinoma, cervix adenocarcinoma and hepatocarcinoma origin were modified to express proteasomes with mCherry-tagged β5i subunit, which is a catalytic subunit of iPs and intPs. Importantly, the chimeric gene expression in modified cells is under control of endogenous regulatory mechanisms and was increased following IFN-γ and/or TNF-α stimulation. Fluorescent proteasomes are catalytically active and were distributed within the nucleus and cytoplasm of the modified cells. RNAseq revealed marginal differences in gene expression profile between modified and initial cell lines, predominate metabolic pathways and a pattern of expressed receptors were determined for each cell line. Using obtained cell-lines it was shown that anti-cancer drugs ruxolitinib, vincristine and gefetinib stimulate the expression of β5i-containing proteasomes which might have an impact on the disease prognosis. Taken together, obtained cell lines represent convenient platform to study the immunoproteasome gene expression, localization of iPs and intPs, association of proteasomes with other proteins and many other aspects of proteasome biology under the influence of various drugs and conditions in real time and in living cells.

overall_design: RNA-seq analysis of modified and control cell lines


title: Transcriptomic analysis of the seminal vesicle response to the  reproductive toxicant acrylamide

city: Callaghan

first: John

middle: Even

last: Schjenken

state: NSW

address: Life Sciences Building, Level 4, Office 39. The University of Newcastle, University Drive

institute: University of Newcastle, Australia

summary: The seminal vesicles synthesise bioactive factors that support gamete function, modulate the female reproductive tract to promote implantation, and influence developmental programming of offspring phenotype. Despite the significance of the seminal vesicles in reproduction, their biology remains poorly defined. Here, to advance understanding of seminal vesicle biology, we analyse the mouse seminal vesicle transcriptome under normal physiological conditions and in response to acute exposure to the reproductive toxicant acrylamide. Mice were administered acrylamide (25 mg/kg bw/day) or vehicle control daily for five consecutive days prior to collecting seminal vesicle tissue 72 h following the final injection.

overall_design: Seminal vesicle tissue (n= 4 / grp) from acrylamide treated (25 mg/kg bw/day) or vehicle control (PBS) male Swiss mice was examined using RNA-sequencing. Mice were treated for 5 consecutive days prior to collecting tissue 72 h following the final injection. Note that seminal vesicle fluid was removed from tissue prior to storage and isolation of RNA

first: David

last: Skerrett-Byrne

first: Brett

last: Nixon

first: Elizabeth

middle: G

last: Bromfield

first: James

last: Breen

first: Natalie

last: Trigg

first: Simone

last: Stanger

first: Ilana

middle: R

last: Bernstein

first: Amanda

last: Anderson

first: Tessa

last: Lord

first: R

last: Aitken

first: Shaun

middle: D

last: Roman

first: Sarah

last: Robertson


title: Human blastoids model blastocyst development and implantation [scRNAseq]

first: Giovanni

last: Sestini

summary: Shortly after fertilization, human embryos implant into the uterus. This requires the formation of a blastocyst consisting of a sphere encircling a cavity lodging the embryo proper. Stem cells can form blastocyst models, which we termed blastoid. Here we show that naïve human pluripotent stem cells (hPSCs) triply inhibited for the Hippo, TGFb, and ERK pathways consistently and efficiently (>70%) form blastoids that generate transcriptional pre-implantation (>95%) analogs of the three founding lineages (trophoblast, epiblast, hypoblast), and in the sequential and timely manner of blastocysts. Blastoids spontaneously form an axis marked by maturation of the polar region, which acquires the potential to specifically attach to hormonally-stimulated endometrial cells, as during in utero implantation. Such human blastoids are scalable, versatile, and ethical models to explore human implantation and development.

overall_design: Blastoids and stem cells with various treatments

first: Harunobu

last: Kagawa

first: Alok

last: Javali

first: Heidar

middle: H

last: Khoei

first: Theresa

middle: M

last: Sommer

first: Yvonne

last: Reimer

first: Nicolas

last: Rivron


title: Spatial transcriptomics (Visium, Spatial 3' V1, 10x Genomics) analysis of heart tissues in a murine model of Kawasaki disease vasculitis

city: Boston

first: David

last: Zemmour

address: 77 avenue Louis Pasteur

institute: Harvard Medical Schol

summary: Spatial transcriptomics (Visium, Spatial 3' V1, 10x Genomics) analysis of heart tissues from Lactobacillus casei cell wall extract (LCWE)-injected mice and control PBS-injected mice

overall_design: Heart from one LCWE-injected and one PBS-injected mice were harvested one week after injection and sectioned to 10µm and used for Visium Spatial Transcriptomics 3' V1

first: Rebecca

last: Porritt

first: Magali

last: Rivas

first: Moshe

last: Arditi


title: Single-cell RNAseq (10x Genomics) analysis of the abdominal aorta in a murine model of Kawasaki disease vasculitis

summary: Single-cell RNAseq (10x Genomics) analysis of the abdominal aorta in a murine model of Kawasaki Disease vasculitis (Lactobacillus casei cell wall extract, LCWE) and in control PBS-injected mice

overall_design: Cells were isolated from the abdominal  aorta of WT C57BL/6J mice 2 weeks after peritoneal injection of LCWE or PBS. The PBS group was composed of a pool of 9 abdominal aortas, whereas the LCWE group was composed of a pool of 7 abdominal aortas. Single cell suspension were encapsulated for 10x Genomics Single Cell 3′ Reagent Kit (V3 chemistry).

last: Noval Rivas


title: CD4+ T cells contribute to neurodegeneration in Lewy body dementia [CSF_and_PBMCs_Healthy and LBD]

city: Chicago

last: Gate

state: IL

address: 303 E Chicago Ave

institute: Northwestern University

summary: Recent studies indicate that the adaptive immune system plays a role in Lewy body dementia (LBD). However, the mechanism regulating T cell brain homing in LBD is unknown. Here, we observed T cells adjacent to Lewy bodies and dopaminergic neurons in post-mortem LBD brains. Single-cell RNA sequencing of cerebrospinal fluid (CSF) identified upregulated expression of C-X-C Motif Chemokine Receptor 4 (CXCR4) in CD4+ T cells in LBD. CSF protein levels of the CXCR4 ligand, C-X-C Motif Chemokine Ligand 12 (CXCL12) were associated with neuroaxonal damage in LBD. Furthermore, we observed clonal expansion and upregulated Interleukin 17A expression by CD4+ T cells stimulated with a phosphorylated α-synuclein epitope. Thus, CXCR4-CXCL12 signaling may represent a mechanistic target for inhibiting pathological interleukin-17-producing T cell trafficking in LBD.

overall_design:
scRNAseq of CSF cells from healthy (n=11) and LBD (n=11) subjects. Live cells were sorted with a live/dead SYTOX blue dye. Sequencing libraries were prepared using the 10X Genomics platform.

PBMCs from some of these subjects were also sequenced by scRNAseq.

first: Olivia

last: Leventhal

first: Tobias

last: Fehlmann


title: mRNA vaccination induces tick resistance and prevents transmission of the Lyme Disease agent

status: Public on Sep 13 2021

city: Hangzhou

first: Xiaotian

last: Tang

state: Zhejiang

address: Agro-Bio-Environ Building, Zhejiang University

department: Entomology

country: China

institute: Zhejiang University

summary: An Ixodes scapularis saliva mRNA vaccine induces tick resistance and prevents Borrelia burgdorferi infection in guinea pigs

overall_design: Examination of the immune responses associated with mRNA vaccination and host protection, using RNA-seq

first: Sajid

last: Andaleeb

first: Matias

last: Jaqueline

first: Arora

last: Gunjan

first: Fikrig

last: Erol


title: Gene expression profiles of Lgr5-GFP high and Lgr5-GFP low cells in small intestines of Itgb7+/+ Lgr5-GFP mice and Itgb7-/- Lgr5-GFP mice

city: Shanghai

first: Chen

last: Shiyang

address: Room 413, New biochemistry building, YueYang Road

institute: Shanghai Institute of Biochemistry and Cell Biology (CAS)

summary: Intestinal stem cell (ISC) differentiation is regulated precisely by a niche in the crypt, where lymphocytes may interact with stem and transient amplifying (TA) cells. However, whether and how lymphocyte-stem/TA cell contact affects ISC differentiation is largely unknown. Here, we uncover a novel role of T cell-stem/TA cell contact in ISC fate decisions. We show that intestinal lymphocyte depletion results in skewed ISC differentiation in mice, which can be rescued by T cell transfer. Mechanistically, integrin αEβ7 expressed on T cells binds to E-cadherin on ISCs and TA cells, triggering E-cadherin endocytosis and the consequent Wnt and Notch signaling alterations. Blocking αEβ7—E-cadherin adhesion suppresses Wnt signaling and promotes Notch signaling in ISCs and TA cells, leading to defective ISC differentiation. Thus, αEβ7+ T cells regulate ISC differentiation at single-cell level through cell-cell contact-mediated αEβ7—E-cadherin adhesion signaling, highlighting a critical role of the T cell-stem/TA cell contact in maintaining intestinal homeostasis.

overall_design: Gene expression profiles of Lgr5-GFP high (stem) and Lgr5-GFP low ( transient amplifying) cells in small intestines of Itgb7+/+ Lgr5-GFP mice and Itgb7-/- Lgr5-GFP mice

first: Jianfeng

last: Chen


title: PARP-1/PARP-2 double deficiency results in recurrent early pregnancy loss and decidualization failure in mice

first: W. Lee

last: Kraus

state: TX

address: 5323 Harry Hines Blvd.

institute: UT Southwestern Medical Center

summary: Specifically, we found inhibition of PARP-1 and PARP-2 catalytic activity, or tissue-specific genetic knockout, results in a severe fertility defect. Transcriptomic analyis uncovered that  PARP-1;PARP-2-deficient mice have defects in the progression of stromal cell decidualization manifesting in pregnancy failure

overall_design: RNAseq libraries were generated from control and knockout implantation sites on gestational days 4, 5, and 6.

first: Andrew

last: Kelleher

first: Rohit

last: Setlem

first: Lee


title: RNA-seq analysis of MEF cells with stably overexpression of IDH2 mutation and MEF cells treated with 2-HG

city: Minato-ku, Tokyo

first: XUN

last: LIU

address: Shirokanedai 4-6-1

country: Japan

institute: The University of Tokyo

summary: Isocitrate dehydrogenase 2 (IDH2) mutations and its key effector 2-hydroxyglutarate (2-HG) have been reported to promote oncogenesis in various human cancers. To elucidate molecular mechanism(s) associated with IDH2 mutations, we established mouse embryonic fibroblasts (MEF) cells stably expressing cancer-associated IDH2R172S mutation and treated MEF cells with or without 2-HG. The expression of genes in these cells were analyzed by RNA-seq.

overall_design: Expression profiles of MEF cells stably expressing mutant IDH2 or mock vector, and MEF cells treated with 300 μM of 2-HG or DMSO for 24 h

first: Xun

last: Liu

first: Tsuneo

last: Ikenoue

first: Yoichi

last: Furukawa

first: Kiyoshi

last: Yamaguchi


title: Exploring the Effect and Mechanisms of Telomerase Inhibition as A Novel Therapeutic Strategy for Glioma [2224, DBTGR-05MG, A172]

city: Durham

first: Gao

last: Zhang

state: NC

address: 203 Research Dr | MSRB1, Rm 179

department: The Preston Robert Tisch Brain Tumor Center

institute: Duke University School of Medicine

summary: The obejctive of this study is to establish the therapeutic role of 6-thio-dG for gliomas in vitro and in vivo and to elucidate the anti-tumor mechanisms of 6-thio-dG in glioma cells. And the ultimate goal of this proposal is to demonstrate the potential of 6-thio-dG as a novel strategy for primary and recurrent glioma.

overall_design: RNAseq of Glioma Cells treated with 6-thio-dG or DMSO

first: Shengnan

last: Yu


title: Spiny mice (Acomys cahirinus) activate unique transcriptional programs after severe kidney injuries and regenerate organ function without fibrosis

first: Daryl

last: Okamura

state: Washington

department: Center for Developmental Biology & Regenerative Medicine

institute: Seattle Children’s Research Institute

summary:
Purpose: The goal of this study was to characterize the kidney transcriptome of Mus musculus and Acomys cahirinus after unilateral ureteral obstruction (UUO) kidney injury.
Methods: Kidney mRNA-seq profiles of 10 week old mouse and spiny mouse were generated at 2 day and 5 days after unilateral ureteral obstruction injury in triplicate, using Illumina NovaSeq 6000. The sequence reads that passed quality filters were analyzed at the transcript isoform level with Salmon, edgeR and a limma-voom pipeline in R.
Results: For both Mus musculus and Acomys cahirinus kidneys, we mapped about 50 million sequence reads per sample to the mouse transcriptome and identified 20580 transcripts in kidneys of Mus musculus and 54075 transcripts in the kidneys of Acomys cahirinus. Using 1.5 fold change and FDR < 0.05, number of transcripts that are significantly different between Mus musculus samples: 3915 between normal and day 2 after UUO, 5365 between normal and day 5 after UUO. For Acomys cahirinus: 1765 between normal and day 2 after UUO, 2499 between normal and day 5 after UUO.
Conclusions: Our study demonstrate there were many conserved responses to kidney injury between M. musculus and A. cahirinus despite the divergent outcomes for kidney fibrosis.

sample_organism:
Acomys cahirinus
Mus musculus

platform_organism:
Acomys cahirinus
Mus musculus

overall_design: Kidney mRNA profiles of 10 week old mouse and Spiny mouse 2 and 5 days after unilateral ureteral obstruction kidney injury.

first: Mark

last: Majesky


title: Exploring the Effect and Mechanisms of Telomerase Inhibition as A Novel Therapeutic Strategy for Glioma


title: STL-seq reveals distinct pause release and termination kinetics for promoter-proximal paused RNA polymerase II transcripts

city: Orange

first: Matthew

last: Simon

state: CT

address: West Campus, 100 West Campus Drive, Ste MIC312A

department: MBB

institute: Yale University

summary: Despite the critical regulatory function of promoter-proximal pausing, the influence of pausing kinetics on transcriptional control remains an active area of investigation. Here, we present Start-TimeLapse-seq (STL-seq), a method that captures the genome-wide kinetics of short, capped RNA turnover and reveals principles of regulation at the pause site. By measuring the rates of release into elongation and premature termination through inhibition of pause release, we determine that pause-release rates are highly variable and most promoter-proximal paused RNA Polymerase II molecules prematurely terminate (~80%). The preferred regulatory mechanism upon a hormonal stimulus (20-hydroxyecdysone) is to influence pause-release rather than termination rates. Transcriptional shutdown occurs concurrently with induction of promoter-proximal termination under hyperosmotic stress but paused transcripts from TATA box-containing promoters remain stable, demonstrating an important role for cis-acting DNA elements in pausing. STL-seq dissects the kinetics of pause release and termination, providing an opportunity to identify mechanisms of transcriptional regulation.

sample_organism:
Drosophila melanogaster
Homo sapiens

platform_organism:
Drosophila melanogaster
Homo sapiens

overall_design: RNA isolated from Drosophila S2 cells treated +/- ecdysone hormone, +/- flavopiridol inhibitor, and +/- s4U or with triptolide +s4U was either subjected to oxidative-nucleophilic-aromatic-substitution chemistry and enrichment of short, capped RNA species or biotinylation and enrichment of s4U-labeled RNA followed by oxidative-nucleophilic-aromatic-substitution chemistry. RNA isolated from human HEK293T cells treated +/- KCl and +/- s4U was subjected to oxidative-nucleophilic-aromatic-substitution chemistry and enrichment of short, capped RNA species. Sequencing libraries were prepared from all RNA.

first: Joshua

middle: T

last: Zimmer

first: Nicolle

last: Rosa-Mercado

first: Daniele

last: Canzio

first: Joan

last: Steitz


title: Chemical capping improves template switching and enhances sequencing of small RNAs

city: Ipswich

first: Madalee

last: Wulf

address: 240 County Road

institute: New England Biolabs

type: Non-coding RNA profiling by high throughput sequencing

summary: We deployed CapTS-seq for sequencing synthetic miRNAs, human total brain RNA, and liver FFPE RNA, and demonstrated that chemical capping in conjunction with template switching consistently reduces sequencing bias and improves library quality in comparison with commercially available RNA-seq kits. Finally, we showed the simultaneous detection of miRNAs and mRNAs in FFPE derived samples, underscoring the potential of this workflow for dissecting regulatory networks between miRNAs and their target gene transcripts.

sample_organism:
Homo sapiens
synthetic construct

platform_organism:
Homo sapiens
synthetic construct

overall_design: Examined small RNA and transcriptome content in synthetic miRNAs and total brain RNA and liver FFPE RNA using either CapTS-seq or commercially available SMARTer smRNA Seq Kit from Takara.

first: Zhiyi

last: Sun

first: Sean

last: Maguire


title: Human blastoids model blastocyst development and implantation

first: Giovanni

last: Sestini

summary: About one week after fertilization, human embryos implant into the uterus. This necessitates the formation of a blastocyst consisting of a sphere encircling a cavity lodging the embryo proper. Stem cells can form a blastocyst model, which we termed blastoid. Here we show that naive human pluripotent stem cells (PXGL hPSCs) efficiently (>70%) form blastoids generating blastocyst-stage analogs of the 3 founding lineages (>97% trophectoderm, epiblast, and primitive endoderm) according to the sequence and to the pace of blastocyst development. Blastoids form the first axis and we observe that the epiblast induces the maturation of the polar trophectoderm that consequently acquires the specific and transient potential to attach to hormonally-stimulated endometrial cells. Such human blastoids are faithful, scalable, versatile, and ethical models to explore human implantation and development.

overall_design: Refer to individual Series


title: Microglia-specific overexpression of alpha-Synuclein leads to severe dopaminergic neurodegeneration by phagocytic exhaustion and oxidative toxicity [single-cell RNA-Seq]

city: Milano

first: Luca

last: Massimino

state: MI

address: Via Olgettina, 58

department: Gastroenterology

country: Italy

institute: Ospedale San Raffaele

summary: Increasing evidence from recent findings in human samples and animal models support  the involvement of inflammation in the development of Parkinson’s disease (PD).  Nevertheless, it is currently unknown whether microglia activation could also be a  primary event leading to neurodegeneration in PD. We have generated a new mouse  model with strictly selective alpha-Synuclein (aSYN) accumulation in microglial cells by  inducible gene expression through lentiviral transduction in the nigral tissue.  Surprisingly, these mice develop progressive and restricted degeneration of  dopaminergic (DA) neurons without any sign of aSYN endogenous aggregation. aSYN  accumulating microglial cells exhibit a strong inflammatory status with production of  pro-inflammatory cytokines and chemokines. Bulk and scRNA-sequencing of the  immune cellular infiltrate identify a complex cellular composition with numerous  peripheral populations of the innate and adaptive immune system with specific  transcriptional signatures. Molecular profiling and functional assessment reveal that  intoxicated microglia develop exhausted phagocytosis and high production of oxidative  molecules creating a molecular feed-forward vicious cycle with IFNg secreting immune  cells infiltrated in the brain parenchyma. In these animals, pharmacological inhibition  of the oxidative and nitrosative molecule production is sufficient to attenuate  neurodegeneration. These results suggest that aSYN accumulation in microglia exerts  selective DA neuronal degeneration by promoting phagocytic exhaustion, excessive  toxic environment and selective recruitment of peripheral immune cells.

overall_design: 1x CTRL (infected with GFP), 1x SNCA3 (CX3-Cre infected with Synuclein-expressing vector)

first: Simone

last: Bido

first: Vania

last: Broccoli


title: RRBS-based blood methylome in chronic nociceptive (NOCI) and neuropathic (NEURO) pain suffering patients and controls (CTL).

city: Geneva

first: Ludwig

last: Stenz

address: Rue Michel-Servet 1

department: Department of Genetic Medecine and Development

country: Switzerland

institute: University of Geneva

type: Methylation profiling by high throughput sequencing

summary:
Nociceptive pain implies the activation of the nociceptors without damage to the somatosensory nervous system.
Neuropathic pain implies an injury or a disease of the central or peripheral nervous system.
Epigenomic may participate in chronic pain conditions as well as in the transition from acute to chronic pain.
The purpose of the present study is to analyze how blood methylome differs in both mentionned types of chronic pain, by comparing them together and to controls.
The study protocol was approved by the local ethic committee (CCVEM 034/12) and was conducted according to the recommendation of the Declaration of Helsinki.
Patients were enrolled at the Clinique romande de réadaptation (CRR) in Sion, in Switzerland.
Data were hosted and analyzed at the University of Geneva, Faculty of Medicine, Department of Genetic Medicine and Development, in Switzerland.
Data were processed in the high performance computing cluster (HPC) nammed "Baobab" in the Geneva University.

overall_design: Three comparisons were performed: 1) CTL versus NOCI, 2) CTL versus NEURO and 3) NOCI versus NEURO.

first: Ariane

last: Giacobino

first: Bertrand

last: Leger


title: Phenotypic drug screening in a human fibrosis model identified a novel class of antifibrotic therapeutics

city: Neuherberg

first: Johannes

last: Beckers

address: Ingolstaedter Landstr. 1

department: Institute of Experimental Genetics

institute: Helmholtz Zentrum Muenchen

summary: Fibrogenic processes instigate fatal chronic diseases leading to organ failure and death. Underlying biological processes involve induced massive deposition of extracellular matrix (ECM) by aberrant fibroblasts. We subjected diseased primary human lung fibroblasts to an advanced 3D phenotypic high-content assay and screened a library of FDA/EMA approved small molecules for inhibiting ECM deposition. Fibrotic Pattern Detection by Artificial Intelligence (FANTAIL) identified Tranilast as an effective inhibitor, however, by structure-activity relationship studies we found N-(2-butoxyphenyl)-3-(phenyl)acrylamides (N23Ps) as a novel and highly potent compound class. N23Ps suppressed myofibroblast transdifferentiation, ECM deposition, cellular contractility, and altered cell shapes, thus advocating a unique mode of action. Mechanistically, transcriptomics identified SMAD (de)ubiquitination/Smurf2 as a potential therapeutic target network. Antifibrotic activity of N23Ps was verified by proteomics in a human ex vivo tissue fibrosis disease model, suppressing profibrotic markers SERPINE1/PAI1 and CXCL8/IL8. Conclusively, these data suggest N23Ps as a novel class of highly potent compounds with implications for inhibiting organ fibrosis in patients.

overall_design: We performed gene expression microarray analysis in primary human pulmonary fibroblasts from patients with idiopathic pulmonary fibrosis. Cells were treated with TGFb1, cmp4 or with a combination of both

first: Michael

last: Gerckens

first: Kenji

last: Schorpp

first: Francesco

last: Pelizza

first: Melanie

last: Wögrath

first: Kora

last: Reichau

first: Huilong

last: Ma

first: Armando-Marco

last: Dworsky

first: Arunima

last: Sengupta

first: Mircea-Gabriel

last: Stoleriu

first: Katharina

last: Heinzelmann

first: Juliane

last: Merl-Pham

first: Martin

last: Irmler

first: Hani

last: Alsafadi

first: Eduard

last: Trenkenschuh

first: Lenka

last: Sarnova

first: Marketa

last: Jirouskova

first: Wolfgang

last: Frieß

first: Steffanie

last: Hauck

first: Nikolaus

last: Kneidinger

first: Jürgen

last: Behr

first: Anne

last: Hilgendorff

first: Kamyar

last: Hadian

last: Lindner

last: Königshoff

first: Oliver

last: Eickelberg

last: Gregor

last: Plettenburg

first: Ali-Önder

last: Yildirim

first: Gerald

last: Burgstaller


title: Microglia-specific overexpression of alpha-Synuclein leads to severe dopaminergic neurodegeneration by phagocytic exhaustion and oxidative toxicity

first: Luca

last: Massimino

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


title: Microglia-specific overexpression of alpha-Synuclein leads to severe dopaminergic neurodegeneration by phagocytic exhaustion and oxidative toxicity [bulk RNA-Seq]

overall_design: 3xCTRL (infected with GFP), 3xCX3-Cre (infected with Synuclein-expressing vector), 3xDAT-Cre (infected with Synuclein-expressing vector)

first: Simone

last: Bido

first: Vania

last: Broccoli


title: Alpha-ketoglutarate induces lipidome remodeling and mitochondrial metabolism to inhibit regulatory T cell differentiation

city: Bethesda

first: Amir

last: Foroushani

state: MD

address: 10 CENTER DR RM 11N306

department: NIAID

institute: NIH

summary: The differentiation of CD4 T cells is regulated by glycolysis and mitochondrial oxidative phosphorylation (OXPHOS), integrating the cell’s metabolism with transcriptional and epigenetic changes. However, the precise mechanisms coupling tricarboxylic (TCA) acid cycle intermediates to CD4 T cell differentiation remain unclear. Here we demonstrate that a-ketoglutarate (aKG) increased Th1 polarization while significantly inhibiting regulatory T cell (Treg) generation. Consistent with these data, aKG promoted the effector profile of Treg-polarized chimeric antigen receptor-engineered T cells against the ErbB2 tumor antigen. Mechanistically, aKG altered the transcription of >250 membrane-related genes, decreasing membrane fluidity and inducing a robust lipidome-wide remodelling. The massive increase in storage and mitochondria lipids was associated with a significantly augmented OXPHOS. Notably, inhibition of succinate dehydrogenase activity, the bridge between the TCA cycle and the electron transport chain, enforced Treg generation. Thus, our study identifies novel connections between aKG, lipidome remodeling and OXPHOS in CD4 T cell fate decisions.

overall_design: mRNA profiles of mouse naïve T cells, as well as T cells under Treg and Th polarizing conditions with and without alpha-ketoglutarate.

first: Madeline

last: Wong

middle: K

first: Stefan

last: Muljo

first: Naomi

last: Taylor

first: Valerie

last: Dardalhon

first: Carmen

last: Yong

first: Maria

middle: F

last: Matias


title: G protein-coupled estrogen receptor stimulates human trophoblast cell invasion via YAP-mediated ANGPTL4 expression

status: Public on Sep 12 2021

city: Zhengzhou

first: Jung-Chien

last: Cheng

state: Henan

address: 40, Daxue Road

institute: The First Affiliated Hospital of Zhengzhou University

summary: RNA sequencing was applied to HTR-8/SVneo human trophoblast cells in response to G1, an agonist of GPER, to identify the target genes of GPER.

overall_design: HTR-8/SVneo cells were treated with 1 µM G1 for 24 hours.


title: CDC7 inhibition in PLC/PRF/5 cells

first: Cun

last: Wang

address: Xietu Road

institute: Shanghai Cancer Institute

summary: Purpose: to analyze gene expression in XL413 treated liver cancer cells.  Methods: PLC/PRF/5 cells are treated with XL413 (10uM) for 96 hours .For RNA sequencing, the library was prepared using TruSeq RNA sample prep kit according to the manufacturer’s protocol (Illumina). Gene set enrichment analysis was performed using gene set enrichment analysis software.

overall_design: RNA seq data of PLC/PRF/5-control, PLC/PRF/5_ XL413 treatment,  to check the effects of XL413 on globle gene expression.


title: DLEU1 promotes oral squamous cell carcinoma progression through activating interferon-stimulated genes

city: Sapporo

first: Hiromu

last: Suzuki

state: Hokkaido

address: S1, W17, Chuo-ku

department: Department of Molecular Biology

institute: Sapporo Medical University


title: IFN-gamma is essential for alveolar macrophage driven pulmonary inflammation in macrophage activation syndrome

city: Cincinnati

first: Nathan

last: Salomonis

address: 3333 Burnet Avenue

department: Biomedical Informatics

institute: Cincinnati Children's Hospital

summary: Macrophage activation syndrome (MAS) is a life-threatening cytokine storm syndrome complicating systemic juvenile idiopathic arthritis (SJIA) and driven by IFN-gamma. SJIA and MAS are also associated with an unexplained emerging inflammatory lung disease (SJIA-LD), with our recent work supporting pulmonary activation of IFN-gamma pathways as a pathologic link between SJIA-LD and MAS. Our objective was to mechanistically define the novel observation of pulmonary inflammation in the TLR9 mouse model of MAS. In acute MAS, lungs exhibit mild but diffuse CD4-predominant, perivascular interstitial inflammation with elevated IFN-gamma, IFN-induced chemokines, and alveolar macrophage expression of IFN-gamma-induced genes. Single-cell RNA-sequencing confirmed IFN-driven transcriptional changes across immune and parenchymal lung cell types. Resolution of MAS was associated with increased alveolar macrophage and interstitial lymphocytic infiltration. alveolar macrophage microarrays confirmed IFN-gamma-induced proinflammatory polarization during acute MAS, which switches towards an anti-inflammatory phenotype during MAS resolution. Interestingly, recurrent MAS led to increased alveolar inflammation and lung injury, and reset alveolar macrophagepolarization towards a proinflammatory state. Furthermore, in mice bearing macrophages insensitive to IFN-gamma, both systemic feature of MAS and pulmonary inflammation were attenuated. These findings demonstrate that experimental MAS induces IFN-gamma-driven pulmonary inflammation replicating key features of SJIA-LD, and provides a model system for testing novel treatments directed towards SJIA-LD.

overall_design: Single-cell RNA-sequencing of lung immune and parenchymal cell types in macrophage activation syndrome

first: Denny

last: Gao

first: Maggie

last: Henderlight

first: Christopher

last: Woods

first: Alexei

last: Grom

first: Sherry

last: Thornton

first: Michael

middle: B

last: Jordan

first: Kathryn

last: Wikenheiser-Brokamp

first: Grant

middle: S

last: Schulert


title: DLEU1 promotes oral squamous cell carcinoma progression through activating interferon-stimulated genes [ChIP-Seq]

summary: Long noncoding RNAs (lncRNAs) are deeply involved in cancer development, and we have previously reported that DLEU1 (deleted in lymphocytic leukemia 1) is frequently overexpressed in oral squamous cell carcinoma (OSCC) cells, where it plays an oncogenic role. In this study, we aimed to further clarify the molecular function of DLEU1 in the pathogenesis of OSCC. Chromatin immunoprecipitation-sequencing (ChIP-seq) analysis revealed that knockdown of DLEU1 induced substantial changes in the levels of histone H3 lysine 4 trimethylation (H3K4me3) and H3K27 acetylation (H3K27ac) in OSCC cells. Notably, depletion of DLEU1 significantly downregulated levels of H3K4me3/H3K27ac and expression of a number of interferon-stimulated genes (ISGs) including IFIT1, IFI6 and OAS1, while ectopic DLEU1 expression activated these genes. Western blot analysis and reporter assay suggested that DLEU1 upregulates ISGs through activating the JAK-STAT signaling in OSCC cells. Moreover, IFITM1, one of the ISGs induced by DLUE1, is frequently overexpressed in primary OSCC tumors, and knockdown of IFITM1 attenuated OSCC cell proliferation, migration and invasion. Our data suggest that DLEU1 exerts its oncogenic function through, at least in part, activating a series ISGs in OSCC cells.

overall_design: HSC3 cells were transfected with siDLEU1 or Control siRNA and were incubated for 72 hours.

first: Takeshi

last: Niinuma


title: DLEU1 promotes oral squamous cell carcinoma progression through activating interferon-stimulated genes [microarray]

overall_design: Ca9-22 cells were infected with a DLEU1 lentiviral vector or a control vector and were selected with 6mg/ml of Blasticidin for 2 weeks.


title: The viral oncoproteins Tax and HBZ control the host mRNA splicing landscape

city: Liège

first: Jean-Claude

last: Twizere

state: Liège

address: CHU - B34 Quartier Hôpital Avenue de l'Hôpital,11

department: Molecular Biology of Diseases

country: Belgium

institute: GIGA Institute

summary: We demonstrate that the HTLV-1 oncoviral proteins Tax and HBZ differentially alter the transcriptome landscape in T-cells

overall_design: RNA sequencing was performed on control and Jurkat cells, conditionally expressing Tax or HBZ viral proteins. Three biological replicates were preared for each condition.

first: Charlotte

last: Vandermeulen

first: Tina

middle: O

last: Glady

first: Benoit

last: Charloteaux


title: Transcriptome analyses reveal tau isoform-driven changes in transposable element and gene expression

city: La Jolla

first: Robert

last: Rissman

state: California

address: 9500 Gillman Drive

institute: University of California, San Diego

summary:
Purpose: Overexpression of tau isoforms has been linked to neurodegenerative diseases, and abnormal tau expression is correlated with genomic instability. The purpose of this study is to use RNA-sequencing and molecular biological techniques to determine if there are any unique genomic consequences associated with overexpression of one or more tau isoforms.
Methods: SHSY-5Y cells were infected in triplicate with lentiviruses to induce overexpression of 3R and 4R tau isoforms, were treated with amyloid-beta oligomers or DMSO, and were differentiated into neurons using retinoic acid. After RNA collection via TRIzol, poly(A)-selected, 100 bp, paired-end library preparation and sequencing were completed by the IGM Genomics Center, University of California, San Diego, La Jolla, CA, using an Illumina NovaSeq 6000 that was purchased with funding from a National Institutes of Health SIG grant (#S10 OD026929). Trimmed FASTQ files were aligned to GRCH38 using STAR and differential expression of genes and transposable elements was determined using DESeq2 and software from TEToolkit (available from the Hammell Lab at Cold Spring Harbor Laboratory).
Results: Our RNA-seq analysis revealed similar gene and transposable element expression patterns between cells overexpressing tau isoforms and patients with Alzheimer's disease or progressive supranuclear palsy. We found widespread differential expression particularly in L1 and Alu transposable elements and upregulation of genes related to extracellular matrix organization, focal adhesions, and cell junctions, among other biological processes.
Conclusions: Our study is the first to our knowledge to examine locus-specific transposable element expression in the context of tauopathies, and to our knowledge is the first to specifically examine transposable element expression stemming from overexpression of tau isoforms. Our study provides additional evidence that transposable elements may play a role in tauopathies and provides further insight into the locations of transposable element expression in the genome in the context of neurodegenerative diseases.

overall_design: Lentiviral infection to produce tau isoform overexpression in neuronally differentiated SH-SY5Y cells, along with treatment with DMSO or amyloid-beta oligomers to examine transcriptomic effects (N=3).

first: Jennifer

last: Grundman

first: Brian

last: Spencer

first: Floyd

last: Sarsoza

middle: A


title: Epigenome analysis of primary intracranial sarcoma

city: Hamburg

first: Martin

last: Mynarek

address: Martinistr. 52

department: Department for Pediatric Hematology and Oncology

institute: University Medical Center Hamburg-Eppendorf

type: Methylation profiling by genome tiling array

summary: Genome wide DNA methylation profiling of DNA extracted from formalin-fixed parffin embedded tumor samples from patients with primary intracranial sarcoma from Peru. The Illumina MethylationEPIC array was used to obtain DNA methylation profiles of 850.000 CpG sides

overall_design: Bisulphite converted DNA from 28 tumor samples

first: Ulrich

last: Schüller


title: Next Generation Sequencing facilitates Quantitative Analysis of mouse CD4+ T cells affected by Gpr120

status: Public on Sep 11 2021

city: Galveston

first: Yingzi

last: Cong

address: 301 University Boulevard

institute: University of Texas Medical Branch at Galveston

summary: The goal of this study is to analyze the effect of Gpr120 in mouse CD4+ T cells.

overall_design: Examine the different gene profiles between CD4+ T cells treated with or without Gpr120 agonist CpdA using RNA seq


title: Inosine monophosphate and inosine differentially regulate endotoxemia and bacterial sepsis

city: New York

first: George

last: Hasko

state: NY

address: 630 West 168th Street

department: Anesthesiology

institute: Columbia University Medical Center

summary: Inosine monophosphate (IMP) plays a central role in intracellular purine metabolism. How IMP regulates inflammation induced by bacterial products or bacteria is unknown. In this study, we demonstrate that IMP fundamentally alters gene expression in the liver.To assess how IMP affects inflammatory cell activation in vivo, we subjected liver, the organ containing the highest numbers of macrophages in the body, to RNAseq analyses. High quality RNA was isolated from the livers of 3 vehicle- and 3 IMP-treated endotoxemic mice and submitted for RNAseq analysis. Our analysis identified 105 genes that were differentially expressed, determined by adjusted p values (42 downregulated, 63 upregulated; padj ≤ 0.05).

overall_design: RNAseq analyses in the liver tissue of vehicle- and IMP-treated endotoxemic mice

first: Marianna

last: Lovaszi


title: Lin28A/CENPE promoting the proliferation and chemo-resistance of acute myeloid leukemia

status: Public on Sep 10 2021

city: zhengzhou

first: Mingyue

last: Shi

state: China

address: no.7 weiwu road

institute: Henan Provincial People’s Hospital

summary: Peripheral blood samples of 3 refractory/relapsed AML patients (R/R-AML, relapsed/refractory AML patients who failed to achieve complete remission/CR after 2 courses of induction chemotherapy), 3 refractory secondary AML patients (S-AML, MDS or MPN derived AML patients did not reach CR after 2 rounds of induction chemotherapy), 4 de novo AML patients (AML, CR after standard “3+7” induction chemotherapy), and 3 healthy controls (HC) were collected. Nanodrop was applied to quantify the total RNA samples. Illumina kits were used to prepare the RNA-seq library.

overall_design: RNA-seq of de novo AML, secondary AML, and refractory/relapsed AML

first: Yuqing

last: Chen

first: Kai

last: Sun


title: Negative Feedback Regulation of Cholesterol Biosynthesis by MMAB

city: New Haven

first: Rolando

last: Garcia-Milian

address: 333 Cedar St

department: CWML

summary: Intricate regulatory networks govern the net balance of cholesterol biosynthesis, uptake and efflux; however, the mechanisms surrounding cholesterol homeostasis remain incompletely understood. Here, we develop an integrative genomic strategy to detect regulators of LDLR activity and identify 250 genes whose knockdown affects LDL-cholesterol uptake and whose expression is modulated by intracellular cholesterol levels in human hepatic cells. From these hits, we focus on MMAB, an enzyme which catalyzes the conversion of vitamin B12 to adenosylcobalamin, and whose expression has previously been linked with altered levels of circulating cholesterol in humans. We demonstrate that hepatic levels of MMAB are modulated by dietary and cellular cholesterol levels through SREBP2, the master transcriptional regulator of cholesterol homeostasis.  Knockdown of MMAB decreases intracellular cholesterol levels and augments SREBP2-mediated gene expression and LDL-cholesterol uptake in human and mouse hepatic cell lines. Reductions in total sterol content were attributed to increased intracellular levels of propionic and methylmalonic acid and subsequent inhibition of HMGCR activity and cholesterol biosynthesis. Moreover, mice treated with antisense inhibitors of MMAB display a significant reduction in hepatic HMGCR activity, hepatic sterol content and increased expression of SREBP2-mediated genes. Collectively, these findings reveal an unexpected role for the adenosylcobalamin pathway in regulating LDLR expression and identify MMAB as an additional control point by which cholesterol biosynthesis is regulated by its end product.

overall_design: Human hepatic cells (Huh7) were cultured in Dulbecco’s Modified Eagle Medium (DMEM) containing 10% FBS and 120 μg/ml native LDL (nLDL, to cholesterol enrich), 5 μM statin (to cholesterol deplete) or vehicle control for 24 h. Following this, cells were harvested in Trizol and RNA was purified using the RNeasy Isolation Kit (Qiagen). The purity and integrity of total RNA was verified using the Agilent Bioanalyzer and hybridized to the Affymetrix GeneChip Human Genome U133 Plus 2.0 Array according to the manufacturer’s instructions. Two biological replicates for each condition were used for analysis. Raw data were normalized and analyzed by GeneSpring GX software version 11.5 (Agilent Technologies).

first: Leigh

last: Goedeke

first: Carlos

last: Fernandez-Hernando


title: Chronic mineral oil administration increased hepatic inflammation in wild type mice compared to lipocalin 2 null mice

city: Cologne

first: Ali

last: Abdallah

address: Joseph Stelzmann Str. 26

department: Medical Faculty

institute: University of Cologne

summary: Concerning the roles of LCN2 in chronic inflammation and fibrosis, we investigated chronic liver inflammation and fibrosis using repeated carbon tetrachloride (CCl4) in mineral oil injection. We found that mice treated with the mineral oil vehicle alone also showed liver inflammation and more severe in wild-type mice compared to lipocalin 2 null mice.

overall_design: Wild type and lipocalin 2 null mice were subjected to intraperitoneal injection twice weekly for 8 weeks with carbon tetrachloride (CCl4) in mineral oil or mineral oil vehicle, liver leukocyte isolation, immunohistochemistry (IHC) of leukocyte surface antigens, and RNA sequencing (RNA-Seq)

first: Erawan

last: Borkham-Kamphorst

first: Ute

last: Haas

first: Manuela

last: Pinoé-Schmidt

first: Ralf

last: Weiskirchen


title: FAM72A promotes mutagenic repair during antibody maturation

city: Toronto

first: Jason

last: Moffat

state: ON

address: 686 Bay Street, Room 13.9705

department: Molecular Genetics

country: Canada

institute: University of Toronto

summary: Genome-wide CRISPR screen in murine CH12F3-2 cell line to identify novel factors involved in antibody class switch recombination. FAM72A, a protein with no ascribed function, was identified in this screen as a major determinant during mutagenic uracil repair by antagonizing base excision repair protein UNG2.

overall_design: CH12F3-2 cells were transduced with the CRISPR gRNA library, followed by C.I.T cocktail treatment to allow antibody class switching to occur from IgM to IgA. Cells were stained for surface IgA expression and FACS sorting was done to separate IgA+ population from IgA- population. gRNA representation was compared from IgA+ or IgA- population to the unsorted to determine genes involved in CSR.

last: Feng

first: Alberto

last: Martin


title: The Hsp90 cochaperone TTT promotes cotranslational maturation of PIKK kinases prior to complex assembly

city: Montpellier

first: Dominique

last: Helmlinger

address: 1919 route de Mende

department: CRBM

country: France

institute: CNRS

summary: PIKKs are a family of kinases that control fundamental processes, including cell growth, DNA damage repair, and gene expression. Although their regulation and activities are well characterized, little is known about how PIKKs fold and assemble into active complexes. Previous work identified an Hsp90 cochaperone, the TTT complex, which specifically stabilizes PIKKs. Here we describe a mechanism by which TTT promotes their de novo maturation in fission yeast. We show that TTT recognizes newly synthesized PIKKs during translation. Although PIKKs form multimeric complexes, we found that they do not engage in cotranslational assembly with their partners. Rather, we accumulated evidence that TTT protects nascent PIKK polypeptides from misfolding and degradation and that PIKKs acquire their native state after translation is terminated. Thus, PIKK maturation and assembly are temporally segregated, suggesting that the biogenesis of large complexes requires both dedicated chaperones and cotranslational interactions between subunits.

overall_design: mRNA profiles from S. pombe strains of various genotypes were generated by deep sequencing, in triplicate, in one lane of an Illumina HiSeq 4000, with 1x 50bp single reads.

first: Dom


title: Role of 53BP1 in End Protection and DNA synthesis at DNA breaks [SAR-seq]

city: Shanghai

first: Wei

last: Wu

state: Shanghai

address: 320 yueyang road

department: Center for Excellence in Molecular Cell Science

summary: Double-strand break (DSB) repair choice is greatly influenced by the initial processing of DNA ends. 53BP1 limits the formation of recombinogenic single strand DNA (ssDNA) in BRCA1-deficient cells leading to defects in homologous recombination (HR). However, the exact mechanisms by which 53BP1 inhibits DSB resection remain unclear. Previous studies have identified two potential pathways: protection against exonucleases presumably through the Shieldin (SHLD) complex binding to ssDNA, and localized DNA synthesis through the (CTC1-STN1-TEN1) CST and DNA polymerase alpha (Polα) to counteract resection. We present evidence here that 53BP1-mediated exonuclease protection plays a more significant role than CST/Polα synthesis in countering hyper-resection at DSBs in G1 phase. Using a combinatorial approach of END-seq, SAR-seq, and RPA ChIP-seq, we directly assessed the extent of resection, DNA synthesis, and ssDNA, respectively, at AsiSI-induced DSBs. We show that in the presence of 53BP1, Polα-dependent DNA synthesis reduces the fraction of resected DSBs and the resection lengths. However, in the absence of 53BP1, Polα activity is sustained on ssDNA yet does not substantially counter resection. In contrast, Exo1 nuclease activity is essential for hyperresection in the absence of 53BP1. Thus, 53BP1 inhibits resection mainly through end-protection rather than by promoting fill-in.

overall_design: We used a combinatorial approach of END-seq, SAR-seq, and RPA SSDS to characterize at nucleotide resolution the in vivo molecular events that regulate DSB resection.

first: Jacob

last: Paiano

first: Nicholas

last: Zolnerowich

first: Raphael

last: Pavani

first: Chen

last: Wang

first: Hongzhi

last: Li

first: Li

last: Zheng

first: Binghui

last: Shen

first: Barry

last: Sleckman

first: Bo-Ruei

first: André

last: Nussenzweig


title: Inhibition of CBP synergizes with the RNA-dependent mechanisms of azacitidine by limiting protein synthesis

city: Badalona

first: Jeannine

last: Diesch

state: Barcelona

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

department: Research

institute: Josep Carreras Leukaemia Research Institute

summary: The nucleotide analogue azacitidine interferes with RNA and DNA metabolism and is currently the best treatment option for a subset of patients with high-risk myelodysplastic syndromes. However, only half of treated patients respond and almost all patients that initially respond eventually relapse. We performed an optimized loss-of-function shRNA screen in combination with azacitidine treatment in an MDS-derived acute myeloid leukemia cell line to identify chromatin regulators affecting drug response. We identified CBP, as a major regulator of azacitidine sensitivity. Compounds inhibiting the enzymatic activity of CBP synergistically reduced viability of MDS-derived AML cell lines when combined with AZA. Surprisingly, this effect was specific for the RNA-dependent functions of AZA and not observed with the related compound decitabine that is limited to DNA incorporation. The identification of immediate target genes suggested that the effect of CBP inhibition is mediated by downregulation of genes encoding the translational machinery, which could be confirmed in proteomic analysis of nascent proteins. Furthermore, proteins most affected by CBP inhibition include key drivers of cell cycle progression.

overall_design:
Nascent mRNA transcripts in the sAML cell line MOLM-13 in response to C646 treatment (2h, 10µM) identified by SLAM-seq
MOLM-13 cells were either treated with 10 µM C646 or DMSO (3 replicates each) for 1 hour, labelled with 100 µM 4sU for another hour and SLAM-seq performed using the SLAMseq Anabolic Kinetics Kit (Lexogen Gmbh, Austria) followed by library preparation using the QuantSeq 3’mRNA-Seq for Illumina (FWD) and PCR Add-on Kit for Illumina (Lexogen Gmbh, Austria) according to the manufacturer’s instructions. The subsequent data analysis was performed using the software SLAM-Dunk and the R package DEseq2.

first: Marcus

last: Buschbeck


title: Role of 53BP1 in End Protection and DNA synthesis at DNA breaks [END-seq]

institute: Center for Excellence in Molecular Cell Science

overall_design:
We used a combinatorial approach of END-seq, SAR-seq, and RPA SSDS to characterize at nucleotide resolution the in vivo molecular events that regulate DSB resection.
Please note that the 'PreB-Lig4-18hrs-AsisI-ENDseq-control-for-HU' sample was used as control for both HU and Lig4-/-53BP1-/-.

last: Chen


summary: The nucleotide analogue azacitidine interferes with RNA and DNA metabolism and is currently the best treatment option for a subset of patients with high-risk myelodysplastic syndromes. However, only half of treated patients respond and almost all patients that initially respond eventually relapse. We performed an optimized loss-of-function shRNA screen in combination with azacitidine treatment in an MDS-derived acute myeloid leukemia cell line to identify chromatin regulators affecting drug response. We identified CBP, as a major regulator of azacitidine sensitivity. Compounds inhibiting the enzymatic activity of CBP synergistically reduced viability of MDS-derived AML cell lines when combined with AZA. Surprisingly, this effect was specific for the RNA-dependent functions of AZA and not observed with the related compound decitabine that is limited to DNA incorporation. The identification of immediate target genes suggested that the effect of CBP inhibition is mediated by downregulation of genes encoding the translational machinery, which could be confirmed in proteomic analysis of nascent proteins. Furthermore, proteins most affected by CBP inhibition include key drivers of cell cycle progression.

overall_design: SKK-1 cells were transduced with the Epi9 shRNA library, selected with puromycin and half of the cells treated with 0.25µM azacitidine every two days for two weeks. 14 days after initiation of the treatment the cells were harvested, the gDNA extracted and sequenced using a custom primer recognizing the shRNA guide strands


title: Induced Pluripotent Stem Cell Derived Liver Model for the Study of PNPLA3 Associated Non-Alcoholic Fatty Liver Disease

first: Samantha

middle: Grace

last: Tilson

state: Maryland

address: 10 Center Dr., Building 10, Room 10S257

department: Liver Diseases Branch

institute: NIH, NIDDK

summary: In order to identify molecular mechanisms associated with PNPLA3 associated non-alcoholic fatty liver disease, we ran RNA sequencing on human induced pluripotent stem cell-derived hepatocyte like cells that were homozygous for the wild type PNPLA3 allele, homozygous for the risk PNPLA3 allele (I148M), or had a complete knock-out of the PNPLA3 gene. The cells were then treated with either oleic acid or palmitic acid to induce a steatotic or a lipotoxic phenotype, respectively. The cells were treated with control medium or medium supplemented with 0.25 mM oleic acid or palmitic acid for 24 hours before samples were collected for RNA sequencing.

overall_design: mRNA profiles of hepatocyte like cells of 3 PNPLA3 genotypes (WT, I148M, and KO) treated with 3 lipid treatments (Control, OA, and PA).


title: Male biased islet b cell dysfunction is caused by the MODY MAFA S64F variant by inducing premature aging and senescence

city: Nashville

first: Xin

last: Tong

state: TN

address: 2213 Garland Ave

institute: Vanderbilt University

summary: A heterozygous missense mutation producing a variant of the islet β-cell-enriched MAFA transcription factor (p.Ser64Phe (S64F)) was identified in patients who developed adult-onset, β-cell dysfunction (diabetes or insulinomatosis), with men more prone to diabetes than women. This mutation engenders increased stability to the normally unstable MAFA protein. To obtain insight into how this variant impacts β cell function, we developed a mouse model expressing S64F MafA and found sex-dependent phenotypes, with heterozygous mutant males (MafAS64F/+) displaying impaired glucose tolerance while females were slightly hypoglycemic with improved blood glucose clearance. Only MafAS64F/+ males showed transiently higher MafA protein levels preceding the onset of glucose intolerance and sex-dependent, differential expression of genes involved in Ca2+ signaling, DNA damage, aging, and senescence. Functional changes in islet Ca2+ handling and signs of islet aging and senescence processes were uniquely observed in male animals. In addition, MAFAS64F production in male human β cells accelerated cellular senescence and increased production of senescence-associated secretory proteins compared to cells expressing MAFAWT. Together, these results implicate a conserved mechanism of accelerated islet aging and senescence in promoting diabetes in MAFAS64F carriers in a sex-dependent manner.

overall_design: Bulk RNAseq was performed in 4 groups of isolated mouse islets: Male Wildtype, Male MafA-S64F heterozygous, Female Wildtype and Female MafA-S64F heterozygous. Each group includes 4-5 replicates.

first: Emily

last: Walker

first: Jeeyeon

last: Cha

first: Roland

last: Stein


title: RNA-sequencing of tumor-infiltrated T cells in colorectal tumors

city: San Francisco

first: Bushra

last: Samad

state: CA

address: 505 Parnassus Ave

institute: USCF Immunoprofiler

summary: The goal of this study is to identify differentially expressed genes in colorectal tumor samples enriched for exhausted T cells. CD38 expression was used as a representative marker for T cell exhaustion. The comparison was made between CD38high (top 33rd percentile of all samples) and CD38low (bottom 33rd percentile of all samples) samples.

overall_design: Tumor samples for the Immunoprofiler was transported from various cancer operating rooms (ORs) as well as from outpatient clinics. All patients consented by the UCSF IPI clinical coordinator group for tissue collection under a UCSF IRB approved protocol (UCSF IRB# 20-31740). Tumor or metastatic tissue was thoroughly chopped with surgical scissors and transferred to GentleMACs C Tubes (Miltenyi Biotec) containing 20 uL/mL Liberase TL (5 mg/ml, Roche) and 50 U/ml DNAse I (Roche) in RPMI 1640 per 0.3 g tissue. GentleMACs C Tubes were then installed onto the GentleMACs Octo Dissociator (Miltenyi Biotec) and incubated for 45min according to the manufacturer’s instructions. Samples were then quenched with 15 mL of sort buffer (PBS/2% FCS/2mM EDTA), filtered through 100 um filters and spun down. Red blood cell lysis was performed with 175 mM ammonium chloride if needed.   Cells were then incubated with Human FcX (Biolegend) to prevent non-specific antibody binding. Cells were then washed in DPBS and incubated with Zombie Aqua Fixable Viability Dye (Thermo). Following viability dye, cells were washed with sort buffer and incubated with cell surface antibodies mix diluted in the BV stain buffer (BD Biosciences) following manufacturer instruction for 30 minutes on ice in the dark and subsequently fixed in either Fixation Buffer (BD Biosciences) or in Foxp3/Transcription Factor Staining Buffer Set (eBioscience) if intracellular staining was required.

first: Alexis

middle: J

last: Combes

first: Matthew

last: Krummel


title: Transcriptome analysis of CD14+ monocytes from SSc patients

city: Zürich

first: Peter

last: Leary

address: Winterthurerstrasse 190

department: Functional Genomics Center Zürich

institute: ETH Zürich

summary:
Purpose: To compare transcriptome profile of CD14+ blood monocytes from systemic sclerosis (SSc) patients and healthy controls.
Methods: CD14+ monocytes were isolated from peripheral blood of lcSSc (n=5, age=54.4±6.7), dcSSc patients (n=5, age=51.8±7.2) and age- and sex-matched healthy controls (HC) (n=5, age=50.8±9.7). Total RNA was isolated and polyA libraries were prepared using TruSeq Stranded mRNA kit. Next Generation Sequencing was performed using Illumina HiSeq 4000 platform. Differentially expressed genes were computed using DESeq2 algorithm.
Results: We detected 1440 differentially expressed genes between dcSSc vs HC and 225 between lcSSc and HC respectively (p≤0.01; log2 ratio≥0.5, Fig 1). Among those, in dcSSc 1076 were upregulated (e.g. MMP9, IL1R2, FLT3, MIF, TLR9) and 364 were downregulated (e.g. TGFBR1, CD44, CD244, HLA-DRA, HLA-G). In lcSSc 160 transcripts were upregulated (e.g. CCL2, WNT5B, MMP17) and 65 were downregulated (e.g. KLF11, IRAK2). We identified 123 commonly deregulated genes between SSc subgroups (e.g. CCL3, CD14, IL27, MMP17). Principal component analysis showed close clustering within SSc subgroups and clear separation from healthy controls. Pathway analysis revealed alterations in several biological processes important in fibrogenesis including antigen presentation, MIF-induced immune responses, TGF-β, NOTCH and WNT signalling pathways. qPCR analysis further confirmed differences in gene expression on mRNA level (n HC=8, n SSc=25, p≤0.05).
Conclusions: To our knowledge, this is the first global transcriptome analysis of peripheral blood CD14+ monocytes in SSc. Our results suggest a primary activation of monocytes in peripheral blood, which might be further translated into novel cellular biomarker of the disease and potentially used for distinguishing between responders and non-responders to a novel treatment in future clinical trials.

overall_design: Examination of CD14+ monocytes isolated from lcSSc, dcSSc and healthy controls.

first: Michal

last: Rudnik

first: Gabriela

last: Kania


title: Small intestinal immune cell distributions and transcriptomes in mice lacking H2-Ab1 on intestinal epithelial cells.

status: Public on Sep 09 2021

city: Salt Lake City

first: W. Zac

last: Stephens

state: UT

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

department: Pathology

institute: University of Utah

summary: Intestinal epithelial cells (IEC) express large amounts of major histocompatibility complex II (MHCII) molecules. Despite this long-appreciated observation, the function of epithelial MHCII-mediated signaling on gut homeostasis remains enigmatic. As IECs serve as the primary cellular barrier between intestinal microbes and underlying host immune cells, we reasoned that IEC-intrinsic antigen presentation may play a role in tolerogenic responses towards the microbiota. Mice with an IEC-intrinsic defect in MHCII expression (IECΔMHCII) develop elevated T cell-dependent IgA responses, but a reduction in microbiota responsive regulatory T cells (Tregs). Despite elevated IgA levels in IECΔMHCII mice, immunoglobulin repertoires exhibit less selection and IgA has reduced reactivity to the microbiota, which is associated with increased inter-individual variability in microbiota composition. Consistent with reductions in microbiota-responsive Tregs, IECΔMHCII mice develop worsened colitis. A striking difference observed in the absence of IEC-MHCII is that while transcription of MHCII is similar, underlying mononuclear phagocytes had reduced surface MHCII. Macrophages were found to be capable of acquiring MHCII molecules from IECs, and macrophages isolated from IECΔMHCII mice had decreased capacity to stimulate Treg development. Thus, epithelial-myeloid interactions govern development of adaptive responses to microbial antigens within the gastrointestinal tract.

overall_design: Ileum lamina propria cells or Peyer's patches from 4 adult female mice each of WT (H2-Ab1 fl/fl; Villin:Cre-) and IECΔMHCII (H2-Ab1 fl/fl; Villin:Cre-) were isolated and MACS enriched for CD45+ cells, then pooled by genotypes for each tissue.

first: W

middle: Z


title: Peyer's Patch BCR repertoire analysis in mice lacking H2-Ab1 on intestinal epithelial cells

overall_design: Peyer's patches from 4 adult female mice each of WT (H2-Ab1 fl/fl; Villin:Cre-) and IECΔMHCII (H2-Ab1 fl/fl; Villin:Cre-) were isolated and MACS enriched for CD45+ cells, then pooled by genotypes.


title: CD45- EpCAM+ Intestinal Epithelial Cell Transcriptomes

overall_design: Sorted IECs from 5 WT (H2-Ab1 fl/fl; Villin:Cre-) and 5 IECΔMHCII (H2-Ab1 fl/fl; Villin:Cre-) mice were sorted and RNA extracted in bulk from each animal for RNAseq.


title: Distal Ileum Transcriptomes of Mice with DSS-colitis

overall_design: Total RNA from 2 cm of distal ileum tissue from 6 WT (H2-Ab1 fl/fl; Villin:Cre-) and 5 IECΔMHCII (H2-Ab1 fl/fl; Villin:Cre-) mice.


title: Repeated exposure with short-term behavioral stress resolves pre-existing stress-induced depressive-like behavior in mice

city: Seoul

first: Eun-Hwa

last: Lee

address: Science building B214, 52 Ewhayeodae-gil, Seodaemun-gu

country: South Korea

institute: Ewha Womans University

summary: Chronic stress induces adaptive changes in the brain via the cumulative action of glucocorticoids, which is associated with mood disorders. Here we show that repeated daily five-minute restraint resolves pre-existing stress-induced depressive-like behavior in mice. Repeated injection of glucocorticoids in low doses mimics the anti-depressive effects of short-term stress. Repeated exposure to short-term stress and injection of glucocorticoids activate neurons in largely overlapping regions of the brain, as shown by c-Fos staining, and reverse distinct stress-induced gene expression profiles. Chemogenetic inhibition of neurons in the prelimbic cortex projecting to the nucleus accumbens, basolateral amygdala, or bed nucleus of the stria terminalis results in anti-depressive effects similarly to short-term stress exposure, while only inhibition of neurons in the prelimbic cortex projecting to the bed nucleus of the stria terminalis rescues glucocorticoid release. In summary, we show that short-term stress can reverse adaptively altered stress gains and resolve stress-induced depressive-like behavior.

overall_design: Two independent sets of control (CON), CRST, and CRST+RS5 groups were prepared. Mice were subjected to CRST treatment followed by RS5, and then sacrificed on post-stress day 22. Total RNA was extracted from the pooled PL tissue of 6 – 7 animals in each group. RNA extraction and hybridization on Illumina microarrays.


title: Homeoprotein SIX1 compromises anti-tumor immunity through TGF-β-mediated regulation of collagens

city: Suzhou

first: Wancheng

last: Liu

state: jiangsu

address: 100 Chongwen Road, Industrial Park

institute: Chinese Academy of Medical Sciences and Peking Union Medical College

summary:
Purpose: The tumor microenvironment (TME), such as non-immune-infiltrated “cold” tumors, has been associated with immune suppression. However, the complex mechanisms regulating tumor immunogenicity have not been clearly elucidated. In this study, we investigated whether Transcription factor sine oculis homeobox 1 (SIX1) expression in the cancer cells regulated anti-tumor immunity by reshaping TME.
Methods: The expression of SIX1 in human tumor tissues was analyzed based on TCGA data. Six1 knockout murine cell lines were generated to analyze the SIX1-associated tumor progression and anti-tumor immune response in tumor xenograft models. Immune cells and cytokines in TME were quantified by immunofluorescence, flow cytometry, quantitative reverse transcription-PCR, and ELISPOT assay. SIX1-associated differentially expressed genes were investigated in cancer cells and tissues with RNA-Seq. At the end, Correlation of SIX1 with TGFBR2 expression was assessed by Western blot and Dual-luciferase reporter assay.
Results:SIX1 was highly upregulated in human tumor tissues, while rarely expressed in matched normal tissues. Deletion of the Six1 in cancer cells inhibited tumor growth, depending on adaptive immunity. Moreover, Six1 deficiency significantly enhanced CD8+ T cells infiltration, leading to eradication of poorly immunogenic tumors and maintaining a long-term protection from tumor re-challenge. Mechanistically, SIX1 as a transcription factor upregulated TGFBR2 expression, which induced collagen genes expression via the Smad2/3 phosphorylation, increased collagens deposition in TME and hampered CD8+ T cells infiltration.
Conclusions: Six1 deletion in cancer cells improved tumor immunogenicity, leading to tumor destruction by enhancing anti-tumor immune responses. Our observations uncovered a crucial role of SIX1 on remodulating tumor immunogenicity and demonstrated a proof of concept for targeting SIX1 in cancer immunotherapy.

overall_design: mRNA profiles of MCA205 wild type (WT) and SIX1-/- cell lines or tumor tissue.

first: M

last: Gao

first: L

last: Li

first: H

last: Yang

first: G

last: Cheng


title: Single cell RNA sequencing of naive and memory B cells

first: Ryan

last: Smolkin

address: 408 E 69th St

institute: MSKCC

summary: The goal of this study is to compare the transcriptomes of memory and naïve B cells, and provide a resource for single cell analysis of murine B cells during influenza infection.

overall_design: Standard droplet-based single cell RNA sequencing of naïve and memory B cells done via 10X and analyzed with Seurat.

first: William

last: Yewdell

first: Kalina

last: Belcheva

first: Jayanta

last: Chaudhuri


title: Assessment of the platelet transcriptome in patients with COVID-19 versus controls

city: New York City

first: MacIntosh

middle: Grant

last: Cornwell

department: School of Medicine

institute: New York University

summary: To understand how COVID-19 alters the platelet transcriptome.

overall_design: A cohort of eight hospitalized COVID-19 patients (n=8) were recruited from NYU Langone Health between May 11-21, 2020. SARS-CoV-2 infection was confirmed by RT-PCR, in accordance with current standards. All COVID-19 patients and , age-, and sex-matched control donors were recruited under study protocols approved by the NYU Langone Health Institutional Review Board. Each study participant or their Legal Authorized Representative gave written informed consent for study enrollment in accordance with the Declaration of Helsinki. For COVID-19 patients, enrollment criteria included age greater than 18, hospital admission, positive SARS-CoV-2 testing, and informed consent. COVID-19 patients were monitored until discharge or death.

first: Tessa

last: Barrett

first: Jeffrey

last: Berger


title: Biofilm Formation and Virulence of Shigella flexneri is Modulated by pH

city: Cleveland

first: Thaddeus

last: Stappnbeck

state: OH

address: 9500 Euclid Ave

department: Inflammation and Immunity

institute: Cleveland Clinic, Lerner Research Institute

summary: In this study, we probed factors that could influence Shigella pathogenesis. We show that in basic pH conditions, deoxycholate-induced biofilm formation and virulence of Shigella are attenuated. We utilized RNA-sequencing to investigate pathways enriched in bacterial cells in biofilms.

overall_design: Transcriptomic profiles of Shigella in different conditions of growth.

first: I-Ling

last: Chiang

first: Yi

last: Wang

first: Satoru

last: Fujii

first: Brian

last: Muegge

first: Qiuhe

last: Lu

first: Phillip

middle: I

last: Tarr

last: Stappenbeck


title: The RNA helicase Ddx21 controls Vegfc-driven developmental lymphangiogenesis by balancing endothelial cell ribosome biogenesis and p53-p21 signalling

city: Melbourne

first: Tyrone

last: Chen

state: VIC

address: 305 Grattan St

department: Bioinformatics Core (RRID: SCR_025901); Organogenesis and Cancer Program

institute: Peter MacCallum Cancer Centre

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

sample_organism:
Danio rerio
Homo sapiens

platform_organism:
Danio rerio
Homo sapiens

overall_design: Refer to individual Series


title: The RNA helicase Ddx21 controls Vegfc-driven developmental lymphangiogenesis by balancing endothelial cell ribosome biogenesis and p53-p21 signalling [human]

summary: The development of a differentiated and functional vasculature requires coordinated control of cell fate specification, lineage differentiation and vascular network growth. Cellular proliferation is spatiotemporally regulated in developing vessel networks but how this is achieved and differentially controlled in specific lineages is unknown. Using a zebrafish forward genetic screen for mutants that form blood vessels but fail to form lymphatic vessels, we uncovered a mutant for the RNA helicase Ddx21. Ddx21 cell-autonomously regulates the early development of lymphatic endothelial cells. Ddx21 is essential for Vegfc-Vegfr3 driven endothelial cell proliferation. Ddx21 is an established regulator of ribosomal RNA transcription and in the absence of Ddx21, mutant lymphatic endothelial cells show reduced ribosome biogenesis. Ultimately, loss of Ddx21 leads to a p53-p21 dependent cell cycle arrest that blocks embryonic lymphangiogenesis. Thus, the RNA helicase Ddx21 coordinates the endothelial cell proliferative response to Vegfc-Vegfr3 signalling by balancing ribosome biogenesis and p53-p21 signalling. This mechanism may have therapeutic potential in diseases of excessive lymphangiogenesis such as in cancer metastasis or lymphatic malformation.

overall_design: HUVEC RNA-seq data on untreated control cells and DDX21-siRNA knockdown cells using two constructs

first: K

last: Koltowska


title: Microvascular Endothelial Cell RNA-Seq: Treatment with Platelet Releasate from COVID-19 & Control Donors

summary: Assessment of the endothelial transcriptome following exposure to platelet releasated factors from COVID-19 and controls.

overall_design: Human cardiac microvascular endothelial cells exposed to platelet releasate (7 controls, 7 COVID-19) originating from patients with COVID-19 or controls.


title: Hairless regulates heterochromatin maintenance and muscle stem cell function as a histone demethylase antagonist (ChIPseq, ATACseq)

city: Palo Alto

first: Ling

last: Liu

address: 300 Pasteur Drive

institute: Stanford University

summary: Skeletal muscle possesses remarkable regenerative ability owing to the resident muscle stem cells (MuSCs). A prominent feature of quiescent MuSCs is a high content of heterochromatin. However, little is known about the mechanisms by which heterochromatin is maintained in MuSCs. We found that the mammalian Hairless (Hr) gene is expressed in quiescent MuSCs. Using a mouse model in which Hr can be specifically ablated in MuSCs, we demonstrate that Hr expression is critical for MuSC function and muscle regeneration. We show that Hr is a histone demethylase antagonist that preserves Histone 3 Lysine 9 trimethylation (H3K9me3) and maintains heterochromatin structure. Loss of Hr results in reduced H3K9me3 levels, reduced heterochromatin, increased susceptibility of MuSCs to genotoxic stress, and the accumulation of DNA damage. Our study not only elucidates the molecular mechanism by which Hr regulates histone demethylases, but also demonstrates the importance of heterochromatin maintenance in stem cell function.

overall_design: Skeletal muscle stem cells (MuSCs) were isolated from conditional Hairless knockout and control mice by FACS and processed for ChIP-seq using H3K9me3-specific antibodies.

first: Thomas

last: Rando


title: Hairless regulates heterochromatin maintenance and muscle stem cell function as a histone demethylase antagonist [RNA-seq]

overall_design: Skeletal muscle stem cells (MuSCs) were isolated from conditional Hairless knockout and control mice by FACS and processed for RNA-seq.


title: Transcriptomic analysis of Spiroplamsa infection induced changes in gene expression in reproductive tissues of female and male Glossina fuscipes fuscipes

status: Public on Sep 08 2021

first: Jae Hak

last: Son

state: CT

address: 60 College St

institute: Yale School of Public Health

summary: We collected female and male Glossina fuscipes fuscipes from the field (Uganda) and determined the Spiroplasma infections status of each individual. We used RNA-seq to investigate the effects of Spiroplasma on the male and female gene expression in the reproductive tissues. We observed that Spiroplasma infection induces sex-biased expressional changes in genes that encode proteins critical for tsetse`s reproductive success.

overall_design: The following Spiroplasma infection status and Glossina fuscipes fuscipes sex combinations were analyzed: 1) Spi- female;  2) Spi+ female; 3) Spi- male; 4) Spi+ male. Three female or male reproductive tracks were pooled into distinct biological replicates, creating two or three biological replicates.

first: Daniela

last: Schneider

middle: L

last: Weiss

first: Serap

last: Aksoy