title: Identifying non-genetic determinants of malignant clonal fitness at single cell resolution (primary leukaemia scRNAseq)

last: Dawson

overall_design: Single-cell RNA-seq of MLL-AF9 leukaemia cells with SPLINTR barcodes


title: Identifying non-genetic determinants of malignant clonal fitness at single cell resolution (NSG vs. BL6 barcode-seq)

status: Public on Nov 08 2021

city: Melbourne

first: Mark

state: VIC

address: 305 Grattan Street

country: Australia

institute: Peter MacCallum Cancer Centre

summary: All cancers emerge following a period of clonal selection and subsequent clonal expansion. Whilst the evolutionary principles imparted by genetic intra-tumour heterogeneity (ITH) are becoming increasingly clear, little is known about the non-genetic mechanisms that contribute to ITH and malignant clonal fitness. Using SPLINTR, a synthetic expressed barcoding strategy, in three clinically relevant mouse models of acute myeloid leukaemia (AML) we find that malignant clonal dominance is a stable and heritable property that is facilitated by the repression of antigen presentation and the increased expression of Slpi, a leukocyte protease inhibitor that has not previously been characterised in AML. Increased transcriptional heterogeneity is a consistent feature enabling clonal fitness in diverse tissue / immune microenvironments and in the context of clonal competition between genetically distinct clones within a uniform microenvironment. Compared to extramedullary sites, leukaemia initiating capacity is most enriched in malignant cells resident within the bone marrow microenvironment and leukaemia stem cells (LSC), like normal haematopoietic stem cells, display heritable clone-intrinsic properties of high, and low clonal output that contribute to the overall tumour mass. Finally, we demonstrate that clonal output does not dictate sensitivity to chemotherapy and both high and low output LSC clones retain marked cellular plasticity enabling them to survive potent therapeutic challenge and persist as minimal residual disease. Together these data provide fundamental insights into the non-genetic transcriptional processes that underpin malignant clonal fitness which may inform future therapeutic strategies.

overall_design: DNA barcode-seq of MLL-AF9 leukaemia cells transplanted into Ptprca or NSG mice

middle: A

first: Katie

last: Fennell

first: Dane

last: Vassiliadis


title: Identifying non-genetic determinants of malignant clonal fitness at single cell resolution (primary leukaemia barcode-seq)

sample_organism:
Mus musculus
synthetic construct

platform_organism:
Mus musculus
synthetic construct

overall_design: DNA barcode-seq of MLL-AF9 leukaemia cells


title: Individual Human Cortical Progenitors Can Produce Excitatory and Inhibitory Neurons

status: Public on Nov 06 2021

city: San Francisco

first: Tomasz

last: Nowakowski

state: Ca

country: USA

institute: UCSF

type: Expression profiling by high throughput sequencing

summary: The cerebral cortex is a cellularly-complex structure comprised of a rich diversity of neuronal and glial cell types. Cortical neurons can be broadly categorized into two classes—glutamatergic excitatory neurons and GABAergic inhibitory interneurons. Previous developmental studies in rodents have led to the prevailing model that while excitatory neurons are born from progenitors located in the cortex, cortical interneurons are born from a separate population of progenitors located outside of the developing cortex in the ganglionic eminences1-5. However, the developmental potential of human cortical progenitors has not been thoroughly explored. Here we show that in addition to excitatory neurons and glia, human cortical progenitors are also capable of producing GABAergic neurons with the transcriptional characteristics and morphologies of cortical interneurons. By developing a cellular barcoding tool called “ScRNAseq-compatible Tracer for Identifying Clonal Relationships” (STICR), we were able to perform clonal lineage tracing of 1912 primary human cortical progenitors from six specimens and capture both the transcriptional identities and clonal relationships of their resulting progeny. A subpopulation of cortically-born GABAergic neurons were transcriptionally similar to cortical interneurons born from the caudal ganglionic eminence and these cells were frequently related to excitatory neurons and glia.  Thus, our results demonstrate that individual human cortical progenitors can generate both excitatory neurons and cortical interneurons, providing a new framework for understanding the origins of neuronal diversity in the human cortex.

overall_design: Sequencing of Synthetic Plasmid and Lentiviral STICR libraries for quantitative analysis

first: Ryan

middle: N

last: Delgado


title: Identifying non-genetic determinants of malignant clonal fitness at single cell resolution

type: Methylation profiling by high throughput sequencing

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

overall_design: Refer to individual Series


title: Genome wide mapping of DDX41 chromatin binding using a targeted nuclease approach in U2OS cells after expression

status: Public on Nov 17 2021

city: Mainz

first: Petra

last: Beli

address: Ackermannweg 4

country: Germany

institute: Institute of Molecular Biology (IMB)

type: Genome binding/occupancy profiling by high throughput sequencing

summary:
Purpose: Investigation of DDX41 chromatin binding sites in U2OS cells
Method: Expression of DDX41-GFP was induced for 48 hours before crosslinking using 1µg/ml docycycline. Control cells were not induced with doxycline. Cells were mildly crosslinked using 1% FA for 2 min at RT. Concanavalin A-coated beads were activated in Binding Buffer (20 mM Hepes-KOH pH 7.9, 10 mM KCl, 1mM CaCl2, 1 mM MnCl2).1*10^6  U2OS cells were washed twice with Wash Buffer (Hepes-NaOH pH7.5, 150 mM NaCl, 0.5 mM Spermidine, 1 mM protease inhibitor) at room temperature and afterwards immobilized on the activated beads in 1 ml Wash Buffer. Cells were permeabilized with 0.05% digitonin in Wash buffer for 4 min at RT. 1 µg Nanobody-GFP-MNase (in-house protein production, PMID:25362362) binding was performed at 4°C for 30 min in 0.05% digitonin-Wash buffer. After 2x washing, the MNase was activated by adding 3mM CaCl2 on ice for 30 min.  2x Stop Buffer (68 µl 5M NaCl, 40 µl 0.5 M EDTA, 20 µl 0.2 M EGTA, 10 µl 5% digitonin, 5 µl 10mg/ml RnaseA, )) was mixed with the samples to stop the reaction. Chromatin fragments were released by incubating the samples for 20 minutes at 37°C and centrifugation at 16.000×g for 10 minutes at 4°C. Supernatants were incubated at 70°C in the presence of 0.1% SDS and 5 µg proteinase K. Before library preparation, the DNA was recovered by phenol-chloroform extraction
Results: We succesfully mapped DDX41 binding sites on cromatin in U2OS cells. DDX41 preferentially binds in the promoter region of genes.

overall_design: U2OS cells that dox-inducible express GFP-tagged DDX41 protein were either induced or not with 1 µg/ml doxycycline for 48h. Control cells did not express GFP-tagged DDX41. Cells were mildly crosslinked. Nanobody-GFP-Mnase was use to perform  CUT&RUN.

first: Thorsten

last: Mosler

first: Joan

last: Barau

first: Nastasja

last: Kreim


title: Canonical WNT signaling-dependent gating of MYC requires a non-canonical CTCF function at a distal binding site

city: Stockholm

first: Anita

last: Göndör

address: Nobels väg 16, KI Solna Campus, Karolinska Institutet, Box 280

department: MTC

country: Sweden

institute: Karolinska Institutet

summary: Abnormal WNT signaling increases MYC expression in colon cancer cells in part via oncogenic super-enhancer-(OSE)-mediated gating of the active MYC to the nuclear pore in a poorly understood process. We show here that that the principal tenet of the WNT-regulated MYC gating, facilitated nuclear export of the MYC mRNA, converged on a single CTCF binding site (CTCFBS) within the OSE to confer increased growth advantage. To achieve this, the CTCFBS directed in a stepwise manner the WNT-dependent trafficking of the entire OSE to the nuclear pore from intra-nucleoplasmic positions. Once the OSE reached a perinuclear position, triggered by CTCFBS-mediated CCAT1 eRNA activation, its final stretch (<1micrometer) to the nuclear pore required the recruitment of AHCTF1, a key nucleoporin, to the CTCFBS. Thus, a novel WNT-CTCF-eRNA circuit converges on the ability of the OSE to promote pathological cell growth by coordinating the trafficking of MYC within the 3D nuclear architecture.

overall_design: Examination of CTCF binding in D3, E4 and WT. All the cell line is HCT-116. D3 and E4 are two clones of CTCF mutation.


title: The WNT-dependent gating of MYC requires non-canonical functions of a distal CTCF binding site (RNA-Seq II)


title: Diagnosis of Taenia solium infections based on “mail order” RNA-sequencing of single tapeworm egg isolates from stool samples

city: Berlin

first: Markus

last: Schuelke

state: Berlin

address: Augustenburger Platz 1

department: Neuropediatrics

institute: Charite

summary: Combined community health programs aiming at health education, preventive antiparasitic chemotherapy, and vaccination of pigs have proven their potential to regionally reduce and even eliminate Taenia solium infections that are associated with a high risk of neurological disease through ingestion of T. solium eggs. Yet it remains challenging to target T. solium endemic regions precisely or to make exact diagnoses in individual patients. One major reason is that the widely available stool microscopy may identify Taenia ssp. eggs in stool samples as such, but fails to distinguish between invasive (T. solium) and less invasive Taenia (T. saginata, T. asiatica, and T. hydatigena) species. The identification of Taenia ssp. eggs in routine stool samples often prompts a time-consuming and frequently unsuccessful epidemiologic workup in remote villages far away from a diagnostic laboratory. Here we present “mail order” single egg RNA-sequencing, a new method allowing the identification of the exact Taenia ssp. based on a few eggs found in routine diagnostic stool samples. We provide first T. solium transcriptome data, which show extremely high mitochondrial DNA (mtDNA) transcript counts that can be used for subspecies identification. “Mail order” RNA-sequencing can be administered by health personnel equipped with basic laboratory tools such as a microscope, a Bunsen burner, and access to an international post office for shipment of samples to a next generation sequencing facility. Our suggested workflow combines traditional stool microscopy, RNA-extraction from single Taenia eggs with mitochondrial RNA-sequencing, followed by bioinformatic processing with a basic laptop computer. The workflow could help to better target preventive healthcare measures and improve diagnostic specificity in individual patients based on incidental findings of Taenia ssp. eggs in diagnostic laboratories with limited resources.

overall_design:
Sequencing of the transcriptome of single Taenia solium eggs isolated from a fresh stool sample. Sequencing of whole genomic DNA isolated from a proglottid of a Taenia solium tapeworm excreted after treatment with niclosamide.

Grant Information:

Grant ID: EXC-2049-390688087
Title: NeuroCure
Funding Agency: Germany's Excellence Strategy German Research Foundation (DFG)
Recipient of the grant: Markus Schuelke

Grant ID: 01KA1618
Title: CYSTINET-Africa
Funding Agency: German Federal Ministry of Education and Research
Recipient of the grant: Andrea S. Winkler

first: Henrik

last: Sadlowski


title: Gut microbiota analysis of colitic SD rats

city: Hong Kong

first: Pengchao

last: Zhao

address: G10A, RRSSB

department: Biomedical Engineering

country: China

institute: Chinese University of Hong Kong

summary: Fecal samples collected on day 5 from randomly selected colitic SD rats were analyzed for gut microbiota by sequencing the V4 region of the 16S rRNA gene. The orally administered Dex-P-laden NPA2 coacervate (Dex-P/NPA2) significantly restores the diversity of gut microbiota compared with colitic SD rats in the Dex-P/PBS group and the untreated colitic rats (Control).

overall_design: The fresh fecal samples in autoclaved sterile vials were properly packaged with dry ice cooling agents and shipped to BGI Genomics Co., Ltd (Hong Kong) for microbiota analyses.

first: Xianfeng

last: Xia


title: CXCR3 regulates stem and proliferative CD8+ T cells during chronic infection by promoting interactions with DCs in splenic bridging channels

status: Public on Nov 05 2021

city: Berkeley

first: Zoë

last: Steier

state: CA

address: 378 Stanley Hall

institute: UC Berkeley

summary: Production of armed effector CD8+ T cells during persistent infection requires a stable pool of stem-like cells that can give rise to short-lived effector T cells via a proliferative intermediate population. In chronic infection models marked by T cell exhaustion, this process can be transiently induced by check-point blockade, but it occurs spontaneously and continuously in the spleens of mice chronically infected with the protozoan intracellular parasite, Toxoplasma gondii, providing a unique opportunity to examine the steps in the differentiation process. Using this model, we observed distinct locations and gene expression patterns for stem-like memory cells, proliferative intermediate cells, and terminally differentiated effector cells, implying a link between differentiation and discrete anatomical niches in the spleen. Loss of the chemokine receptor CXCR3 on parasite-specific T cells did not impair their white pulp to red pulp migration but reduced their interactions with CXCR3 ligand-producing type 2 conventional DCs in the bridging channels and impaired their transition from memory to intermediate states, leading to a build up of memory T cells localized to the red pulp. These  results demonstrate a critical role for CXCR3 during chronic infection by increasing T cell exposure to differentiation-inducing signals during red pulp migration, providing a dynamic and flexible mechanism for modulating effector differentiation in response to environmental signals.

overall_design: RNA-seq was performed on GRA6-specific CD8+ T cell subsets isolated from four mice chronically infected with T. gondii (biological replicates). T cells were isolated using FACS sorting on infected splenocytes and divided into three functionally distinct subsets using CXCR3 and KLRG1 staining: Memory cells (TMEM: CXCR3+ KLRG1-), Intermediate cells (TINT: CXCR3+ KLRG1+), and Effector cells (TEFF: CXCR3- KLRG1+). One TMEM sample and one TINT sample were removed from the analysis due to low RNA quality, resulting in 10 samples in total.

first: Derek

middle: J

last: Bangs

first: Alexandra

last: Tsitsiklis

first: Shiao

middle: W

last: Chan

first: James

last: Kaminski

first: Aaron

last: Streets

first: Nir

last: Yosef

first: Ellen

last: Robey


title: Analysis of renal protective effects of ARB and SGLT2i at single cell level.

city: New York

first: Zeguo

last: Sun

address: 1468 Madison Avenue

institute: Ichan School of Medicine at Mount Sinai

summary: Angiotensin receptor blockade (ARB) and sodium-glucose co-transporter 2 inhibitor (SGLT2i) have been used as the standard therapy for patients with diabetic kidney disease (DKD). However, how these two drugs possess additive renal protective effects remains unclear. Here, we conducted single cell RNA-sequencing to profile the kidney cell transcriptome of db/db mice treated with vehicle, ARB, SGLT2i, or both drugs and db/m mice. We identified 10 distinct clusters of kidney cells with predominant proximal tubular (PT) cells. We found that ARB has more anti-inflammatory and anti-fibrosis effects while SGLT2i affects more mitochondrial function. We also identified a new PT subcluster  which was increased in DKD but reversed by treatments.This new subcluster was also confirmed by Immunostaining of mouse and human kidneys with DKD. Together, our study reveal kidney cell-specific gene signatures in response to ARB and SGLT2i and also identified a new PT subcluster which provides new insight into DKD.

overall_design: Male db/db mice (BKS.Cg-Dock7m+/+Leprdb/Nju) were purchased from the Model Animal Research Center of Nanjing University. Male lean controls db/m mice were purchased from The Jackson Laboratory. We used only male mice here because these mice develop DKD more consistently than female mice. Mice were housed in a specific pathogen-free facility with free access to food and water and a 12:12h night-day cycle. Male db/db mice develop DKD defined by development of albuminuria at 10 weeks of age. The db/db mice were randomly assigned to four groups as follow: treated vehicle control (phosphate-buffered saline, PBS) (n = 8), treated with 3mg/kg/day of dapagliflozin (n = 8), treated with 20mg/kg/day of irbesartan (n = 8), and combination therapy with both dapagliflozin and irbesartan (n = 8). Vehicle and drugs were administra- ted daily by oral gavage for 8 weeks and all mice were sacrificed at age of 18 weeks. All animal studies were performed in accordance with the protocol approved by the Animal Care Committee at the Chongqing Medicine University. BW and fasting BG levels were monitored weekly by glucometer readings. Urine samples were collected weekly.

first: Jinshan

last: Wu


title: RNA-seq of liver tissue of Lifrfl/fl and Lifrfl/fl;Crealb/alb mice

city: Houston

first: Yalan

last: Deng

state: Texas

address: MD Anderson Cancer Center, Zayed Building, Z7.2036, 6565 MD Anderson Blvd

institute: MD Anderson Cancer Center

summary: Loss of LIFR promotes liver tumorigenesis and confers resistance to ferroptosis, which can be targeted by anti-LCN2 therapy.

overall_design: RNA samples were extracted from liver of 3 months old Lifrfl/fl and Lifrfl/fl;Crealb/alb mice

first: Fan

last: Yao

first: Mutian

last: Zhang

first: Li

last: Ma


title: PRC1.6 and SETDB1-mediated repression of germline genes in the early embryo precedes DNA methylation-mediated silencing.

city: Vancouver

first: Kentaro

last: Mochizuki

state: BC

address: 2350 Health Sciences Mall

department: Medical Genetics

country: Canada

institute: University of British Columbia

summary: Silencing of a subset of germline genes is dependent upon DNA methylation (DNAme) post-implantation. However, these genes are generally hypomethylated in the ICM, implicating alternative silencing pathways before implantation. Indeed, in embryonic stem cells (ESCs), an overlapping set of genes, including germline “genome-defence” (GGD) genes, are upregulated following deletion of the H3K9 methyltransferase SETDB1 or subunits of the non-canonical PRC1 complex PRC1.6. Here, we show that in pre-implantation embryos and naïve ESCs (nESCs), hypomethylated promoters of germline genes bound by the PRC1.6 DNA-binding subunits MGA/MAX/E2F6 are enriched for RING1B-dependent H2AK119ub1 and H3K9me3. Accordingly, silencing of these genes in nESCs shows a greater dependence on PRC1.6 than DNAme. In contrast, GGD genes are hypermethylated in epiblast-like cells and their silencing is dependent upon SETDB1, PRC1.6/RING1B and DNAme, with H3K9me3 and DNAme establishment dependent upon MGA binding. Thus, GGD genes are initially repressed by PRC1.6, with DNAme subsequently engaged in post-implantation embryos.

overall_design: ChIP-seq from E9.5 PGCs and WT nESCs, EpiLCs and d4 PGCLCs, and Mga dHLH, Setdb1 cKO, Pcgf6 cKO or Dnmt cTKO nESCs and EpiLCs. RNA-seq from WT and Dnmt cTKO, Mga ΔHLH, Setdb1 cKO, Pcgf6 cKO, Ring1b cKO or Setdb1/Ring1b cDKO nESCs and EpiLCs.

first: Jafar

last: Sharif

first: Kenjiro

last: Shirane

first: Kousuke

last: Uranishi

last: Bogutz

first: Sanne

last: Janssen

first: Ayumu

last: Suzuki

first: Akihiko

last: Okuda

first: Haruhiko

last: Koseki

first: Matthew

middle: C

last: Lorincz


title: Early transcriptional changes in rabies virus-infected neurons and extended effects on neuronal functions

status: Public on Nov 04 2021

city: Paris

first: Rachel

last: Legendre

address: 28, rue du docteur Roux

department: Research and Resource Center for Scientific Informatics

country: France

institute: Institut Pasteur

summary: Street-strain rabies virus primarily replicates in central nervous system without inducing significant immune response or structural damages on neurons, but the manifested symptoms of rabies indicate inherent neuronal dysfunctions in CNS. To understand the underlying state of rabies virus-infected neurons and find probable mechanisms for the neuronal dysfunction, we performed RNA-Seq at multiple time-points. This dataset provides RNA-Seq results of wild-type and mutant rabies virus-infected neuron transcriptome, with clear differential expressions between conditions. Through comparative analysis of different time-points, we have found that the matrix protein of rabies virus plays an important role in early suppression of host gene expression and maintaining control over immune response and other processes. The signaling pathways previously known to interact with rabies virus were confirmed to be modulated in this dataset, and contribute to neuronal function-associated processes. We have verified the regulation of gene expressions that could impact neuronal functions collectively, and demonstrated in calcium imaging that indeed the oscillation of calcium trace in neurons are influenced by rabies virus infection.

overall_design: Total mRNA profile comparison of non-infected and Tha / Th4M rabies virus-infected neurons at multiple time-points after infection

first: Seonhee

last: Kim

first: Hugo

last: Varet

first: Florence

last: Larrous

first: Regis

last: Grailhe

first: Hervé

last: Bourhy


title: Cell-specific alterations in the activation of the Pitx1 regulatory landscape caused by the loss of a single enhancer (scRNA-seq)

city: Geneva

first: Guillaume

last: Andrey

address: Rue Michel-Servet 1

department: Department of Genetic Medicine and Development

country: Switzerland

institute: University of Geneva

summary: Developmental genes often reply on multiple transcriptional enhancers to achieve correct expression during embryogenesis. These enhancers can have partially redundant activities, thus it is difficult to ascertain the direct impact of the loss of a single one of them as often only mild, if any, phenotypes are seen. In this work we use the testbed locus Pitx1 to characterise the regulatory and cell specific alterations following the loss of one of its enhancers Pen in vivo. We combine single cell transcriptomics and in embryo cell tracing and observe the global decrease perviously observed in deletion of Pen hindlimbs results from an accumulation of low to non-expressing cells that affects more proximal rather than distal limb and leads to a clubfoot phenotype product of a localised heterochrony and loss of irregular connective tissue. We also analyse the contribution of the enhancer to the overall locus structure and activity and we suggest that in some cases redundant enhancers may locally enforce a robust activation of their gene’s regulatory landscape.

overall_design: Whole forelimb and hindlimb buds were prepared for single cell RNA-seq analysis to identify which cell types make up the developing limbs of Mus musculus.

first: Olimpia

last: Bompadre

first: Raquel

last: Rouco


title: Cell-specific alterations in the activation of the Pitx1 regulatory landscape caused by the loss of a single enhancer (ChIP-seq)

overall_design: We used chromatin immunoprecipitation against H3K27ac chromatin marks in Mus musculus hindlimb cells. We analysed Pitx1-GFP postive (total positive, highly-positive and mildly-positive) and negative cells and Pitx1-GFP-ΔPen GFP postive and negative cells from hindlimb tissue to compare changes in activity level at the locus.


title: Cell-specific alterations in the activation of the Pitx1 regulatory landscape caused by the loss of a single enhancer (RNA-seq)

overall_design: We analysed two biological replicates of Pitx1-GFP postive and negative cells and two biological replicates of Pitx1-GFP-ΔPen GFP postive and negative cells from hindlimb tissue to compare the experession differences between Pitx1 expressing and non-expressing cells in the two genetic backgrounds.


title: A Novel Non-Classical Monocyte-derived Macrophage Subset That Provides A Splenic Replication Niche For Intracellular Salmonella [Bulk RNA-Seq I]

city: Rehoboth

first: Dotan

last: Hoffman

address: Herzel St 234, Biological regulation

department: Biological regulation

country: Israel

institute: Weizmann institute of science

summary: Interactions between intracellular bacteria and mononuclear phagocytes give rise to diverse cellular phenotypes that may determine the outcome of infection. Recent advances in single-cell RNA-seq (scRNA-seq) have identified multiple subsets within the mononuclear population, but implications to their function during infection remain unknown. Here, we applied microscopy, flow cytometry and scRNA-seq to survey the mononuclear niche of intracellular Salmonella Typhimurium (S.Tm) during early systemic infection in mice. We describe eclipse-like growth kinetics in the spleen, with a first phase of bacterial control mediated by tissue-resident red-pulp macrophages. A second phase involved extensive bacterial replication within a newly identified macrophage population characterized by CD9 expression. Using Nr4a1e2-/- mice we established that CD9+ macrophages originated from non-classical monocytes (NCM), and NCM-depleted mice were more resistant to S.Tm infection. Our study defines a novel host-pathogen interface, with macrophage subset-specific interactions that determines early infection dynamics and the infection outcome of the entire organism.

overall_design: Splenocytes were isolated from 6 mice 24 hours post infection with Salmonella enterica serovar Typhimurium. RNA was extracted from CD9 Mf and iNOS Mf

first: Yaara

last: Tevet

first: Sebastien

last: Trzebanski

first: Gili

last: Rosenberg

first: Leia

last: Vainman

first: Aryeh

last: Solomon

first: Noa

middle: B

last: Ben-Moshe

first: Shelly

last: Hen-Avivi

first: Roi

last: Avraham


title: Transcriptomics of Kidney Transplant Recipients after Cytomegalovirus DNAemia

city: Los Angeles

first: Richard

middle: Sungho

last: Ahn

address: 611 Charles E. Young Drive East, UCLA, Boyer Hall, 510C

department: Microbiology, Immunology, and Molecular Genetics

institute: University of California Los Angeles

summary: To study the gene expression dynamics in cytomegalovirus (CMV) DNAemic patients after kidney transplantation,  we performed RNA sequencing (RNAseq) on 31 DNAemic patients at 4 timepoints (baseline, 1 week post-DNAemia, 1 month post-DNAemia, and a long-term timepoint 9-18 months post-DNAemia) matched with 31 non-DNAemic patients at 2 timepoints (baseline and long-term).

overall_design: Whole blood samples were obtained from kidney transplant patients that a) developed CMV DNAemia or b) did not develop CMV DNAemia. Baseline, 1 week post-DNAemia, 1 month post-DNAemia, and 9-18 months post transplant samples were collected from CMV DNAemic patients. Baseline and 9-18 months post transplant samples were collected from non-DNAemic patients. All samples were sequenced on the Illumina HiSeq3000 platform.


title: A Novel Non-Classical Monocyte-derived Macrophage Subset That Provides A Splenic Replication Niche For Intracellular Salmonella


title: ADME gene variants in multiple myeloma susceptibility

status: Public on Nov 03 2021

city: Catanzaro

first: Mariamena

last: Arbitrio

address: via Tommaso Campanella, 115

country: Italy

institute: Institute of Biomedical Research and Innovation (IRIB), National Council of Research (CNR)

summary: The metabolism of xenobiotics is regulated by phase I and II enzymes, and by transporters encoded by the absorption, distribution, metabolism, and excretion (ADME) genes. It is known that the activity of these proteins is influenced by the presence of polymorphic variants in the corresponding gene that can account for the inter-individual variability in both xenobiotic response/toxicity and disease predisposition. Exposure to pesticides and toxic substances, many of which are substrates of ADME-associated proteins, has been demonstrated to increase the risk of Multiple Myeloma (MM). To investigate the inter-individual variability of ADME genes as a risk factor for MM risk, we compared DMET Plus genotyping data from 65 MM patients with 59 CEU Hapmap controls (GPL17860).

overall_design: Affymetrix DMET Plus arrays were performed according to the manufacturer's directions on DNA extracted from peripheral blood samples. Association analysis was performed using DMET-Analyzer software.

first: Francesca

last: Scionti

first: Pierosandro

last: Tagliaferri


title: Congenital iRHOM2 deficiency causes ADAM17 dysfunction and environmentally directed immunodysregulatory disease [whole blood]

city: Ar-Rayyan, Doha,

first: Melanie

last: Makhlouf

address: Al Gharrafa Street,

country: Qatar

institute: Sidra Medicine

summary: We report a pleiotropic disease due to loss-of-function mutations in RHBDF2, the gene encoding iRHOM2, in 2 kindreds with recurrent infections in different organs. One patient had recurrent pneumonia but no colon involvement, another had recurrent infectious hemorrhagic colitis but no lung involvement, and the other two experienced recurrent respiratory infections. Loss of iRHOM2, a rhomboid superfamily member that regulates the ADAM17 metalloproteinase, caused defective ADAM17-dependent cleavage and release of cytokines, including TNF and amphiregulin. To understand the diverse clinical phenotypes, we challenged Rhbdf2-/- mice with  Pseudomonas aeruginosa by nasal gavage and observed more severe pneumonia whereas infection with Citrobacter rodentium caused worse inflammatory colitis than wild-type mice. The fecal microbiota in the colitis patient had characteristic oral species that can predispose to colitis. Thus, a new human immunodeficiency arising from iRHOM2 deficiency causes divergent disease phenotypes that can involve the local microbial environment.

overall_design: Whole blood mRNA-seq.

first: Satoshi

last: Kubo

first: Ahmet

last: Ozen

first: Bernice

last: Lo

first: Michael

last: Lenardo


title: Congenital iRHOM2 deficiency causes ADAM17 dysfunction and environmentally directed immunodysregulatory disease

overall_design: Refer to individual Series


title: Expression data from BE2C cells treated with GSK-J4 and KDM6B knockdown

city: Memphis

first: JUN

last: YANG

state: TN

address: 262 Danny Thomas Place

department: Surgery

institute: St Jude Children's Research Hospital

summary:
KDM6B inhibition shows anticancer activity in neuroblastoma models. However, the acting mechanism of KDM6B inhibition remains poorly understood.
We used microarray to detail the global programme of gene expression underlying anticancer activity of KDM6B inhibition in neuroblastoma cells.

overall_design: BE2C cells were treated with GSK-J4 and KDM6B knockdown for RNA extraction and hybridization on Affymetrix microarrays.

first: Jun

last: Yang


title: Genetic determinants of endophytism in the Arabidopsis root mycobiome

city: Cologne

first: Fantin

last: Mesny

address: Carl-von-Linné-Weg 10

department: Plant-Microbes interaction

institute: Max Planck Institute for Plant Breeding Research

summary: Roots of Arabidopsis thaliana do not engage in symbiotic association with mycorrhizal fungi but host taxonomically diverse fungal communities that influence health and disease states. We sequenced the genomes of 41 isolates representative of the A. thaliana root mycobiota for comparative analysis with 79 other plant-associated fungi. We report that root mycobiota members evolved from ancestors having diverse lifestyles and retained diverse repertoires of plant cell wall-degrading enzymes (PCWDEs) and effector-like small secreted proteins. We identified a set of 84 gene families predicting best endophytism, including families encoding PCWDEs acting on xylan (GH10) and cellulose (AA9). These genes also belong to a core transcriptional response induced by phylogenetically-distant mycobiota members in A. thaliana roots. Recolonization experiments with individual fungi indicated that strains with detrimental effects in mono-association with the host not only colonize roots more aggressively than those with beneficial activities but also dominate in natural root samples. We identified and validated the pectin degrading enzyme family PL1_7 as a key component linking aggressiveness of endophytic colonization to plant health.

sample_organism:
Arabidopsis thaliana
Macrophomina phaseolina
Truncatella angustata
Halenospora varia
Paraphoma chrysanthemicola
Phaeosphaeria sp.
Chaetomium sp.
Chaetomium sp. MPI-CAGE-AT-0009
Phaeosphaeria sp. MPI-PUGE-AT-0046c

overall_design: We report the transcriptomes of six root mycobiota members in interaction in mono-association with their host Arabidopsis thaliana in gnotobiotic system. The six fungal strains used in this experiment were isolated from the roots of healthy Arabidopsis plants in the wild (Duran et al. 2018). To identify the genes they express in planta, we conducted a RNA-sequencing experiment. A. thaliana sterile seeds were inoculated with mycelium, then cultivated for 28 days on a phosphate-deficient agar-based medium. Roots were harvested and crushed, then total RNA was extracted and sequenced. As control conditions, mycelia were grown on the same plant culture medium and fungal RNA was extracted with the same method, after 28 days in culture. With this experiment, we identified a core set of genes - mostly composed of genes encoding carbohydrate active enzymes - which are over-expressed by all six fungi in the roots of A. thaliana. We believe they constitute a toolbox for root colonization and the endophytic lifestyle.

first: Bruno

last: Hüttel

first: Stephane

last: Hacquard


title: In vitro generation of mature murine plasma cells uncovers adaptive Prdm1 expression

city: York

first: Matthew

middle: Anthony

last: Care

state: North Yorkshire

address: Wentworth Way

department: Department of Biology

country: United Kingdom

institute: The University of York

summary: Upon encounter with antigen, B cells undergo a sequential process of differentiation to become antibody-secreting plasma cells. While the key drivers of differentiation have been identified, research has been limited by the lack of in vitro models recapitulating the full process for murine B cells. Here we describe methodology using BCR or TLR ligation to obtain plasma cells that are phenotypically mature, have exited cell cycle and express a gene signature concordant with long-lived plasma cells.  Dependent on the initial stimuli, the transcriptomes also show variation, suggestive of unique functional properties for the resultant plasma cells. Moreover, using the new culture conditions we demonstrate that alternative promoter choice regulating the expression of the master transcription factor Blimp-1 can be observed. Thus, our novel system provides a readily tractable model for furthering our understanding of plasma cell biology.

overall_design: mRNA profiles of 45 samples, 3 replicates for each time/condition

first: Gina

middle: M

last: Doody

first: Emily

last: Robinson


title: Microskeletal stiffness promotes aortic aneurysm by sustaining pathological vascular smooth muscle cell mechanosensation via Piezo1

status: Public on Nov 01 2021

first: Michele

last: Silvestro

state: NY

address: 550 1st Avenue

institute: New York University Langone Medical Center

summary: Mechanical overload of the vascular wall is a pathological hallmark of life-threatening abdominal aortic aneurysms (AAA). However, how this mechanical stress resonates at the unicellular level of vascular smooth muscle cells (VSMC) is undefined. Here, we combined novel tweezers-based micromechanical system and single-cell RNA sequencing to map defective mechano-phenotype signatures of VSMC in AAA. Notably, theoretical modelling predicted that cytoskeleton alterations fueled cell membrane tension of VSMC, thereby modulating their mechanoallostatic responses which were validated by live micromechanical measurements. Mechanistically, VSMC gradually adopted a mechanically solid-like state by upregulating CSK crosslinker, α-actinin2, in the presence of AAA-promoting signal, Netrin-1, thereby directly powering the activity of mechanosensory ion channel Piezo1. Inhibition of Piezo1 prevented mice from developing AAA by alleviating pathological vascular remodeling. Our findings demonstrate that deviations of mechanosensation behaviors of VSMC is detrimental for AAA and identifies Piezo1 as a novel culprit of mechanically fatigued aorta in AAA.

overall_design: Single cell RNA-sequencing datasets of abdominal aortas from Apolipoprotein E ko mice infused with Angiotensin II (4 hashtagged samples + 1 sample + 1 pooled sample (n=3)) or PBS (3 hashtagged samples + 1 pooled sample (n=3)) for 28 days


title: Congenital iRHOM2 deficiency causes ADAM17 dysfunction and environmentally directed immunodysregulatory disease [T cells]

overall_design: mRNA-seq profiling of activated T cells


title: Gardnerella vaginalis activates host responses necessary for Escherichia coli recurrent UTI from bladder reservoirs

city: St. Louis

first: Nicole

last: Gilbert

state: MO

address: 425 S Euclid Ave

institute: Washington University School of Medicine

summary: Recurrent urinary tract infections (rUTI) are a costly clinical problem affecting millions of women worldwide each year. The majority of rUTI cases are caused by uropathogenic Escherichia coli (UPEC). Data from humans and mouse models indicate that some instances of rUTI are caused by UPEC emerging from latent reservoirs in the bladder. Some studies have reported that women with vaginal dysbiosis, typically characterized by high levels of Gardnerella vaginalis and other anaerobes, are at increased risk of UTI. Multiple studies have detected G. vaginalis in urine collected by transurethral catheterization (to limit vaginal contamination), suggesting that some women experience routine urinary tract exposures. We recently reported that inoculation of G. vaginalis into the bladder triggers rUTI from UPEC bladder reservoirs in a mouse model. Here we performed whole bladder RNAseq to identify host pathways involved in G. vaginalis-induced rUTI. We identified multiple host pathways differentially expressed following G. vaginalis exposure. At the gene and transcript level, we identified upregulation of the orphan nuclear receptor Nur77 (aka Nr4a1) and Nur77-regulated genes. Pilot data from Nur77 knockout mice suggests that Nur77 is necessary for G. vaganalis exposure to trigger rUTI.

overall_design: A total of 20 mouse bladders, all containing quiescent UPEC reservoirs, were analyzed. 5 bladders from each of the following groups: 12 hours (h) after one PBS exposure (PBS-1), 12h after one G. vaginalis exposure (Gvag-1), 12h after 2 PBS exposures given 12h apart (PBS-2), 12h after 2 G. vaginalis exposures given 12h apart (Gvag-2)

first: Valerie

middle: P

last: O'Brien

first: Amanda

middle: L

last: Lewis


title: Comparison of tumor microenvironment of murine lung cancer ASB-XIV and LLC1.

city: Osaka

first: Koji

last: Nagaoka

address: 377-2 Ohno-Higashi, Osaka-Sayama

department: Department of Immunology

country: Japan

institute: Kindai University Faculty of Medicine

summary: Murine lung cancer cell lines, ASB-XIV and LLC1 are sensitive and resistant to immune checkpoint blockade, respectively. To compare their tumor microenvironment, we performed RNA-Seq analysis.

overall_design: BALB/c or C57BL/6 mice were inoculated with 1×106 ASB-XIV or LLC1 cells, respectively. CpG oligodeoxinucleotide (30 microgram/mouse) was subcutaneously injected the day before tumor inoculation. Seven, 14 and 21 days later, tumors were harvested and subjected to RNA-Seq.

first: Changbo

last: Sun

first: Kazuhiro

last: Maejima

first: Hidewaki

last: Nakagawa

last: Kakimi


title: Maternal cannabis use is associated with suppression of immune gene networks in placenta and increased anxiety phenotypes in offspring

city: New York City

first: Yoko

last: Nomura

address: 1470 Madison Ave

institute: Icahn School of Medicine at Mt. Sinai

summary: While cannabis is among the most used recreational drugs during pregnancy, the impact of maternal cannabis use (mCB) on fetal and child development remains unclear. Here, we assessed the effects of mCB on psychosocial and physiological measures in young children along with the potential relevance of the in-utero environment reflected in the placental transcriptome. Children (~3-6 years) were assessed for hair hormone levels, neurobehavioral traits on the behavioral assessment system for children (BASC-2) survey, and heart rate variability (HRV) at rest and during auditory startle. For a subset of children with behavioral assessments, placental specimens collected at birth were processed for RNA-sequencing. Hair hormone analysis revealed increased cortisol levels in mCB children. In addition, mCB was associated with greater anxiety, aggression, and hyperactivity. Children with mCB also showed a reduction in the high frequency component of HRV at baseline, reflecting reduced vagal tone. In the placenta, there was reduced expression of many genes involved in immune system function including type I interferon, neutrophil, and cytokine signaling pathways. Furthermore, cannabinoid receptor 1 (CNR1) was among the many differentially expressed genes and inversely correlated with mCB.  Finally, several of these mCB-linked immune genes organized into co-expression networks that correlated with child anxiety and hyperactivity. Overall, our findings reveal a relationship between mCB and immune response gene networks in placenta as a potential mediator of risk for anxiety-related problems in early childhood.

overall_design: Bulk RNA-sequencing of term human placenta

first: Gregory

middle: R

last: Rompala


title: RNA-seq from ileal lamina propria CD4+ cell transcripts in germ-free and E. lenta monocolonized mice

city: San Francisco

first: Margaret

last: Alexander

state: California

address: 513 Parnassus Ave

institute: UCSF

summary: The goals of this study were to assess the differentially expressed transcripts in CD4+ cell isolated from the ileal lamina propria of germ-free (GF) and E. lenta strain 2243 monocolonized mice.

overall_design: 4 mice per group (4 germ-free, 4 E. lenta monocolonized)


title: A Novel Strategy for Combination of Clofarabine and Pictilisib is Synergistic in Gastric Cancer

city: miami

first: RIMPI

last: KHURANA

state: FL

address: Room 6112B, Rosenstiel Medical Science Building (RMSB),1600 NW 10th Avenue

institute: University of miami

summary: Purpose: We aimed to identify a novel therapeutic approach in gastric cancer using high-throughput screening and our computational SynerySeq pipeline. We investigated the therapeutic efficiency of clofarabine as a single agent and in combination with the PI3K inhibitor pictilisib in treating in vitro and in vivo models of gastric cancer.

overall_design: Examination of 2 gastric adenocarcinoma cell lines AGS and MKN45 treated with Clofarabine compound

first: Shayan

last: Khalafi

first: Shoumin

last: Zhu

first: Rimpi

last: Khurana

first: Ines

last: Lohse

first: Silvia

last: Giordano

first: Hassan

last: Al-Ali

first: Shaun

last: Brothers

first: Claes

last: Wahlestedt

first: Stephan

last: Schürer

first: Wael

last: El-Rifai


title: Skeletal muscle derived Musclin protects the heart during pathological overload

status: Public on Nov 02 2021

city: Mannheim

first: Joerg

last: Heineke

state: Baden Wuttemberg

address: Ludolf-Krehl-Str. 7-11

department: – Department of Cardiovascular Research (HKF)

institute: Medizinische Fakultät Mannheim

summary: Cachexia is associated with poor prognosis in patients with chronic heart failure. The underlying mechanisms of cachexia triggered heart failure progression, however, are not well understood. Here, we investigated whether the dysregulation of myokine expression from wasting skeletal muscle and impaired inter-organ crosstalk during advanced heart failure might contribute to progression of the disease.   RNA sequencing analysis from wasting skeletal muscles of mice with cardiac cachexia during long-term pressure overload revealed a strongly reduced expression of Ostn. Ostn encodes for the skeletal muscle derived myokine Musclin, which had been previously implicated in the enhancement of natriuretic peptide (NP) signaling. Using newly developed skeletal muscle specific, inducible Ostn knock-out mice, we demonstrated that reduced skeletal muscle Musclin levels exaggerated cardiac dysfunction and myocardial fibrosis compared to littermate control mice after TAC. Restoration of Musclin deficiency during cardiac cachexia via AAV6-mediated skeletal muscle specific Musclin overexpression, in turn, attenuated left ventricular dysfunction and myocardial fibrosis. Mechanistically, we found that Musclin enhanced CNP/NPR2/cGMP signaling in cardiomyocytes, which led to improved contractility by inhibition of the cAMP degrading phosphodiesterase (PDE)3 and augmented cAMP/protein kinase A signaling. In addition, Musclin directly acted on cardiac fibroblasts to inhibit their activation.   Together, our study indicates the therapeutic potential of targeting interorgan cross-talk during heart failure, for example by counteracting the impaired secretion of the Musclin from wasting skeletal muscle.

overall_design: At the age of 8 weeks C57Bl6 mice were exposed to transverse aortic constriction (TAC) around a 27 gauge needle or sham surgery. 12 weeks after surgery, the experiment was terminated and RNA was extracted.

first: Badder

last: Kattih

first: Julio

last: Cordero


title: A single-cell transcriptomic analysis of cranial neural crest development reveals timing of lineage decisions and novel regulators

city: Irvine

first: Thomas

middle: F

last: Schilling

address: 4462 Natural Sciences II

department: Developmental and Cell Biology

institute: University of California Irvine

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


title: Epigenetic analysis of sperm DNA methylation from a randomized placebo controled study of zinc and folic acid suplementation

city: Provo

first: Tim

last: Jenkins

state: UT

address: 4005 Life Sciences Building

institute: Brigham Young University

summary: Sperm DNA methylation was assessed in pateints who were randomly assigned to a folic acid suplementation group or a palcebo group. Basic differential methylation analysis was performed to identify sites of differential methylation.

overall_design: Bisulfite converted sperm DNA was hybridized to Illumina's EPIC array to determine the impact of 5 mg of folic acid and 30 mg of elemental zinc suplementation daily for six months


title: The developmental origins of heterodonty and acrodonty as revealed by reptile dentitions

city: University of Helsinki

first: Nicolas

last: Di-Poi

address: Viikinkaari 1

department: Institute of Biotechnology, Helsinki Institute of Life Sciences

country: Finland

institute: University of Helsinki

summary: Owing to their exceptional diversity and central role in vertebrate evolution, teeth are key biological systems. We exploited the diversity of bearded dragon dentitions to unreveal new aspects of major dental characters only represented in non-mammalian lineages.

overall_design: RNA sequencing of dental tissues from wild-type and Scaleless mutant bearded dragon lizards, using NextSeq 500 platform (Illumina, U.S.A.).

first: Lotta

last: Salomies

first: Julia

last: Eymann

first: Joni

last: Ollonen

first: Imran

last: Khan

last: Di-Poï


title: scRNA-seq timeline of first pharyngeal arch migratory stream in zebrafish

summary: We have performed single-cell RNA-seq of cranial NC cells in zebrafish over several stages during their migration

overall_design: single cells were captured via FACS and 10X Chromium platform at 6 stages across neural crest migration.

first: David

last: Tatarakis

first: Zixuan

last: Cang

first: Xiaojun

last: Wu

first: Matt

last: Karikomi

first: Adam

last: MacLean

first: Qing

last: Nie


title: Heterochromatin is a quantitative trait locus associated with spontaneous epiallele formation

city: Athens

first: Robert

last: Schmitz

state: GA

address: B416 Davison Life Sciences

department: Genetics

institute: University of Georgia

summary: Epialleles are meiotically inherited variations in expression states that are not linked to changes in DNA sequence. Although they are well documented to persist in plant genomes, their molecular origins are unknown. Here, we show using a variety of mutant and experimental populations that epialleles in Arabidopsis thaliana result from feedback regulation of pathways that primarily function to maintain DNA methylation at heterochromatin. Perturbations to maintenance of heterochromatin methylation leads to feedback regulation of DNA methylation in genes, with a preference for genes with pre-existing DNA methylation. Using epigenetic recombinant inbred lines (epiRIL), we show that epiallelic variation is enriched in euchromatin, yet, associated with QTL primarily located in heterochromatin. Mapping three-dimensional chromatin contacts reveals that genes that are hotspots for epiallelic variation have increased contact frequencies with regions possessing H3K9me2. Altogether, these data show that feedback regulation of pathways that evolved to maintain heterochromatin silencing leads to the origins of spontaneous epialleles.

overall_design: MethylC-seq


title: High-throughput sequencing analysis of mouse FABP5 KO BMDM cell transcriptomes with overexpression of FABP5 WT or FABP5 C127S after LPS treatment

city: Hangzhou

first: Yuxian

last: Guo

address: Department of Basic Medical Sciences,   School of Medicine, Zhejiang University,  388 Yuhangtang Rd

institute: Zhejiang University School of Medicine

summary:
Purpose: Mouse BMDM is the universal cell type of studying innate immunity. This study was designed to analyze LPS induced innate immune response and the gene expression in FABP5 KO BMDMs with overexpression of FABP5 WT or FABP5 C127S.
Methods: FABP5 KO BMDMs were nucleofected with 5 μg pXJ40-3xFlag-FABP5 WT or C127S plasmid using the Amaxa Mouse Macrophage Nucleofector Kit (Lonza, VPA-1009) following the manufacturer’s instructions. Replace medium 6 hours post Nucleofection and add 500 ng/mL LPS to the fresh medium. 24 hours after treatment, harvest cells by using GenElute Single Cell RNA Purification Kit (Sigma, RNB300) and perform RNA-seq. Then mRNA profiles of these samples were generated by high-throughput sequencing analysis, using Illumina NovaSeq6000. And the differential mRNA profiles were analyzed.
Results: mRNA profiles were analyzed, and differential expression profiles were compared.
Conclusions: Our study demonstrated the transcriptional profiles of FABP5 KO BMDMs with overexpression of FABP5 WT or FABP5 C127S upon LPS treatment, with biologic replicates, generated by RNA-seq technology.

overall_design: mRNA profiles of FABP5 KO BMDMs with overexpression of FABP5 WT or FABP5 C127S upon LPS treatment were generated by high-throughput sequencing, with biologic replicates, using Illumina NovaSeq6000.

first: Xue

last: Zhang

first: Yuehai

last: Ke

first: Yaru

last: Liu


title: Bulk RNA-seq comparing expression profiles between neural crest cells experiencing high and low levels of Wnt signaling

first: Thomas

last: Schilling

summary: The purpose of this study was to obtain an unbiased list of wnt-regulated genes in zebrafish neural crest.

overall_design: NC cells were collected from 7xTCF:mCherry transgenic Wnt reporter zebrafish embryos. 3 samples each were collected for high and low levels of the transgene by FACS.


title: Single Cell Transcriptomic Landscape of Diabetic Foot Ulcers

city: Atlanta

first: Debasree

last: Sarkar

address: 615 Michael Street

department: Pediatrics

institute: Emory University

summary: Diabetic foot ulcers (DFUs) are a devastating complication of diabetes. In order to identify systemic and local factors associated with DFU healing, we examined the cellular landscape of DFUs by single-cell RNA-seq analysis of foot and forearm skin specimens, as well as PBMC samples, from 10 non-diabetic subjects, and 17 diabetic patients, 11 with, and 6 without DFU. Our analysis shows enrichment of a unique inflammatory fibroblast population in DFU patients with healing wounds. The patients with healing DFUs also depicted enrichment of macrophages with M1 polarization, as opposed to more M2 macrophages in non-healing wounds. These findings were verified using Immunohistochemistry and Spatial Transcriptomics.

overall_design:
Single-cell RNA sequencing of foot and forearm skin cells, as well as  peripheral blood mononuclear cells (PBMCs), from 10 samples from non-diabetic subjects, and 17 diabetic patients, 11 with, and 6 without DFU.

***Raw data not available due to patient privacy concerns***

first: Manoj

last: Bhasin

first: Aristidis

last: Veves


title: Next generation sequencing of WT or Mir139KO MLL-AF9 cells

city: Rotterdam

first: Stefan

middle: J.

last: Erkeland

state: Zuid-Holland

address: Dr. Molewaterplein 40

country: Netherlands

institute: Erasmus University Medical Center

overall_design: Refer to individual Series


title: Next generation sequencing of RNA of WT, KO and MLL-AF9 transformed HSPCS

summary: MIR139 is a tumor suppressor and commonly silenced in Acute myeloid leukemia (AML). Reactivating the expression of MIR139 eliminates AML  cells. Here, we investigated the mechanism of MIR139 gene inactivation in AML expressing the Mixed-Lineage Leukemia (MLL)-AF9 oncogene. We found that MLL-AF9-mediated repression of MIR139 is a selective event in leukemogenesis. Analyses of Histone marks revealed two well-conserved enhancer regions, which are epigenetically silenced by the Polycomb-Repressive Complex-2 (PRC2) downstream of MLL-AF9. Genomic deletion of these enhancer regions abolished transcriptional regulation of MIR139. Genome-wide knockout screens revealed the transcriptional pausing factor of RNA Polymerase-II, POLR2M, as a critical MIR139 silencing factor. Furthermore, POLR2M-binding to the MIR139 transcriptional start site induces paused transcription, which is abrogated upon PRC2 inhibition. Together, we present evidence for a POLR2M-mediated MIR139 silencing mechanism, downstream of MLL-AF9 and PRC2. The findings in this study highlight the importance of the transcriptional deregulation in malignant transformation.

overall_design: RNAs-seq of 4 WT HSPCs, 4 Mir139KO and 6 MLL-AF9 transformed HSPCs

first: Christiaan

last: Stavast


title: Next generation sequencing of DNA of WT or Mir139KO MLL-AF9 cells transduced with lenticrisprv2 gecko library

overall_design: NGS of WT MLL-AF9 cells tranduced with CRISPR sgRNA library 24hrs post selection and 14 days post selection and Mir139KO MLL-AF9 cells  tranduced with CRISPR sgRNA library 14 days post selection.

first: Wilfred

last: van IJcken

first: Rutger

last: Brouwer


title: Single-cell RNAseq of the human normal/healthy lung

first: Tien

last: Peng

address: 513 Parnassus Ave, Health Sciences East 1350

department: Department of Pulmonary Medicine

institute: University of California San Francisco

summary: To study adult human lung, we conduct single-cell RNAseq of the human healthy lung using 10X scRNAseq technique.

overall_design: Adult human lung epithelial and mesenchymal cells were sorted based on EpCAM+ and EpCAM-/CD11b-/CD45-/CD31-, respectively. Dead cells were excluded using DRAQ7 negative selection. The sequencing sample were mixed with half epithelial cells andhalf mesenchymal cells.

first: Chaoqun

last: Wang


title: Next generation sequencing of RNA of MLL-AF9-139 cells treated with and without Doxycycline

overall_design: RNAs-seq of 3 MLL-AF9-i139 and 1 MLL-AF9-ieGFP clones treated with Doxycycline and 3 MLL-AF9-i139 and 1 MLL-AF9-ieGFP clones treated without Doxycycline


title: 9-ING-41, a small molecule inhibitor of GSK-3β, potentiates the effects of chemotherapy in KRAS mutated colorectal cancer

status: Public on Oct 31 2021

city: Moscow

first: Sergey

last: Nikulin

address: 7, Vavilova st.

department: Faculty of Biology and Biotechnology

country: Russia

institute: HSE University

summary: Despite the prevalence of KRAS mutations in colorectal cancer, there is no K-Ras targeted therapies for these tumors and available treatment options are limited for metastatic disease. GSK-3β has been demonstrated to be a critically important kinase for survival and proliferation of K-Ras-depended pancreatic cancer cells. In this study we tested 9-ING-41, a small molecule inhibitor of GSK-3β, in patient-derived tumor organoid models of colorectal cancer. We demonstrated that addition of 9-ING-41 to the standard of care drugs 5-FU and oxaliplatin could significantly enhance inhibition of the growth of colorectal cancer cells harboring KRAS mutations. The results of the transcriptomic analysis support our findings of cell cycle arrest and DNA repair deficiency in 9-ING-41-treated colorectal cancer cells. Moreover, we found substantial similarity in the changes of transcriptomic profile after inhibition of GSK-3β and suppression of STK33, another critically important kinase for K-Ras-dependent cells. Overall, the results of this study provide a rationale for the inclusion of patients with mutant KRAS colorectal cancer in clinical studies of 9-ING-41 and other GSK-3 inhibitors.

overall_design: Total mRNA profiles of 9-ING-41 treated and control colorectal tumor organoids.


title: Single-cell transcriptomics revealed a critical role of SPP1/CD44-mediated crosstalk between macrophages and cancer cells in glioma

city: Shanghai

first: Deshui

last: Jia

address: 650 Xin Songjiang Road

institute: Shanghai General Hospital, Shanghai Jiao Tong University School of Medicine

summary: a single-cell RNA sequencing analysis of one high-grade glioma with wide-type IDH genes, revealing the landscape of tumor and immune cells in the ecosystem of this tumor

overall_design: one sample from glioma tissue

first: Yaohua

first: Cong

last: He

first: Xiaojun

last: Ma


title: Bulk RNAseq of fetal human lung fibroblasts (MRC5) and primary adult human lung mesenchyme (AHLM)

summary: The fetal human lung fibroblasts (MRC5) and primary adult human lung mesenchyme (AHLM) have different capabilities in supporting human alveolar type 2 cell transdifferentiation into basal KRT5+ cells.Therefore, we examined the transcriptome of AHLM vs. MRC5 by bulk RNAseq.

overall_design: MRC5 and AHLM cells were cultured in DMEM/F12 with 10% FBS for 2 days. Then the total RNA were extracted from technical triplicates of MRC5 and AHLM cells, respectively. The RNA of the samples were sent to GENEWIZ company for sequencing. Raw sequence data (.bcl files) generated from Illumina HiSeq was converted into fastq files and de-multiplexed using Illumina's bcl2fastq 2.17 software.


title: Peri-transplant Glucocorticoids Redistribute Donor T-cells to the Bone Marrow and Prevent Relapse after haploidentical SCT

city: Pittsburgh

first: Warren

middle: David

last: Shlomchik

state: PA

address: 3550 Terrace Street

department: Medicine

institute: University of Pittsburgh

summary: Patients with acute leukemia who are unable to achieve complete remission prior to allogeneic hematopoietic stem cell transplantation (SCT) have dismal outcomes with relapse rates well in excess of 60%. Haplo-identical SCT (haplo-SCT) may allow enhanced graft-versus-leukemia (GVL) effects by virtue of HLA class I/II donor-host disparities but typically requires intensive immune-suppression with post-transplant cyclophosphamide (PT-Cy) to prevent lethal graft-versus-host disease (GVHD). Here we demonstrate in preclinical models that glucocorticoid administration from day -1 to +5 inhibits alloantigen presentation by professional recipient antigen presenting cells in the gastrointestinal tract and prevents donor T-cell priming and subsequent expansion therein. In contrast, direct glucocorticoid signaling of donor T-cells promotes chemokine and integrin signatures permissive of preferential circulation and migration into the bone marrow, promoting donor T-cell residency.

overall_design: 6 samples (3 controls)

first: Scott

last: Furlan

first: Geoff

last: Hill


title: RNA-Seq expression analysis of thymic epithelial cells (TEC) lacking expression of Hoil

city: Parkville

first: Waruni

last: Abeysekera

state: Victoria

address: 1G Royal Parade

department: Bioinformatics

institute: Walter and Eliza Hall Institute of Medical Research

summary: To investigate the role of LUBAC in the development and maintenance of the thymic epithelail cells, we analysed the transcriptional changes that result from loss of HOIL - a critical component of the LUBAC protein complex rnecessary for complex assembly, stability and therefore function - in purified mouse thymic epithelial cell populations.

overall_design: RNA was isolated from freshly purified cortical and medullary thymic epithelial cells of control mice (Foxn1cre+/+ Hoillox/lox) and mice carrying TEC-specific deletion of Hoil (Foxn1creT/+ HOILlox/lox) for comparison by RNA-Seq.

first: Reema

last: Jain

first: Julie

last: Sheridan

first: Melanie

last: Heinlein

first: Gordon

middle: H

last: Smyth

first: Daniel

last: Gray


title: Developmental Programming: Impact of prenatal testosterone excess on liver and muscle transcriptome and the relationship between non-coding and coding RNA networks in female sheep

city: Ann Arbor

first: Kelly

last: Bakulski

state: MI

address: 1415 Washington Heights

department: Department of Epidemiology

institute: University of Michigan

type: Non-coding RNA profiling by high throughput sequencing


title: Developmental Programming: Impact of prenatal testosterone excess on liver and muscle transcriptome and the relationship between non-coding and coding RNA networks in female sheep [ncRNA-seq]

summary: This study seeks to assess the effects of prenatal exposure of female sheep to execessive testosterone in metabolically relevant liver and muscle tissue. The goals are to 1) determine noncoding RNA in the control animals and 2) assess the effects of prenatal T-treatment on  non-coding RNA. 3) Finally identify putative ncRNA-totalRNA interactions.

overall_design: 8 Female Sheep. 4 Controls and 4 Treatment. 2 Tissues-Liver and Muscle. 16 Samples total- 8 Samples Liver(4C, 4T) 8 Samples Muscle(4C, 4T)

first: Muraly

last: Puttabyatappa

first: Nadia

last: Saadat

first: Venkateswaran

last: Elangovan

first: John

last: Dou

first: Robert

last: Thompson

first: Vasantha

last: Padmanabhan


title: Early Transcriptomic Response to Burn injury: Severe Burns are Associated with Immune Pathway Shutdown

city: Silver Spring

first: Ruoting

last: Yang

state: MD

address: 503 Robert Grant Ave

department: CMPN

institute: WRAIR

summary: Burn injury induces a systemic hyperinflammatory response with detrimental side effects. Studies have described the biochemical changes induced by severe burns, but the transcriptome response is not well characterized. The goal of this work is to characterize the blood transcriptome after burn injury. Burn patients presenting to a regional center between 2012-2017 were prospectively enrolled.  Blood was collected on admission and at predetermined time points (hours 2, 4, 8, 12, 24). RNA was isolated and transcript levels were measured with a gene expression microarray. To identify differentially regulated genes (FDR≤0.1) by burn injury severity, patients were grouped by total body surface area (TBSA) above or below 20% and statistically enriched pathways were identified. Sixty-eight patients were analyzed, most patients were male with a median age of 41 (IQR, 30.5-58.5) years, and TBSA of 20% (11-34%). Thirty-five patients had %TBSA injury ≥20%, and this group experienced greater mortality (26% vs. 3%, p=0.008). Comparative analysis of genes from patients with </≥20% TBSA revealed 1505, 613, 380, 63, 1357, and 954 differentially expressed genes at hours 0, 2, 4, 8, 12 and 24 respectively.  Pathway analysis revealed an initial upregulation in several immune/inflammatory pathways within the ≥20% TBSA groups followed by shutdown. Severe burn injury is associated with an early proinflammatory immune response followed by shutdown of these pathways. Examination of the immunoinflammatory response to burn injury through differential gene regulation and associated immune pathways by injury severity may identify mechanistic targets for future intervention.

overall_design: Burn patients presenting to a regional center between 2012-2017 were prospectively enrolled.  Blood was collected on admission and at predetermined time points (hours 2, 4, 8, 12, 24). RNA was isolated and transcript levels were measured with a gene expression microarray. patients were grouped by total body surface area (TBSA) above or below 20% and statistically enriched pathways were identified. Sixty-eight patients were analyzed, most patients were male with a median age of 41 (IQR, 30.5-58.5) years, and TBSA of 20% (11-34%).

first: Aarti

last: Gautam

first: Stacy-Anne

last: Miller

first: Rasha

last: Hammamieh

first: Bonnie

last: Carney

last: Keyloun


title: High-resolution mapping of transcribing RNAPII by CRAC in a wt or a sen1T1623E mutant

city: Paris

first: Odil

last: Porrua Fuerte

address: 15 rue Helene Brion

institute: Institut Jacques Monod-CNRS

summary:
Manuscript title: Modulated termination of non-coding transcription partakes in the regulation of gene expression
Here we report high-resolution analyses of transcribing RNAPII in either a wild-type (WT) background or a sen1T1623E mutant, which harbours a substitution that mimics the phosphorylation at threonine 1623. Sen1 is a well-characterized transcription termination factor for RNAPII-dependent non-coding genes in budding yeast. Here we show that the T1623E phosphomimetic mutation induces moderate but widespread termination defects at target non-coding genes, strongly suggesting that phosphoryation at T1623 modulates negatively Sen1 transcription termination activity.

overall_design: Analysis of the effect of the Sen1 T1623E mutation on RNAPII-dependent transcription genome-wide using CRAC

last: Porrua


title: Transcriptional Pathology Evolves Over Time in Rat Hippocampus Following Lateral Fluid Percussion Traumatic Brain Injury

city: Davis

first: Alex

middle: S.

last: Nord

state: CA

address: 1544 Newton Court

department: Center for Neuroscience

institute: University of California Davis

summary: Traumatic brain injury (TBI) causes acute and lasting impacts on the brain, driving pathology along anatomical, cellular, and behavioral dimensions. Rodent models offer the opportunity to study TBI in a controlled setting, and enable analysis of the temporal progression that occurs from injury to recovery. We applied transcriptomic and epigenomic analysis, characterize gene expression and in ipsilateral hippocampus at 1 and 14 days following moderate lateral fluid percussion (LFP) injury. This approach enabled us to identify differential gene expression (DEG) modules with distinct expression trajectories across the two time points. The major DEG modules represented genes that were up- or downregulated acutely, but largely recovered by 14 days. As expected, DEG modules with acute upregulation were associated with cell death and astrocytosis. Interestingly, acutely downregulated DEGs related to neurotransmission mostly recovered by two weeks. Upregulated DEG modules related to inflammation were not necessarily elevated acutely, but were strongly upregulated after two weeks. We identified a smaller DEG module with delayed downregulation at 14 days including genes related to cholesterol metabolism and amyloid beta clearance. Finally, differential expression was paralleled by changes in H3K4me3 at the promoters of differentially expressed genes at one day following TBI. Following TBI, changes in cell viability, function and ultimately behavior are dynamic processes. Our results show how transcriptomics in the preclinical setting has the potential to identify biomarkers for injury severity and/or recovery, to identify potential therapeutic targets, and, in the future, to evaluate efficacy of an intervention beyond measures of cell death or spatial learning.

overall_design: Examination of transcriptional differences between TBI and sham control rat hyppocampus samples at 1 and 14 days after lateral fluid percussion injury, and epigenetic perturbations 1 day after TBI.

first: Rinaldo

last: Catta-Preta

first: Iva

last: Zdillar

first: Bradley

last: Jenner

first: Emily

middle: T

last: Doisy

first: Kayleen

last: Tercovich

middle: S

first: Gene

middle: G

last: Gurkoff


title: Developmental Programming: Impact of prenatal testosterone excess on liver and muscle transcriptome and the relationship between non-coding and coding RNA networks in female sheep [RNA-seq]

first: Kelly

last: Bakulski

summary: This study seeks to assess the effects of prenatal exposure of female sheep to execessive testosterone in metabolically relevant liver and muscle tissue. The goals are to 1) determine genes and gene pathways in the control animals and 2) assess the effects of prenatal T-treatment on gene expression and gene pathways. 3) Finally identify putative ncRNA-totalRNA interactions.

overall_design: 10 Female Sheep. 5 Controls and 5 Treatment. 2 Tissues-Liver and Muscle. 9 Samples Liver(4C, 5T) 10 Samples Muscle(5C, 5T)

first: Muraly

last: Puttabyatappa

first: Nadia

last: Saadat

first: Venkateswaran

last: Elangovan

last: Dou

last: Thompson


title: Pre and post Classical Swine Fever (CSF) vaccination PBMC transcriptomes of Indian Ghurrah and Landrance pigs

city: Bareilly

first: Amit

last: Kumar

state: UP

address: Izatnagar

department: Animal Genetics

country: India

institute: Indian Veterinary Research Institute

summary: In this study, we generated the PBMC transcriptome data of 3 Landrace, and 3 indigenous Ghurrah pigs before and 7 days after Classical Swine fever vaccination. The piglets were 8-12 weeks of age. The sequencing was done on Illumina X-Ten platform, generating  average of 39 million 150 bp paired end reads for each sample.

overall_design: Across and within breed comparison of PBMC transcriptomes, before and after CSF vaccination.

first: Arnav

last: Mehrotra


title: 10x single-cell RNAseq profiling of influenza Virus infected Human PBMCs treated with Dimethyl itaconate

city: Hannover

first: Frank

last: Pessler

address: Feodor-Lynen-Str. 7-9

department: Experimental Infection Research

institute: Center for Experimental and Clinical Infection Research GmbH

summary: Itaconic acid and its variant Dimethyl itaconate (DI) have shown an anti-inflammatory effect in different human cell lines and PBMCs during Infleunza A virus (IAV) infection.  In this experiment, we further explored which cell types in PBMCs respond to IAV infection and DI treatment.

overall_design: Human PBMCs were isolated from blood and cultured in medium. PBMCs were infected with IAV and treated with DI.

first: Robert

last: Geffers


title: Expression data from differentiated THP-1 (dTHP-1) cell line infected with influenza A virus (IAV) and treated with/without Itaconate

summary:
Analysis of gene expression indTHP-1 cells infected with influenza A virus or Mock and treated with Itaconic acid (IA), or Dimethyl itaconate (DI) to investigate the effects itaconate have on transcriptional response in dTHP-1 cells.
We particularly looked at anti-inflammatory effects of itaconate in inflammation followed by Influenza A virus infection.

overall_design: THP1-1 cells were differentiated by phorbol 12-myristate-13-acetate. dTHP-1 cells were infected with influenza A viruses, or non-infected (Mock) for 12 hours. Infected dTHP-1 cells were pre-treated with itaconic acid, or dimethyl itaconate for 24 hours and then an additional 12 hours treatment along with IAV infection. RNA was extracted, and subjected to transcriptome profiling array  Clariom™ S Assay, human.


title: Differential regulation of cranial and cardiac neural crest by Serum Response Factor

status: Public on Oct 30 2021

city: NEW YORK

first: Philippe

last: Soriano

address: 1468 Madison Avenue, Annenberg 25-70, Box 1020

department: Cell, Development, and Regenerative Biology

institute: Icahn School of Medicine at Mount Sinai

summary: The goal of this experiment was to identify genes regulated by Serum Response Factor (SRF) in the neural crest in the craniofacial region, including whether there were different targets in different craniofacial prominences. SRF is an essential transcription factor that influences many cellular processes including cell proliferation, migration, and differentiation. SRF directly regulates and is required for immediate early gene (IEG) and actin cytoskeleton-related gene expression. SRF coordinates these competing transcription programs through discrete sets of cofactors, the Ternary Complex Factors (TCFs) and Myocardin Related Transcription Factors (MRTFs). The relative contribution of these two programs to in vivo SRF activity and mutant phenotypes is not fully understood. To study how SRF utilizes its cofactors during development, we generated a knock-in SrfaI allele in mice harboring point mutations that disrupt SRF-MRTF-DNA complex formation but leave SRF-TCF activity unaffected. Homozygous SrfaI/aI mutants die at E10.5 with notable cardiovascular phenotypes, and neural crest conditional mutants succumb at birth to defects of the cardiac outflow tract but display none of the craniofacial phenotypes associated with complete loss of SRF in that lineage.

overall_design: 16 total samples were anlyzed from eight E11.5 mouse embryos. These consist of four control and four conditional Srf mutant mice. Each mouse had two samples, one from the mandible and one from the frontonasal prominence (i.e. lateral and medial nasal processes)

first: Colin

middle: J

last: Dinsmore


title: Expression data from A549 cell line infected with influenza A virus (IAV) and treated with/without Itaconate

summary:
Analysis of gene expression in A549 cells infected with influenza A virus or Mock and treated with Itaconic acid (IA), or Dimethyl itaconate (DI) to investigate the effects itaconate have on transcriptional response in A549 cells.
We particularly looked at anti-inflammatory effects of itaconate in inflammation followed by Influenza A virus infection.

overall_design: A549 cells were infected with influenza A viruses, or non-infected (Mock) for 24 hours. Infected A549 cells were pre-treated to itaconic acid, or dimethyl itaconate for 24 hours and then an additional 24 hours treatment along with IAV infection. RNA was extracted, and subjected to transcriptome profiling array  Clariom™ S Assay, human.


title: Gpx4 prevents Treg-cell lipid peroxidation and ferroptosis in promoting immune homeostasis and tumor immune escape

city: Indianapolis

first: Travis

last: Johnson

state: IN

department: Biostatistics

institute: Indiana University School of Medicine

summary:
Purpose: Identification of Gpx4-dependent genes in resting and activated Treg cells
Methods: RNA was purified from resting of stimulated (aCD3-CD38 for 4 h) splenic Treg cells sorted from WT or Foxp3CreGpx4fl/fl mice
Conclusions: Gpx4 prevents lipid peroxidation and ferroptosis of activated Treg cells..

overall_design: In total 12 samples were analyzed: WT (3 Unstimulated and 3 Stimulated); Gpx4 Ko (3 Unstimulated and 3 stimulated)

first: Chengxian

last: Xu

first: Jie

last: Zhang

first: Kai

last: Yang


title: Expression data from PBMCs infected with influenza A virus and treated with/without Itaconate

summary:
Analysis of gene expression in human PBMCs infected with influenza A virus or Mock and treated with Itaconic acid, Dimethyl itaconate or 4-Octyl itaconate to investigate the effects itaconate have on transcriptional response in human PBMCs.
We particularly looked at anti-inflammatory effects of itaconate in inflammation followed by Influenza A virus infection.

overall_design: Human PBMCs were infected with influenza A viruses, or non-infected (Mock) for 12 hours. Infected PBMCs were treated to itaconic acid, dimethyl itaconate or 4-Octyl-itaconate. RNA was extracted, and subjected to transcriptome profiling array  Clariom™ S Assay, human.


title: BCG invokes superior STING-mediated innate immune response over radiotherapy in a carcinogen murine model of urothelial cancer

city: Baltimore

first: Kara

last: Lombardo

state: Maryland

address: 600 N Wolfe St

department: Urology

institute: Johns Hopkins Medical Institutions

summary: Radiation and BCG instillations are used clinically for treatment of urothelial carcinoma, but the precise mechanisms by which they activate an immune response remain elusive.  The role of the cGAS-STING pathway has been implicated in both BCG and radiation-induced immune response however comparison of STING-pathway molecules and immune landscape following treatment in urothelial carcinoma has not be performed. We therefore comprehensively analyzed the local immune response in the bladder tumor microenvironment following radiotherapy and BCG instillations in a well-established spontaneous murine model of urothelial carcinoma to provide insight into activation of STING-mediated immune response.  Mice were exposed to the oral carcinogen, BBN, for 12 weeks prior to treatment with a single 15Gy dose of radiation or 3 intravesical instillations of BCG (1x108 CFU). At sacrifice, tumors were staged by a urologic pathologist and effects of therapy on the immune microenvironment were measured using NanoString Myeloid Innate Immunity Panel and immunohistochemistry. Clinical relevance was established by measuring immune biomarker expression of cGAS and STING on a human tissue microarray consisting of BCG-treated non-muscle invasive urothelial carcinomas.  BCG instillations in the murine model elevated STING and downstream STING-induced interferon and pro-inflammatory molecules, intratumoral M1 macrophage and T-cell accumulation, and complete tumor eradication.  In contrast, radiotherapy caused no changes in STING pathway or innate immune gene expression; rather, it induced M2 macrophage accumulation and elevated FoxP3 expression characteristic of immunosuppression. In human non-muscle invasive bladder cancer, STING protein expression was elevated at baseline in patients who responded to BCG therapy and increased further after BCG therapy. Overall, these results show that STING pathway activation plays a key role in effective BCG-induced immune response and strongly indicate that the effects of BCG on the bladder cancer immune microenvironment are more beneficial than those induced by radiation.

overall_design: In the study presented here, we used the BBN-induced bladder tumor model to compare the tumor immune microenvironment following treatment with BCG and radiotherapy with the goal of understanding mechanisms of immune response and in particular activation of the dna sensing cGAS-STING pathway using gene expression analysis from NanoString innate immunity panel and immnohistochemistry.  We supported our findings from this BBN in-vivo model with in-vitro experiments with wild type and STING KO murine BMDMs treated with BCG and provided clinical relevance using a human tissue microarray stained by IHC for protein expression of cGAS and STING.

middle: A

first: Aleksandar

last: Obradovic

first: Alok

last: Singh

first: James

last: Liu

first: Gregory

last: Joice

first: Max

last: Kates

first: William

last: Bishai

first: David

last: McConkey

first: Alcides

last: Chaux

first: Marie-Lisa

last: Eich

first: M

middle: K

last: Rezaei

first: George

last: Netto

first: Charles

last: Drake

first: Phuoc

last: Tran

first: Andres

last: Matoso

first: Trinity

last: Bivalacqua


title: The Role of Sorting Nexin 17 in Cardiac Development

first: Yufei

last: Wu

address: Siping Campus,1239 Siping Road, Yangpu District, Shangha.

institute: School of Medicine, Tongji University

summary: Homozygous deletion of the SNX17 gene in rats resulted in mid-gestational embryonic lethality which was accompanied by congenital heart defects, including in the double-outlet right ventricle, atrioventricular and ventricular septal defects. To elucidate the potential mechanisms underlying development of cardiac OFT defects induced by SNX17 knockout, we performed RNA-seq analysis in cardiac outflow tract (OFT) tissues isolated from WT and HO embryos at E13.5. A total of 262 differentially expressed genes (DEGs) were identified between WT and HO samples, of which 94 and 168 were up-regulated and down-regulated, respectively. Analysis of biological functions of the DEGs, via Gene Ontology (GO), revealed that the up-regulated genes were mainly involved in heart development, and negative regulation of the intrinsic apoptotic pathway, while the down-regulated DEGs were associated with cell adhesion, extracellular organization, and negative regulation of the Wnt signaling pathway.

overall_design: RNA-seq analysis in cardiac OFT tissues isolated from WT and HO embryos at E13.5.

first: Yaqun

last: Zhou

first: Jian

last: Huang

first: Ke

last: Ma

first: Tianyou

last: Yuan

first: Yong

last: Jiang

first: Maoqing

last: Ye

first: Jun

last: Li


title: RNA sequencing after RNA switch transfection

city: Kyoto

first: Hirohide

last: Saito

state: Kyoto

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

department: Department of Life Science Frontiers

institute: Kyoto University

summary: To reveal the effect of the purification by mRNA switches on the gene expression profiles of the cells.

overall_design: mRNA profiles of HeLa and 293FT cells at 24, 48, and 72 hours after transfection of RNA switches, GFP mRNA and control. miRNA profiles of iPSCs at 72, 96, and 120 hours after transfection of RNA switches and control.

first: Takuya

last: Yamamoto

first: Yoshihiko

last: Fujita

first: Moe

last: Hirosawa


title: Using RNAseq for identifying hepatic genes involved in  multi- (F1 generation) and transgenerational (F2 generation) responses to chronic benzo[a]pyrene  (BaP) and triclosan (TCS) mixture exposure (F0 generation) in Xenopus tropicalis.

city: Saint Martin d'Hères

first: Stéphane

last: REYNAUD

address: 2233 rue de la piscine

institute: Grenoble-Alpes University

summary: We investigated the multi- and transgenerational impact of BaP and TCS mixture on Xenopus females following F0 generation exposure from the tadpole to the mature adult stage at a concentration (50 ng.L-1 each).  5 females from F1 and F2 generations by parentale exposure conditions (control vs Mixture) were used for studying liver transcriptome by RNAseq. RNA-seq libraries were sequenced on an illumina GAIIx sequencer as 75bp single reads

overall_design: mRNAseq from liver of mature F1 and F2 females exposed to BaP and TCS mixture. Processed data file (Cuffdiff) have been performed by using RNAseq sequence from control mRNAseq data deposited on NCBI Gene Expression Omnibus (GEO) (accession no GSE150911)

last: Reynaud


title: Colon cancer cells acquire immune regulatory molecules from tumor-infiltrating lymphocytes by trogocytosis

status: Public on Oct 29 2021

city: Seoul

first: Jaechul

last: Lim

state: CT

address: 1 Gwanak-ro, 1 Gwanak-gu

country: South Korea

institute: Seoul National University

summary: Cancer cells can develop an immunosuppressive tumor microenvironment to control tumor-infiltrating lymphocytes. The underlying mechanisms still remain unclear. Here, we report that mouse and human colon cancer cells acquire lymphocyte membrane proteins including cellular markers such as CD4 and CD45. We observed cell populations harboring both a tumor-specific marker and CD4 in the tumor microenvironment. Sorted cells from these populations were capable of forming organoids, identifying them as cancer cells. Live imaging analysis revealed that lymphocyte membrane proteins were transferred to cancer cells via trogocytosis. As a result of the transfer in vivo, cancer cells also acquired immune regulatory surface proteins such as CTLA4 and Tim3, which suppress activation of immune cells. RNA-sequencing analysis of ex vivo co-cultured splenocytes with trogocytic cancer cells showed reductions in Th1 activation and natural killer cell signaling pathways compared to the non-trogocytic control. Cancer cell trogocytosis was confirmed in the patient-derived xenograft models of colorectal cancer and head and neck cancer. These findings suggest that cancer cells utilize membrane proteins expressed in lymphocytes, which in turn contribute to developing the immunosuppressive tumor microenvironment.

overall_design: Four replicates from co-cultured C57BL/6 mouse splenocytes with the sorted liver metastasized trogocytic colon cancer cells.

first: Jae

last: Shin

first: Alfred

last: Bothwell


title: DLG2 knockout reveals neurogenic transcriptional programs underlying neuropsychiatric disorders and cognition

city: Cardiff

first: Eunju

state: Wales

address: Hadyn Ellis Building,Maindy Road.

department: Neuroscience & Mental Health Research Institute, School of Medicine.

institute: Cardiff University

summary: Brain development requires a complex choreography of cell proliferation, specialisation, migration and network formation, guided by the activation and repression of gene expression programs. It remains unclear how this process is disrupted in neuropsychiatric disorders. Here we integrate human genetics with transcriptomic data from the differentiation of human embryonic stem cells into cortical excitatory neurons. This reveals a cascade of transcriptional programs, activated during early corticoneurogenesis in vitro and in vivo, in which genetic variation is robustly associated with neuropsychiatric disorders and cognitive function. Within these early neurogenic programs, genetic risk is concentrated in loss-of-function intolerant (LoFi) genes, capturing virtually all LoFi disease association. Down-regulation of these programs in DLG2 knockout lines delays expression of cell-type identity alongside marked deficits in neuronal migration, morphology and action potential generation, validating computational predictions. These data implicate specific cellular pathways and neurodevelopmental processes in the aetiology of multiple neuropsychiatric disorders and cognition.

overall_design: mRNA profiles of DLG2-/- and WT lines cultured to days 15, 20, 30 and 60 of cortical differentiation

first: Andrew

last: Pocklington

first: Bret

last: Sanders

first: Daniel

last: D'Andrea


title: Mutations in LRRK2 linked to Parkinson’s disease sequester Rab8a to damaged lysosomes and regulate transferrin-mediated iron uptake in microglia

city: Bethesda

first: Mark

last: Cookson

address: NIHBC 35 - PNRC I BG RM 1A-116

country: USA

institute: National Institutue on Aging, NIH

summary:
Mutations in leucine-rich repeat kinase 2 (LRRK2) cause autosomal dominant Parkinson’s disease (PD) while polymorphic LRRK2 variants are associated with sporadic PD. PD-linked mutations increase LRRK2 kinase activity and induce neurotoxicity in vitro and in vivo. The small GTPase Rab8a is a LRRK2 kinase substrate and is involved in receptor-mediated recycling and endocytic trafficking of transferrin, but the effect of PD-linked LRRK2 mutations on the function of Rab8a is poorly understood. Here, we show that gain-of-function mutations in LRRK2 induce sequestration of endogenous Rab8a to lysosomes in over-expression cell models while pharmacological inhibition of LRRK2 kinase activity reverses this phenotype. Furthermore, we show that LRRK2 mutations drive association of endocytosed transferrin with Rab8a-positive lysosomes. LRRK2 has been nominated as an integral part of cellular responses downstream of proinflammatory signals and is activated in microglia in post-mortem PD tissue. Here, we show that iPSC-derived microglia from patients carrying the most common LRRK2 mutation, G2019S, mistraffic transferrin to lysosomes proximal to the nucleus in proinflammatory conditions. Furthermore, G2019S knock-in mice show a significant increase in iron deposition in microglia following intrastriatal LPS injection compared to wild type mice, accompanied by striatal accumulation of ferritin.
Conclusion: Our data support a role of LRRK2 in modulating iron uptake and storage in response to proinflammatory stimuli in microglia. Neuroinflammation remodels endolysosomal gene expression in microglia, in vitro and in vivo.

overall_design: For in vivo study, LPS striatal injections were administered to WT and LRRK2 KO mice, followed by microglia isolation from adult brain and single-cell RNA Seq.

first: Adamantios

last: Mamais

first: Jillian

last: Kluss

first: Luis

last: Bonet-Ponce

first: Natalie

last: Landeck

first: Rebekah

last: Langston

first: Nathan

last: Smith

first: Alexandra

last: Beilina

first: Alice

last: Kaganovich

first: Manik

middle: C

last: Ghosh

first: Laura

last: Pellegrini

first: Ravindran

last: Kumaran

first: Ioannis

last: Papazoglou

first: George

last: Heaton

first: Rina

last: Bandopadhyay

first: Nunziata

last: Maio

first: Changyoun

last: Kim

first: Matthew

last: LaVoie

first: David

last: Gershlick


title: KDM6B promotes activation of the oncogenic CDK4/6-pRB-E2F pathway by maintaining enhancer activity in MYCN-amplified neuroblastoma [CUT&TAG]

city: Memphis

first: Beisi

last: Xu

state: Tennessee

address: 262 Danny Thomas Pl

department: Center for Applied Bioinformatics

institute: St Jude Children's Research Hosipital

summary: The H3K27me2/me3 histone demethylase KDM6B is over-expressed in neuroblastoma and is essential to neuroblastoma cell survival. While the KDM6B inhibitor, GSK-J4, has shown activity in in vitro and in vivo preclinical models, the mechanism of action remains poorly defined. We demonstrate that genetic and pharmacologic inhibition of KDM6B downregulates the pRB-E2F transcriptome and MYCN expression. Chemical genetic analyses show that high expression of the E2F transcriptome is positively correlated with sensitivity of cancer cells to GSK-J4. Mechanistically, inhibition of KDM6B activity 1) reduces the chromatin accessibility of E2F target genes and MYCN, 2) selectively leads to an increase of H3K27me3 but a decrease of the enhancer mark H3K4me1 at the CTCF and BORIS binding sites, which may, consequently, disrupt the long-range chromatin interaction of MYCN and E2F target genes, and 3) phenocopies the transcriptome induced by the specific CDK4/6 inhibitor palbociclib. Overexpression of CDK4/6 or Rb1 knockout confers neuroblastoma cell resistance to both palbociclib and GSK-J4. A gene signature targeted by KDM6B inhibition is associated with poor survival of patients with neuroblastoma regardless of the MYCN status. These data indicate that KDM6B activity  promotes an oncogenic CDK4/6-pRB-E2F pathway in neuroblastoma cells via H3K27me3-dependent enhancer-promoter interactions, providing a rationale to target KDM6B for high-risk neuroblastoma.

overall_design: Examination of chromatin changes with KDM6B siRNA knockdown and GSK-J4 treatment.

first: Jie

last: Fang

first: Hongjian

last: Jin

first: Jun

last: Yang


title: Expression (Gene) in the endothelial cells of the mouse male prefrontal cortex (PFC) following 10-day chronic social stress exposure [male CSDS]

city: Quebec City

first: Caroline

last: Menard

state: QC

address: CERVO Brain Research Center, 2601 de la Canardiere

department: Psychiatry and Neuroscience

institute: Université Laval

summary: Cell-specific transcriptomic analysis were performed on PFC endothelial cells of unstressed controls (CTRL), stress-susceptible (SS) and resilient (RES) mice to identify novel genes and pathways involved in stress responses

overall_design:
Animals were subject to 10 days of chronic social defeat stress, phenotype using a social interaction test then the brains were collected 24h later and PFC punched for magnetic-activated cell sorting and transcriptomic analyses.
Genome Quebec (extracted RNA, assessed quality using the bioanalyzer, performed the Clariom S mouse assay according to the manufacturer's instructions).

first: Laurence

last: Dion-Albert

first: Manon

last: Lebel


title: Expression (Gene) in the endothelial cells of the mouse female prefrontal cortex (PFC) following 10-day chronic social stress or 6-day subchronic variable stress exposure was compared to male [Male_SS_vs_Female_SS_or_SCVS]

summary: Cell-specific transcriptomic analysis were performed on PFC endothelial cells of stress-susceptible (SS) and resilient (RES) mice along with females subjected to 6 days of subchronic variable stress to identify novel genes and pathways involved in stress responses. These gene lists were compared to published findings obtained in the male NAc following 10 days of chronic social stress exposure (Dudek et al., 2020)

overall_design:
Animals were subject to either 10 days of chronic social defeat stress and phenotype deteremined using a social interaction test or 6 days of subchronic variable stress then the brains were collected 24h later and PFC punched for magnetic-activated cell sorting and transcriptomic analyses. Nac gene lists produced in Dudek et al. PNAS (2020)
Genome Quebec (extracted RNA, assessed quality using the bioanalyzer, performed the Clariom S mouse assay according to the manufacturer's instructions).


title: Sex-specific blood-brain barrier alterations and vascular biomarkers underlie chronic stress responses in mice and human depression.

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

overall_design: Refer to individual Series


title: SAGA core module subunits regulate gene expression during oogenesis [ChIP-seq]

city: Kansas City

first: Hua

last: Li

state: MO

address: 1000 E 50th Street

institute: Stowers Institute for Medical Research

summary: ChIP-nexus performed in Kc167 cells for subunits of the SAGA coactivator complex;ChIP performed in ovary homogenate for subunits of the SAGA coactivator complex

overall_design: Genome-wide binding profiles ofhte SAGA complex were establised in cells and ovary tissue using ChIP-nexus and reguar ChIP by chromatin immunoprecipitating Ada2b, Spt3, SAF6 and WDA, and Sgf11

first: Jelly

last: Soffers

first: Sergio

last: Alcantara

first: Christopher

middle: W

last: Seidel

first: Julia

last: Zeitlinger

first: Jerry

last: Workman


title: Single cell transcriptomics captures features of developing and mature DA  neurons in human brain organoids and reveals more precise and reproducible patterning in silk-bioengineered culture

city: Lund

first: Petter

last: Storm

address: Sölvegatan 19, BMC A11

department: Developmental and Regenerative Neurobiology

institute: Lund University

summary: The inaccessibility of human tissue and the difficulty to achieve neuronal maturation and function in 2D cultures makes the study of human brain development, function and disease  challenging. Here, we differentiated human pluripotent stem cells into three-dimensional (3D) regionalized human brain organoids which, when patterned towards a ventral midbrain (VM) fate, gave rise to mature and functional pigmented dopamine (DA) neurons. Using single cell transcriptomics, we defined the cellular composition and reconstructed the developmental trajectory of the different cell types, including DA neurons, by analyzing 120,000 cells harvested at different time points during differentiation. We also obtained an unbiased and comprehensive characterization of human DA subgroups, which reveals a molecular signature of DA neuron diversity and provides a valuable asset for the design of more targeted and effective stem-cell based therapies in Parkinson´s disease. However, the value of brain organoids in modeling later developmental stages and more mature neurons in a dish is hampered by their poor reproducibility and incomplete maturation. To address this issue, we designed a novel technological approach to generate bioengineered VM organoids supported by recombinant spider silk microfibers functionalized with full-length human laminin. Silk-VM organoids reproduce key molecular aspects of DA neurogenesis and, unlike conventional 3D culture, sustain the generation of functional DA neurons homogeneously throughout the organoid. Silk-VM organoids represent a significant advancement in the field of 3D culture, by better recapitulating physiologically relevant aspects of developing human brain tissue and thus paving the way toward their use in developmental studies as well as in in vitro disease modeling and drug discovery.

overall_design:
Single-cell RNA-seq of midbrain patterned organoids
NOTE: We have patient derived samples and can not share the raw sequence data due to GDPR concerns.

first: Alessandro

last: Fiorenzano

first: Malin

last: Parmar


title: Expression (Gene) in the endothelial cells of the mouse female prefrontal cortex (PFC) following 10-day chronic social stress or 6-day subchronic variable stress exposure [Female_CSDS_vs_SCVS]

summary: Cell-specific transcriptomic analysis were performed on PFC endothelial cells of stress-susceptible (SS) and resilient (RES) mice along with females subjected to 6 days of subchronic variable stress to identify novel genes and pathways involved in stress responses

overall_design:
Animals were subject to either 10 days of chronic social defeat stress and phenotype deteremined using a social interaction test or 6 days of subchronic variable stress then the brains were collected 24h later and PFC punched for magnetic-activated cell sorting and transcriptomic analyses.
Genome Quebec (extracted RNA, assessed quality using the bioanalyzer, performed the Clariom S mouse assay according to the manufacturer's instructions).


title: Expression (Gene) in the endothelial cells of the mouse female prefrontal cortex (PFC) following 6-day subchronic variable stress (SCVS) exposure [Female_SCVS]

summary: Cell-specific transcriptomic analysis were performed on PFC endothelial cells of unstressed controls (CTRL) and mice exposed to SCVS to identify novel genes and pathways involved in stress responses

overall_design:
Animals were subject to 6 days of subchronic variable stress then the brains were collected 24h later and PFC punched for magnetic-activated cell sorting and transcriptomic analyses.
Genome Quebec (extracted RNA, assessed quality using the bioanalyzer, performed the Clariom S mouse assay according to the manufacturer's instructions).


title: Expression (Gene) in the endothelial cells of the mouse female prefrontal cortex (PFC) following 10-day chronic social stress exposure [Female_CSDS]