title: MicroRNA profiling of extracellular vesicles isolated from rhesus macaques vaccinated with DNA/ALVAC/gp120 vaccine

summary: We contrasted innate and adaptive immune responses elicited by the DNA/ALVAC/gp120/alum vaccine in non-human primates differing in age. We performed several analyses. MicroRNA profiling in Extracellular Vesicles (EVs) isolated from plasma collected prior and 1 week following vaccination identified 109 miRNAs that differed significantly between age groups at the end of immunization. Several miRNAs correlated with risk of virus acquisition in young and old animals, but none remained significant following correction for FDR. In young animals, we identified 3 miRNAs which were modified by vaccination and correlated with decreased risk of SIV acquisition (unadjusted p-values). Of these three miRNAs, two, miR-139-5p and miR-29b-1-5p, were increased by vaccination, whereas one, mir 98, decreased by vaccination. MiR-139-5p has several validated targets, including: CXCR4, important for egress of CXCR4+ cells and monocytes from bone marrow and HIV entry in target T-cells; the cAMP-specific PDE4D gene which decreases CREB1-mediated transcription by degrading cAMP; and the IGF-1/AKT/Glut1 gene receptor, central for glucose utilization and cellular metabolism.  The validated target of miR-29b-1-5p is the NF-kB pathway. Mir-98 has several validated targets, inhibits IL-10 production, modulates M1-M2 polarization of macrophages towards the M1 phenotype, and its CCL2-induced downregulation increases IL-6 production. These data suggest the importance of CREB1-mediated transcription in vaccine efficacy, identified in the study, is likely assisted by miR-139-5p targeting the cAMP-degrading PDE4D gene, and downregulation of the NF-kB pathway, targeted by miR-29b-1-5p.

overall_design: Forty juvenile (young, average age 3.45 years, 0.545 SD) and twenty-eight adult (old, average age 7.98 years and 2.60 SD) macaques were assigned to four groups based on their major histocompatibility status, age, and weight. Thirteen young and seventeen old macaques were immunized twice with DNA-SIV at weeks 0 and 4. At weeks 8 and 12 all 30 macaques were boosted with intramuscular inoculations of 10^8 Plaque Forming Units (PFU) of recombinant ALVAC (vCP2432). At week 12, vaccinated macaques also received 200 μg each of SIVmac251 – M766 and SIVsmE660 – CG7V gp120-gD proteins formulated in alum Alhydrogel. Plasma collected at baseline from 7 young animals and 1 week following vaccination (week 13) from all vaccinated animals was used for extracellular vesicles (EVs) isolation. EVs were isolated from plasma as described previously (Thery, C.et al.. Isolation and characterization of exosomes from cell culture supernatants and biological fluids. Curr Protoc Cell Biol Chapter 3, Unit 3 22 (2006)) with minor modifications. EVs isolated from plasma were processed by Exiqon, Inc. (Woburn, MA). The RNA was isolated from EVs by following a proprietary RNA isolation protocol. MicroRNA was used to generate the libraries and was subsequently sequenced on a NextSeq500 sequencing instrument (Illumina, Inc.).

first: Veronica

last: Galli

first: Anush

last: Arakelyan

first: Alexei

last: Lobanov

first: George

middle: W

last: Nelson

first: Margaret

last: Cam

first: Leonid

last: Margolis


title: RNA-Seq of mouse CD69+ regulatory T cells (Treg) in venous blot clots

status: Public on Jan 06 2023

city: Mainz

first: Matthias

last: Klein

address: Langenbeckstrasse 1

department: Inst. for Immunology

country: Germany

institute: University Medical Center Mainz

type: Expression profiling by high throughput sequencing

summary: Transcriptional profiling shows that Treg in venous thrombi take on a repair Treg profile and produce the matricellular protein SPARC

overall_design: CD45+CD4+EGFP[Foxp3]+CD69+ Treg were flow-cytometrically sorted from the spleens, thrombi and vein wall tissue of thrombotic (Inferior Vena Cava)-ligated C57BL/6-Tg(Foxp3-DTR/EGFP)23.2Spar/Mmjax (DEREG) mice at day 14 after thrombosis induction.

first: Fatemeh

last: Zare-Shaneh

first: Christian

last: Becker


title: Human variation impacting MCOLN2 restricts Salmonella Typhi replication by magnesium deprivation

status: Public on Jan 05 2023

city: Durham

first: Liuyang

last: Wang

state: NC

address: 213 Research Drive

country: USA

institute: Duke University

summary: Human genetic diversity can reveal critical factors in host-pathogen interactions. This is especially useful for human-restricted pathogens like Salmonella enterica serovar Typhi (S. Typhi), the cause of Typhoid fever. One key dynamic during infection is competition for nutrients: host cells attempt to restrict intracellular replication by depriving bacteria of key nutrients or delivering toxic metabolites in a process called nutritional immunity. Here, a cellular genome-wide association study of intracellular replication by S. Typhi in nearly a thousand cell lines from around the world—and extensive follow-up using intracellular S. Typhi transcriptomics and manipulation of magnesium concentrations—demonstrates that the divalent cation channel mucolipin-2 (MCOLN2) restricts S. Typhi intracellular replication through magnesium deprivation. Our results reveal natural diversity in Mg2+ limitation as a key component of nutritional immunity against S. Typhi.

overall_design: To investigate how Salmonella enterica Typhi impacts human transcriptome profile, dual RNA-seq of human and pathogen are performed on THP-1 cell line and MCLON2 -/- THP-1 cell line.  THP-1 cells and MCLON2 -/- THP-1 cells were infected with either  S. Typhi grown in tubes to late log phase or the intracellular S. Typhi from infected THP-1.

first: Kyle

middle: D

last: Gibbs

first: Dennis

middle: C

last: Ko


title: RNA profiles of cells in the lungs of MuHV-4 infected mice.

city: Liège

first: Bénédicte

last: Machiels

state: Liège

address: Avenue de Cureghem 7b, B43b

department: DMI

country: Belgium

institute: Université de Liège

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

overall_design: Refer to individual Series


title: Comparison of transcriptomic profiles of macrophages cultures without ILC2, in presence of ILC2 from uninfected mice, and in presence of ILC2 from MuHV-4 infected mice.

summary: The immunology of asthma is complex and involves the loss of immunoregulatory pathways with the adoption of a Western lifestyle inducing changes in the diversity and the time exposure to microorganisms. Gammaherpesviruses (γHVs) are among the most prevalent human viruses. Establishing early life infection, they profoundly imprint the immune system of their hosts. Using Murid herpesvirus 4 (MuHV-4), a mouse model of human γHV infections, we showed that both lung resident and bone marrow-derived group 2 innate lymphoid cells (ILC2s) displayed lasting reduced capacity to expand and to produce type 2 cytokines, in response to house dust mites, in an IFN-γ depend manner. Importantly, we uncovered that pulmonary ILC2s represent essential niche cells that imprint alveolar macrophage tissue-specific identity upon monocyte replenishment in infection settings. In particular, we showed that MuHV-4 infection disrupts the normal ILC2-epithelial cells circuit that imprints monocytes-derived AMs for type 2 functions without affecting their differentiation. These results reveal that persistent γHV infection shapes the alveolar landscape much beyond the initial acute infection through long-term effect on ILC2 niche cells.

overall_design: Comparative gene expression profiling analysis of RNA-seq data of macrophages, in presence or not of ILC2 from uninfected and MuHV-4 infected mice.

first: Pauline

last: Loos

first: Jérôme

last: Baiwir

first: Céline

last: Maquet

first: Justine

last: Javaux

first: Rémy

last: Sandor

first: François

last: Lallemand

first: Thomas

last: Marichal

first: Laurent

last: Gillet


title: Gene expression profile at single cell level of cells from the lungs of Mock and MuHV-4 infected mice after HDM challenge.

overall_design: Cells from lungs of non-infected and MuHV-4 infected mice subjected to different doses of HDM (0, 1, 3) were collected.


title: RNA sequencing of mouse dendritic cells in Aire-reporter strain

city: Kuramoto, Tokushima

first: Minoru

last: Matsumoto

department: Department of Molecular Pathology

country: Japan

institute: Tokushima University Graduate School of Biomedical Sciences

summary: Bulk RNA-seq analysis of mouse dendritic cells (DCs) in Aire-reporter strain (AGF-KI: human AIRE and GFP-Flag tag knock-in). Aire+ DCs and Aire- DCs were isolated from lymph nodes and bone marrow, using Aire-sufficient (+/KI) and Aire-deficient background (-/KI).

overall_design: AGF+ and AGF− LNDCs were isolated from lymph node cells gated for CD45+CD11c+MHCII+ fraction. AGF+ and AGF- BMDCs were isolated from bone marrow cells gated for B220−CD24+CD11b−/low fraction and B220−CD11c+MHCII+ fraction, respectively. All samples were isolated from 6 weeks old male mice on C57BL/6 strain.

first: Ryuichiro

last: Miyazawa

first: Jun-ichi

last: Nagao

first: Ken-ichi

last: Arita-Morioka

first: Junko

last: Morimoto

first: Masaki

last: Yoshida

first: Takeshi

last: Oya

first: Koichi

last: Tsuneyama

first: Hideyuki

first: Yoshihiko

last: Tanaka

first: Mitsuru


title: A modERN Resource: Identification of Drosophila Transcription Factor candidate target genes using RNAi

city: Berkeley

first: Benjamin

middle: Wendell

last: Booth

state: CA

address: 1 Cyclotron Road

department: BioSciences

institute: Lawrence Berkeley National Laboratory

summary: Transcription factors (TFs) play a key role in development and in cellular responses to the environment by activating or repressing the transcription of target genes in precise spatial and temporal patterns.  In order to develop a catalog of target genes of D. melanogaster transcription factors, the modERN consortium systematically knocked down expression of transcription factors (TFs) using RNAi in whole embryos followed by RNA-seq. We generated data for 45 TFs which have 18 different DNA-binding domains and are expressed in 15 of the 16 organ systems.  The range of inactivation of the targeted TFs by RNAi ranged from log2fold change -3.52 to +0.49. The TFs also showed remarkable heterogeneity in the numbers of candidate target genes identified, with some generating thousands of candidates and others only tens.  We present detailed analysis from six experiments, including those for four TFs that have been the focus of previous functional studies (ERR, sens, su(Hw) and zfh2) and two previously uncharacterized TFs (sens-2 and CG32006), as well as short vignettes from selected additional experiments to illustrate the utility of this resource. Transcription Factor and target gene expression patterns can be found here: https://insitu.fruitfly.org. These studies provide data that facilitate scientific inquiries into the functions of individual TFs in key developmental, metabolic, defensive, and homeostatic regulatory pathways, as well as provide a broader perspective on how individual TFs work together in local networks during embryogenesis.

overall_design: Transcription factors were knocked down using RNAi, and the effect of gene expression was evaluated by paired-end RNA-seq in replicate.

first: William

last: Fisher

first: Ann

middle: S

last: Hammonds

first: Richard

last: Weiszmann

first: Jaeda

last: Patton

first: Connor

last: Kubo

first: Robert

middle: H

last: Waterston

first: Susan

middle: E

last: Celniker


title: AKT1-FOXO4 Axis Regulates Hemochorial Placentation [Foxo4-KD-rTSC]

city: Stillwater,

first: Khursheed

last: Iqbal

state: OK

address: 310 N Monroe St,

institute: Oklahoma state university

summary: AKT1 is a serine/threonine kinase implicated in fetal, placental, and postnatal growth. In this study, we investigated roles for AKT1 in placental development using a genome-edited/loss-of-function rat model. Both heterozygous and homozygous Akt1 mutant rats were viable and fertile. Disruption of AKT1 resulted in placental, fetal, and postnatal growth restriction. Akt1 null placentas showed deficits in both junctional zone and labyrinth zone size and their ability to adapt to a physiological stressor. Robust differences in the transcriptome of wild type versus Akt1 null junctional zones were identified. Among the differentially expressed junctional zone transcripts was forkhead box O4 (Foxo4), which encodes a transcription factor and known AKT substrate. FOXO4 expression was prominent in the junctional zone and invasive trophoblast cells of the rat placentation site and enhanced following rat TS cell differentiation. Foxo4 gene disruption using genome-editing resulted in placentomegaly, including an enlarged junctional zone. AKT1 and FOXO4 regulate the expression of many of the same transcripts expressed by trophoblast cells; however, in opposite directions. In summary, we have identified AKT1 and FOXO4 as part of a regulatory network controlling hemochorial placenta development.

overall_design: 1) Rat placentation site compartments were dissected on gd18.5. Dissected junctional zone tissues from wild type, Akt1 null, and Foxo4Xm- fetuses were snap-frozen in liquid nitrogen and stored at -80°C until processing for RNA isolation. 2) Foxo4 was knocked down in rat trophoblast stem cells using lentiviral delivery of shRNA. Following selection by puromycin, cells were differentiated for fifteen days. On day fifteen of differentiation, cells treated with either control shRNA or Foxo4-specific shRNA were collected and RNA was isolated.

first: Keisuke

last: Kozai

first: Ayelen

last: Moreno

first: Michael

middle: J

last: Soares


title: Gene expression profile at single cell level of cells from the lungs of Mock and MuHV-4 infected mice.

first: Bénédicte

last: Machiels

overall_design: Cells from lungs of non-infected and MuHV-4 infected (5, 8, 14, and 28 d.p.i.) mice were collected.

first: Pauline

last: Loos

first: Jérôme

last: Baiwir

first: Céline

last: Maquet

first: Justine

last: Javaux

first: Rémy

last: Sandor

first: François

last: Lallemand

first: Thomas

last: Marichal

first: Laurent

last: Gillet


title: AKT1-FOXO4 Axis Regulates Hemochorial Placentation


title: Type I IFN stimulates lymph node stromal cells from adult and old mice during a West Nile virus infection

city: Chicago

first: Allison

last: Bennett

state: IL

address: 835 S WOLCOTT AVE

institute: University of Illinois at Chicago

summary: Advanced age is a significant risk factor during viral infection due to an age-associated decline in the immune response. While infection by West Nile virus (WNV) is typically mild, older individuals are at an increased risk of severe neuroinvasive disease. Previous studies have characterized age-associated defects in hematopoietic immune cells during WNV infection that culminate in diminished antiviral immunity. Situated amongst immune cells in the draining lymph node (DLN) are structural networks of nonhematopoietic lymph node stromal cells (LNSCs). LNSCs are comprised of numerous, diverse subsets, whose roles in the coordination of robust immune responses are underappreciated. Like lymphocytes, LNSCs mount tailored responses to viral pathogens and are susceptible to aging. However, the contributions of LNSCs to WNV immunity and immune senescence are unclear. Here, we examine LNSC responses to WNV infection within adult and old DLNs. Acute WNV infection was shown to trigger cellular infiltration and LNSC expansion. Comparatively, aged DLNs exhibited diminished leukocyte accumulation and altered LNSC subset composition. Upon recognition of WNV, adult and old LNSCs were activated in a distinct, type I interferon-driven mechanism that promoted antiviral gene expression. Aged LNSCs were found to constitutively upregulate immediate early response genes, several of which are associated with immune suppression. Collectively, these data suggest LNSCs uniquely respond to WNV infection via a novel activation mechanism. We are the first to report age-associated differences in LNSCs on the population- and gene expression-level during WNV infection. These changes may compromise antiviral immunity, leading to increased WNV disease in older individuals.

overall_design: Bulk RNA sequencing (RNA-seq) was performed on adult and old lymph node stromal cells (LNSCs) stimulated with viral inflammatory stimuli for 20 hours to determine differentially expressed transcripts in activated stromal cells. Draining lymph nodes from adult (8-10 weeks) and old (18 month) C57BL/6 mice were digested and plated. LNSCs were then enriched for using CD45-labeled microbeads to magnetically deplete unwanted CD45+ cells. Cultures of adult and old LNSCs were incubated for 20 hours with supernatant from bone marrow derived dendritic cells infected with West Nile Kunjin virus ("BMDC"), recombinant IFNa (2 ng/mL, "Recomb"), or were untreated. RNA was extracted from LNSCs using the RNeasy Mini Kit (QIAGEN) and DNase treated. RNA was quantified using the Qubit™ RNA HS Assay Kit (Invitrogen) on the Quantus Fluorometer (Promega). All RNA samples had a RIN score above 8. Total RNA samples were normalized to 50 ng and  libraries were constructed using the Universal Plus mRNA-Seq kit (Tecan), followed by RNA-seq on the Illumina NovaSeq6000 S4 flow cell, obtaining 150bp paired-end reads. Raw RNA-seq reads were assessed using FastQC, aligned to mouse reference genome mm10 with STAR, and quantified using featureCounts. Gene expression was normalized to counts per million units and averaged across sample groups, followed by principal component analysis to assess sample clustering. Differential gene analysis was performed in edgeR.

middle: K

first: Justin

middle: M

last: Richner


title: Plastin 3 rescues deficient TrkB/BDNF signaling and protects from dysregulations in Ca2+ homeostasis in SMA mouse models

city: Würzburg

first: Silke

last: Appenzeller

address: Schweinfurter Str. 28

institute: University Wuerzburg

summary: Spinal muscular atrophy (SMA) is a lethal pediatric neuromuscular disease caused by loss of the Survival Motor Neuron 1 gene (SMN1). Although current therapies focus very efficiently on SMN restoration, the cellular mechanisms underlying the pathological features are not fully understood. Smn-deficient motoneurons tremendously suffer from functional abnormalities leading to affected motoneuron differentiation and maturation. Based on our study, we could illustrate disturbed F-actin-dependent translocation of the Tropomyosin-kinase receptor B (TrkB) to the cell surface of axonal terminals of Smn-deficient motoneurons. This in turn results in impaired Brain-derived neurotrophic factor-induced TrkB activation and downstream signaling. Focusing on the F-actin bundling properties of the protective SMA modifier Plastin 3 revealed that its restoration significantly compensates altered TrkB surface recruitment and activation and ameliorates Cav2.1/2 cluster formation and cellular excitability in Smn-deficient motor axons. Thus, Plastin 3 supports in general two major cellular mechanisms indispensable for development and functional maintenance of motoneurons.

overall_design: Total number of samples is 12

first: Luisa

last: Hennlein

first: Hanaa

last: Ghanawi

first: Florian

last: Gerstner

first: Eduardo

last: Palominos-Garcia

first: Lena

last: Saal-Bauernschubert

first: Chunchu

last: Deng

first: Mehri

last: Moradi

last: Briese

first: Christian

last: Simon

last: Sendtner

first: Sibylle

last: Jablonka


title: Elongation Factor P is important for sporulation initiation

status: Public on Jan 04 2023

city: Baltimore

first: Allen

middle: R

last: Buskirk

state: MD

address: 725 N. Wolfe St

department: Molecular Biology and Genetics

institute: Johns Hopkins University School of Medicine

summary: The universally conserved protein Elongation Factor P facilitates translation at amino acids that form peptide bonds with low efficiency, particularly poly-proline tracts. Despite its wide conservation, it is not essential in most bacteria and its physiological role remains unclear. Here, we show that EF-P affects the process of sporulation initiation in the bacterium Bacillus subtilis. We observe that lack of EF-P delays expression of sporulation-specific genes. Using ribosome profiling, we observe that expression of spo0A, encoding a transcription factor that functions as the master regulator of sporulation, is lower in a ∆efp strain as compared to the wildtype. Ectopic expression of Spo0A rescues the sporulation initiation phenotype, indicating that reduced spo0A expression explains the sporulation defect in ∆efp cells. Since Spo0A is the earliest sporulation transcription factor, these data suggest that sporulation initiation can be delayed when protein synthesis is impaired.

sample_organism: Bacillus subtilis subsp. subtilis str. 168

overall_design: Ribosome profiling of wild-type B. subtilis and a strain lacking EFP 15 and 60 minutes after induction of sporulation


title: Characterization of DkaiC mutants of Pseudomonas putida KT2440/2 and Pseudomonas protegens CHA0

city: Neuchâtel

first: Diego

last: Gonzalez

address: Rue Emile-Argand

country: Switzerland

institute: Laboratory of Microbiology, University of Neuchâtel

summary: KaiC is the central cog of the circadian clock in Cyanobacteria. Close homologs of this protein are widespread among bacteria not known to have a circadian physiology. The function, interaction network, and mechanism of action of these KaiC homologs are still largely unknown. Here, we focus on KaiC homologs found in environmental Pseudomonas species.  We characterize experimentally the only KaiC homolog present in Pseudomonas putida KT2440 and Pseudomonas protegens CHA0. Through phenotypic assays and transcriptomics, we show that KaiC is involved in osmotic and oxidative stress resistance in P. putida and in biofilm production in both P. putida and P. protegens.

sample_organism:
Pseudomonas putida
Pseudomonas putida KT2440
Pseudomonas protegens CHA0

overall_design: Comparison of the transcriptome of the wild-type and DkaiC mutant of Pseudomonas putida KT2440/2  grown in LB to early, mid, and late stationary phase. Comparison of the transcriptome of the wild-type and DkaiC mutant of Pseudomonas protegens CHA0 grown in LB to late stationary phase.

first: Céline

last: Terrettaz

first: Bruno

last: Cabete


title: The molecular principles governing HCMV infection outcome

city: Rehovot

first: Noam

last: Stern-Ginossar

address: 234 herzel

department: Molecular Genetics

country: Israel

institute: Weizmann Institute

summary: Infection with Human cytomegalovirus (HCMV) can result in either productive or non-productive infection, the latter potentially leading to establishment of latency, but the molecular factors that dictate these different infection outcomes are elusive. Macrophages are known targets of HCMV and considered to be permissive for productive infection, while monocytes, their precursors, are thought to support latent infection. Here we reveal that infection of macrophages is more complex than previously appreciated and can result in either productive or non-productive infection. By analyzing the progression of HCMV infection in c and macrophages using single cell transcriptomics, we uncover that the key characteristics that define productive and non-productive cells are the onset of viral gene expression, and activation of Interferon-stimulated genes (ISGs), respectively. We show that the level of viral gene expression, and specifically the expression of the major immediate early factor, IE1, is the principle barrier for establishing productive infection. On the cellular side, induction of ISGs in response to infection surprisingly does not dictate infection outcome, but we find that the cell intrinsic ISG levels is a main determinant of infection outcome. Indeed, intrinsic ISG level is downregulated with monocyte differentiation. We further show that, compared to monocytes, non-productive macrophages maintain slightly higher levels of viral transcripts and are able to reactivate, raising the possibility that they can serve as latency reservoirs in tissues. Moreover, we find that productive infection perturbs macrophage identity and function, likely affecting their immunological role during active infection. Overall, by harnessing the tractable system of monocyte differentiation we decipher the underlying principles that control HCMV infection outcome, and propose macrophages as a new potential HCMV reservoir in tissues.

overall_design: The MARS-seq protocol was used for single cell RNA-seq of primary CD14+ monocytes and PMA-induced monocyte-derived macrophages infected with TB40/E-GFP strain of HCMV at 5, 12 ,24,48, 72 and 144 hours post infection (Experiment 1) and of PMA-induced monocyte-derived macrophages infected with TB40/E-GFP strain of HCMV at 24, 72 and 144 hours post infection, for which at 72 and 144 hours post infection, only GFP-dim cells were sorted for single cell RNA-seq (Experiment 2). 10X Genomics Chromium Single Cell 3? Gene Expression v3.1 kit was used  for  scRNA-seq of BAL cells, infected with TB40/E-GFP strain of HCMV at 20 and 70 hours post infectionpools (Experiment 3). A modified mcSCRB-seq was used for RNA-seq of BAL macrophages infeced with TB40/E-GFP strain of HCMV at 3 days post infection sorted to GFP-dim and GFP-bright cells (Experiment 4). Bulk RNA-seq libraries were prepared according to Shishkin et al. Nat. Methods 2015, for THP1 cells following CRISPR KO of IRF9 or STAT2 or IGSF8 (Experiment 5).

first: Miri

last: Shnayder

first: Michal

last: Schwartz

first: Aharon

last: Nachshon

first: Yaarit

last: Kitsberg

first: Tamar

last: Arazi


title: Single cell RNA-sequencing analysis of islets from WT NOD mice on different diets

first: Alexander

last: Chervonsky

department: Pathology

institute: University of Chicago

summary: Protein source in diet greatly influences the incidence of type-1 diabetes (T1D) in non-obese diabetic (NOD) mouse colonies.  NOD mice fed a diet containing hydrolyzed casein (HC) as the sole protein source are protected from T1D. Replacing 1/5th of the HC with gluten restores high T1D to NOD mice. We hypothesized that gluten might promote inflammation in the islets. We used single cell RNA sequencing (scRNAseq) to characterize and contrast the transcriptional profiles of endocrine cells and islet infiltrating leukocytes from the pancreatic islets of Langerhans of NOD mice fed a 20% HC diet or a 16% HC + 4% gluten diet.

overall_design: scRNAseq was performed on single cell suspensions from isolated pancreatic islets of Langerhans from 15 week old NOD/ShiLtJ mice fed either 20% HC or 16% HC + 4% gluten diets. Islets from 3 mice per diet were pooled for each sample.

first: Matthew

last: Funsten

middle: V

first: Aly

middle: A

last: Khan

first: Derek

last: Reiman


title: Transcriptomic and proteomic profiling of young and old mice in the Bleo model

city: Biberach

first: Sergio

last: Picart-Armada

address: Birkendorfer Straße 65

institute: Boehringer Ingelheim

type: Non-coding RNA profiling by high throughput sequencing

summary: The most preclinical used in vivo model to study lung fibrosis is the bleomycin-induced lung fibrosis model in 2-3-month-old mice. Although this model resembles key aspects of idiopathic pulmonary fibrosis (IPF), there are limitations in its predictability for the human disease. One of the main differences is the juvenile age of animals that are usually used in experiments, resembling humans of around 20 years. Because IPF patients are usually older than 60 years, aging appears to play an important role in the pathogenesis of lung fibrosis.  Therefore, we here compared young (3 months) and old mice (21 months) 21 days after intratracheal bleomycin instillation. Analyzing lung transcriptomics (mRNAs & miRNAs) and proteomics, we found most pathways to be similarly regulated in young and old mice. However, old mice show an imbalanced protein homeostasis as well as an increased inflammatory state in the fibrotic phase compared to young mice. Comparisons with published human transcriptomic data sets (GSE47460, GSE32537 and GSE24206) revealed that the gene signature of old animals correlates significantly better with IPF patients as well as it turned human healthy individuals better into “IPF patients” using an approach termed predictive disease modelling.   Comprising, young and old animals show similarly molecular hallmarks of IPF. Additionally, old mice resemble several features associated with IPF more closely compared to young animals.

overall_design: Male mice were randomized before the study. Mice were instilled intratracheally (i.t.) under 3-4% isoflurane anesthesia once on day 0. Bleomycin administration was performed in a hanging position with a Vasofix-Braunüle 22G (B.Braun, Melsungen, Germany). Mice were instilled using 0.7 and 0.63 mg/kg of bleomycin sulphate (Merck, Darmstadt, Germany) in saline solution for the young and old mice, respectively. Control mice received saline solution only. Mice were analyzed 21 days after bleomycin administration.

first: Stephan

last: Klee

first: Kathrin

last: Wenger

first: Gerald

last: Birk

first: Karsten

last: Quast

first: Daniel

last: Veyel

first: Wolfgang

last: Rist

first: Coralie

last: Violet

first: Andreas

last: Luippold

last: Haslinger

last: Thomas

first: Francesc

last: Fernandez-Albert

first: Marc

last: Kästle


title: Single cell RNA-sequencing analysis of islets from NOD.SCID mice on chow and hydrolyzed casein diets

last: Chervonsky

summary: Protein source in diet greatly influences the incidence of type-1 diabetes (T1D) in non-obese diabetic (NOD) mouse colonies.  NOD mice fed a diet containing hydrolyzed casein (HC) as the sole protein source are protected from T1D.  Exposure to HC also reduced beta-cell insulin secretion during glucose challenge suggesting that beneficial changes to beta-cell physiology might protect NOD mice from T1D. We used single cell RNA sequencing (scRNAseq) to contrast the transcriptional profiles of beta-cells and from the pancreatic islets of Langerhans of NOD mice with the scid mutation (scid mice were used to avoid confouding influence of infiltrating leukocytes) fed a 20% HC diet or a standard chow diet.

overall_design: scRNAseq was performed on single cell suspensions from isolated pancreatic islets of Langerhans from 8-10 week old NOD.Cg-Prkdcscid/J mice fed either 20% HC standard chow diets. 3 chow fed mice and 2 HC fed mice were sequenced individually.


title: Gene expression profiles at the single cell level of CD8+ T cells from the spleens of LCMV-Armstrong or LCMV-Clone 13 infected mice

city: La Jolla

first: Gene

last: Yeo

address: 9500 Gilman Dr

department: Cellular and Molecular Medicine

institute: University of California, San Diego

summary: The transcriptomes of CD8+ T cells from LCMV-Armstrong and LCMV-Clone 13 infected mice are known to be distinct from one another. We used single cell RNA sequencing (scRNA-seq) to analyze the transcriptomic diversity of splenic CD8+ T cells in these two infection conditions at various timepoints after infection.

overall_design: Adoptively transferred CD8+ P14 T cells from the spleen of C57BL/6 mice were isolated by Fluorescence-activated cell sorting (FACS) according to the presence of CD45.1 congenic signal and analyzed using scRNA-seq.

first: Lauren

last: Quezada

first: Wenhao

last: Jin

first: Zhaoren

last: He


title: Areas of H3K27 tri-methylation in CD8+ T cells from the spleens of LCMV-Armstrong or LCMV-Clone 13 infected mice

type: Genome binding/occupancy profiling by high throughput sequencing

summary: The profiles of H3K27 tri-methylation in  CD8+ T cells from LCMV-Armstrong and LCMV-Clone 13 infected mice are known to be distinct from one another. We used CUT&RUN (Cleavage Under Targets and Release Using Nuclease) to analyze these differences in splenic CD8+ T cells of these two infection conditions.

overall_design: Adoptively transferred CD8+ P14 T cells from the spleen of C57BL/6 mice were isolated by Fluorescence-activated cell sorting (FACS) at day 7 post-infection according to the presence of CD45.1 congenic marker signal and analyzed using CUT&RUN.


title: RNA-Seq of storage cells of tardigrades Ramazzottius varieornatus and Hypsibius exemplaris

city: Tsuruoka

first: Kazuharu

last: Arakawa

state: Yamagata

address: Nihonkoku 403-1, Daihoji

institute: Institute for Advanced Biosciences, Keio University

summary: Tardigrades contain characteristic free-floating body cavity cells that are also called storage cells. To investigate the cell-specific expression patterns of tardigrade-unique genes, the expression levels of dissected storage cells are compared to those of the whole body.

sample_organism:
Ramazzottius varieornatus
Hypsibius exemplaris

overall_design: RNA-Seq has been performed on active whole body (N=3) and extracted storage cells (N=5) from two tardigrades, Ramazzottius varieornatus and Hypsibius exemplaris.

first: Sae

last: Tanaka


title: Full-length RNA-seq of short RNAs from Cuscuta campestris

status: Public on Jan 03 2023

city: University Park

first: Michael

last: Axtell

state: PA

address: 280 Mueller Lab

department: Biology

institute: Pennsylvania State University

summary: Cuscuta campestris is an obligate parasitic plant that attaches to the stems of host plants to obtain water and nutrients. C. campestris produces a specialized set of microRNAs specifically at the host-parasite interface. Many of these interface-induced microRNAs target mRNAs from the host. This experiment was designed to capture full-length primary transcripts that give rise to C. campestris interface-induced microRNAs. C. campestris shoot tips were stimulated to produce haustoria, then harvested to extract total RNA. RNA was then treated with various enzymes to modify 5' ends. Libraries were then made and sequenced. Importantly the library method only captures RNAs with a 5'-monophosphate. Therefore the specific enzymatic treatments can reveal the native 5' ends of the sequenced RNAs. Data were used to identify the primary transcripts for many of the known C. campestris primary microRNA transcripts. The data were also interrogated to tally specific snRNAs (U1, U2, U4, and U5) that are known to have a 2,2,7mGppp cap, and the tally pre-tRNAs, which are known to have a ppp 5' end. The analyzed results indicated that C. campestris interface-induced microRNA primary transcripts likely have a 5' ppp, not a cap, consistent with transcription by RNA polymerase III. The analysis also demonstrated that C. campestris interface-induced microRNA primary transcripts mostly terminate within stretches of polyT and lack polyA tails, also features consistent with Pol III transcription. Finally, the analysis indicated that the transcriptional start sites were located ~ 55 bp downstream of a common ten-base pair promoter element (the upstream sequence element). The 55bp spacing is also consistent with Pol III transcription.

overall_design: Cuscuta campestris were cultivated on beets. Shoot tips 4-6cm in length were cut and stimulated to form haustoria by treatment with far-red light and physical pressure (stack of microscope slides pressing the tips against 3% agar) for 4 days. The regions with visible haustoria were dissected with a scalpel and collected for RNA extractions. 3 RNA samples were collected (numbered 1, 2, and 3).


title: RNA splicing analysis using heterogeneous and large RNA-seq datasets

city: Philadelphia

first: Yoseph

last: Barash

state: Pennsylvania

address: 3700 Hamilton Walk

department: Genetics

institute: University of Pennsylvania

summary: The ubiquity of RNA-seq has led to many methods that use RNA-seq data to analyze variations in RNA splicing. However, available methods are not well suited for handling heterogeneous and large datasets. Such datasets scale to thousands of samples across dozens of experimental conditions, exhibit increased variability compared to biological replicates, and involve thousands of unannotated splice variants resulting in increased transcriptome complexity. We describe here a suite of algorithms and tools implemented in the MAJIQ v2 package to address challenges in detection, quantification, and visualization of splicing variations from such datasets. Here we created a large, realistic synthetic RNA-seq dataset of 150 simulated cerebellum samples and 150 skeletal muscle samples using BEERS. We use this as a benchmark dataset to assess the advantages of MAJIQ v2 compared to existing methods.

overall_design: RNA-seq samples were simulated based on transcript expression profiles derived from publically available GTEx v8 transcript expression levels from 150 cerebellum and 150 skeletal muscle samples in order to perform differential splicing benchmarking analysis.

first: Jorge

last: Vaquero-Garcia

first: Joseph

last: Aicher

first: San

last: Jewell

first: Matthew

last: Gazzara

first: Caleb

last: Radens

first: Anupama

last: Jha

first: Scott

last: Norton

first: Nicholas

middle: F

last: Lahens

first: Gregory

last: Grant


title: Construction ceRNA network of regulated ferroptosis in doxorubicin induced myocardial injury

city: Bengbu City

first: hongwei

last: ye

state: Anhui Province

address: 2600 Donghai Avenue

country: China

institute: Bengbu Medical College

type:
Non-coding RNA profiling by array
Expression profiling by array

summary: Based on the microarray detection results and bioinformatics analysis, we screened ferroptosis related gene Becn1 and constructed the lncRNA/miRNA/mRNA ceRNA network of regulated ferroptosis in DIMI.

overall_design: 7 samples divided into two groups,n=3 in control group and n=4 in DOX group

first: Ye

last: Hongwei

first: Li

last: Yuping

last: Lu

first: Huang

last: Yuhui

first: Wang

last: Jiahui

first: Gao

last: Qin


title: Open chromatin regions at the single cell level of CD8+ T cells from the spleens of LCMV-Armstrong or LCMV-Clone 13 infected mice

first: Gene

last: Yeo

summary: The profiles of open chromatin regions of CD8+ T cells from LCMV-Armstrong and LCMV-Clone 13 infected mice are known to be distinct from one another. We used Assay for Transposase-Accessible Chromatin using sequencing at the single cell level (scATAC-seq) to analyze these differences in splenic CD8+ T cells of these two infection conditions at Division 1 post-infection.

overall_design: Adoptively transferred CD8+ P14 T cells from the spleen of C57BL/6 mice were isolated by Fluorescence-activated cell sorting (FACS) at day 2 post-infection according to the presence of CD45.1 congenic marker and CFSE signal to isolate cells undergoing their 1st division and analyzed using scATAC-seq.

first: Lauren

last: Quezada

first: Wenhao

last: Jin


title: A knockout-first model of H3f3a gene targeting leads to developmental lethality

city: Sacramento

first: Paul

last: Knoepfler

address: 2425 Stockton Blvd, Room 633

department: Cell Biology and Human Anatomy

institute: University of California Davis

summary: Histone variant H3.3 is encoded by two genes, H3f3a and H3f3b, which can be expressed differentially depending on tissue type. Previous work in our lab has shown that knockout of H3f3b causes some neonatal lethality and infertility in mice, and chromosomal defects in mouse embryonic fibroblasts (MEFs). Studies of H3f3a and H3f3b null mice by others have produced generally similar phenotypes to what we found in our H3f3b nulls, but the relative impacts of loss of either H3f3a or H3f3b have varied depending on the approach and genetic background. Here we used a knockout-first approach to target the H3f3a gene for inactivation in C57BL6/J mice. Homozygous H3f3a targeting produced a lethal phenotype at or before birth. E13.5 embryos had some potential morphological differences from WT littermates including smaller size and reduced head size, while an E18.5 null embryo was smaller than its control littermates with several potential defects including small head and brain size as well as small lungs,  which would be consistent with a late gestation lethal phenotype. Despite a reduction in H3.3 and total H3 protein levels, the only histone H3 post-translational modification in the small panel assessed that was significantly altered was the unique H3.3 mark phospho-Serine31, which was consistently increased in null neurospheres. H3f3a null neurospheres also exhibited consistent gene expression changes including in protocadherins. Overall, our findings are consistent with the model that there are differential, cell-type-specific contributions of H3f3a and H3f3b to H3.3 functions in epigenetic and developmental processes.

overall_design: RNA was isolated from 8 independent MEF lines (3 WT and 5 KO lines) and 6 indepdendent neurosphere lines (3 WT and 3 KO  lines) for RNA-seq

first: Kelly

last: Bush

first: Vanessa

last: Cervantes


title: Triple negative breast tumors contain heterogeneous cancer cells expressing distinct KRAS-dependent collective and disseminative invasion programs

first: Joel

last: Bader

address: 3400 North Charles Street

department: Biomedical Engineering

institute: Johns Hopkins University

summary: Inter-patient and intra-tumoral heterogeneity complicate the identification of predictive biomarkers and effective treatments for basal triple negative breast cancer (b-TNBC). Invasion is the initiating event in metastasis and can occur by both collective and single-cell mechanisms. We cultured primary organoids from a b-TNBC genetically engineered mouse model in 3D collagen gels to characterize their invasive behavior. We observed that organoids from the same tumor presented different phenotypes that we classified as non-invasive, collective and disseminative. To identify molecular regulators driving these invasive phenotypes, we developed a workflow to isolate individual organoids from the collagen gels based on invasive morphology and perform RNA sequencing. We next tested the requirement of differentially regulated genes for invasion using shRNA knock-down. Strikingly, KRAS was required for both collective and disseminative phenotypes. We then performed a drug screen targeting signaling nodes upstream and downstream of KRAS. We found that inhibition of EGFR, MAPK/ERK, or PI3K/AKT signaling reduced invasion. Of these, ERK inhibition was striking for its ability to potently inhibit collective invasion and dissemination. We conclude that different cancer cells in the same b-TNBC tumor can express different metastatic molecular programs and identified KRAS and ERK as essential regulators of collective and single cell dissemination.

overall_design: C3(1)-TAg organoids were plated in 3D fibrillar collagen I for 3 days. We classified organoids based on their invasive phenotype (non-invasive, invasive (collective) and disseminative. Organoids were then removed from the gels with sharp forceps and were kept on dry ice. Trizol (Invitrogen) was then added and RNA extracted. Three independent experiments were conducted.

first: Elodie

last: Henriet

first: Hildur

last: Knutsdottir

first: Eloise

last: Grasset

first: Matthew

last: Dunworth

first: Meagan

last: Haynes

first: Andrew

last: Ewald


title: IL-13/IL-4 signaling contributes to fibrotic progression of the myeloproliferative neoplasms

city: Memphis

first: Jeremy

middle: Chase

last: Crawford

state: TN

address: MS 221, 262 Danny Thomas Place

department: Department of Host-Microbe Interactions

institute: St. Jude Children's Research Hospital

summary: Myelofibrosis (MF) is a disease with high unmet medical need since bone marrow transplant is not an option for many patients and JAK inhibitors are generally effective for only 2-3 years. MF is characterized by the presence of dysplastic megakaryocytes and progression to fulminant disease is associated with significantly increased fibrosis in the bone marrow. Despite evidence of an inflammatory state in MF that drives disease progression, the specific factors that promote megakaryocyte growth are poorly understood. Here, we analyzed changes in the cytokine profiles of MF mouse models before and after development of fibrosis coupled with analysis of bone marrow populations by scRNA-seq. We found high IL-13 levels in the bone marrow of MF mice. IL-13 promoted the growth of mutant megakaryocytes and induced the surface expression of TGF-B and collagen biosynthesis. Analysis of samples from patients with myelofibrosis revealed elevated levels of IL-13 in plasma and increased IL-13 receptor expression in megakaryocytes in the bone marrow, indicating that our observations in mice are relevant in human disease. In vivo, IL-13 overexpression promoted disease progression while reducing IL-13/IL-4 signaling reduced several features of the disease including fibrosis. Lastly, we found an increase in T cells and mast cells, which are known sources of IL-13 in MF bone marrow. Together, our data demonstrate that IL-13 is involved in the progression of MF and that inhibition of the IL-13/IL-4 signaling pathway should be investigated as a therapeutic option in MF.

overall_design: Bone marrow cells from mice transplanted with Jak2V167F Vav-Cre+ cells at 2 and 6 months after transplant (n = 2 per timepoint)

first: Johanna

last: Melo-Cardenas

first: John

last: Crispino


title: Adult T-cells impair neonatal cardiac regeneration

first: Tom

last: Gräfenhan

address: Josef-Schneider-Str. 2, Bau D15

department: Core Unit SysMed

institute: University of Wuerzburg

summary: Neonate mice and men display a transient cardiac regenerative potential that rapidly declines postnatally. Thus, patients that survive a myocardial infarction (MI) often develop chronic heart failure (HF) as a consequence of this poor myocardial healing capacity. Herein, we hypothesized that the cardiac “regenerative-to-scarring” transition is driven by the developing adaptive immune system.

overall_design: mRNA profile of neonatal hearts from control and adult T-transferred mice

first: Theresa

last: Dolejsi

first: Murilo

last: Delgobo

first: Thomas

last: Schuetz

first: Luigi

last: Tortola

first: Katrin

last: Heinze

first: Ulrich

last: Hofmann

first: Stefan

last: Frantz

first: Axel

last: Bauer

first: Frank

last: Ruschitzka

first: Josef

last: Penninger

first: Gustavo

last: Ramos

first: Bernhard

last: Haubner


title: Transcriptomic changes induced by IL33 transduction, ST2 gene knockout, and their combination in primary mouse lung fibroblasts and unseparated mouse lung cells

first: JP

last: Courneya

state: United States of America

address: 685 W Baltimore St., HSF2 Room 480

institute: University of Maryland, Baltimore

summary: RNA-Seq of unseparated lung cells and primary mouse lung fibroblasts from wild-type and ST2 gene knockout mice

overall_design: Primary mouse lung fibroblasts and unseparated lung cells from wild-type and ST2 gene knockout mice were infected with either mouse IL-33 isoform-encoding or control non-coding (NULL) adenoviruses (AdV) , and cultured. RNA-Seq analyses were performed 72 h later.

first: Sergei

middle: P

last: Atamas

first: Irina

middle: G

last: Luzina


title: The conserved histone chaperone Spt6 facilitates DNA replication and mediates genome stability [MCM ChIPseq]

city: Boston

first: Catherine

middle: LW

last: Miller

state: MA

address: 77 Ave Louis Pasteur

institute: Harvard Medical School

summary: Histone chaperones are an important class of factors that regulate chromatin accessibility for DNA-templated processes. Spt6 is a conserved histone chaperone and key regulator of transcription and chromatin structure. However, its functions outside of these roles have been little explored. In this work, we demonstrate a role for Spt6 in DNA replication and more broadly as a regulator of genome stability. Spt6 binds the replication machinery and mutations or loss of Spt6 impair DNA replication in vivo. Additionally, spt6 mutants are sensitive to DNA replication stress inducing agents, with increased sensitivity when combined with loss of DNA replication associated factors. Furthermore, spt6 mutants have elevated levels of DNA double strand breaks and are hyper-recombinogenic. R-loops do not appear to be elevated in spt6 mutants. In total, we identify Spt6 as a regulator of genome stability, at least in part through a role for Spt6 in DNA replication.

overall_design: MCM ChIPseq in WT, spt6-YW, and spt6-50 strains in biological replicates. G1-phase samples were harvested with alpha-factor treatment

middle: L

first: Fred

last: Winston


title: Dynamic antagonism between key repressive pathways maintains the placental epigenome (ChIP-BS-seq)

status: Public on Jan 02 2023

city: Berlin

first: Sara

last: Hetzel

address: Ihnestraße 63

department: Genome Regulation

institute: Max Planck Institute for Molecular Genetics

type: Methylation profiling by high throughput sequencing

summary: DNA and Histone-3 Lysine 27 methylation typically function as repressive modifications and operate within distinct genomic compartments. In mammals, the majority of the genome is kept in a DNA methylated state, whereas the Polycomb Repressive Complexes regulate the CpG-rich promoters of developmental genes. In contrast to this general framework, the extraembryonic lineages display noncanonical, globally intermediate DNA methylation levels that includes disruption of local Polycomb domains. To better understand this unusual landscape’s molecular properties, we genetically and chemically perturbed major epigenetic pathways in mouse Trophoblast Stem Cells (TSCs). We find that the extraembryonic epigenome reflects ongoing and dynamic de novo methyltransferase recruitment, which is continuously antagonized by Polycomb to maintain intermediate, locally disordered methylation. Despite its disorganized appearance, our data point to a highly controlled equilibrium between counteracting repressors within extraembryonic cells, one that can seemingly persist indefinitely without bistable features typically seen for embryonic forms of epigenetic regulation.

overall_design: H3K27me3 ChIP followed by bisulfite sequencing in wild type mouse embryonic and trophoblast stem cells.

first: Raha

last: Weigert

first: Nina

last: Bailly

first: Chuck

last: Haggerty

first: Ibrahim

last: Ilik

first: Philip

middle: Y

last: Kwong Yung

first: Carmen

last: Navarro

first: Adriano

last: Bolondi

first: Abhishek

last: Sampath Kumar

first: Chiara

last: Anania

first: Björn

last: Brändl

first: David

last: Meierhofer

first: Darío

last: Lupiáñez

first: Franz-Josef

last: Müller

first: Tugce

last: Aktas

first: Simon

last: Elsässer

first: Helene

last: Kretzmer

first: Zachary

middle: D

last: Smith

first: Alexander

last: Meissner


title: Dynamic antagonism between key repressive pathways maintains the placental epigenome (WGBS)

first: Sara

last: Hetzel

overall_design: Whole-genome profiling of DNA methylation in trophoblast stem cells with and without knockout of epigenetic regulators.

first: Raha

last: Weigert

first: Nina

last: Bailly


title: Dynamic antagonism between key repressive pathways maintains the placental epigenome (Nanopore)

overall_design: Profiling of wild type mouse trophoblast stem cells with nanopore sequencing.

first: Chuck

last: Haggerty

first: Ibrahim

last: Ilik

first: Philip

last: Kwong Yung

first: Adirano


title: Dynamic antagonism between key repressive pathways maintains the placental epigenome (RNA-seq)

overall_design: Gene expression profiling of trophoblast stem cells in normal conditions and treated with DNMT1 or EZH2 inhibitor.

first: Carmen

last: Navarro

first: Adriano

last: Bolondi

first: Abhishek

last: Sampath Kumar

first: Chiara


title: The microbiome buffers tadpole hosts from heat stress: a hologenomic approach to understand host-microbe interactions under warming

city: Pittsburgh

first: Samantha

last: Fontaine

address: 4249 Fifth Ave

department: Biological Sciences

institute: University of Pittsburgh

summary: We exposed two groups of green frog tadpoles that differed in their microbiome composition to heat stress or control conditions. We subsequently used RNAseq to profile gene expression in their gut to understand how the microbiome impacts host responses to heat.

overall_design: We had two microbiome treatments: colonized which were tadpoles with natural microbiomes and depleted which were tadpoles with artificially disrupted microbiomes (less diversity, changed composition). We had two temperature treatments: heat stress (32 Celsius, warm) or control (22 Celsius, cool). There were 5 tadpoles in each group (20 total animals). The heat stress treatment lasted 24 hours after which all tadpoles were dissected to remove the gut for RNAseq.

middle: S

first: Kevin

last: Kohl


title: Dynamic antagonism between key repressive pathways maintains the placental epigenome (ChIP-seq)

overall_design: EED ChIP-seq in wild type mouse trophoblast stem cells and TSCs treated with EZH2i.

last: Anania

first: Björn

last: Brändl

first: David

last: Meierhofer

first: Darío

last: Lupiáñez

first: Franz-Josef

last: Müller

first: Tugce

first: Simon


title: Dynamic antagonism between key repressive pathways maintains the placental epigenome (Hi-C)

first: Sara

overall_design: Hi-C data sets generated for mouse embryonic and trophoblast stem cells.

first: Raha

first: Nina

last: Aktas

last: Elsässer

first: Helene

last: Kretzmer

first: Zachary

last: Smith

first: Alexander

last: Meissner


title: Dynamic antagonism between key repressive pathways maintains the placental epigenome (RRBS)

last: Hetzel

overall_design: Profiling of DNA methylation in trophoblast stem cells in normal conditions and treated with DNMT1 or EZH2 inhibitor.

last: Weigert

last: Bailly

first: Chuck

last: Haggerty

first: Ibrahim

last: Ilik

first: Philip

first: Carmen

first: Adriano


title: A non-hemispheric transtentorial ZFTA-fusion-positive ependymoma in a 6-month-old boy

status: Public on Jan 01 2023

city: Rome

first: Evelina

last: Miele

address: Piazza di Sant'Onofrio 4

department: Oncoematologia

country: Italy

institute: Ospedale Pediatrico Bambino Gesù

type: Methylation profiling by genome tiling array

summary:
We report an infantile case of midline ‘supratentorial ependymoma ZFTA fusion-positive’ involving both the infra- and the supra-tentorial compartments (i.e., cerebellar vermis, pons and midbrain).
The radiological features, i.e., the upward displacement and compression of the supratentorial anatomical structures and the infiltrative margins of the tumour towards the cerebellar vermis, suggested an infratentorial origin of the lesion with secondary involvement of the supratentorial compartment. This case adds to the limited literature of non-hemispheric ‘supratentorial ependymoma ZFTA fusion-positive’ occurring in midline structures, both supratentorial (pineal region, thalamus) and infratentorial (cervico-medullary region and spinal cord), and in the cerebellum.  These cases highlight the need to consider this entity in the differential diagnosis of paediatric non-hemispheric lesions and raise the question of whether the inclusion of the location within the WHO definition of this tumour type may be too restrictive

overall_design: Tumor case report


title: Regulation of Macrophage IFNγ-Stimulated Gene Expression by the 1 Transcriptional Coregulator CITED1 - KnockOut

city: Murfreesboro

first: Rebecca

last: Seipelt-Thiemann

address: 1301 E Main St Box 60

institute: Middle Tennessee State University

summary: Macrophages are highly dynamic innate immune cells that adopt temporary phenotypes, known as polarization states, in response to changing environmental signals to combat microbial infection and contribute to the maintenance of tissue homeostasis. During the early stages of an infection, exposure to interferon-gamma (IFNγ) and microbial ligands induce M1 polarization, a heightened proinflammatory anti-microbial state, by regulating the expression of interferon stimulated genes (ISGs). While this response must be sufficiently vigorous to ensure the successful clearance of invading pathogens, it must also be spatially and temporally restricted to prevent uncontrolled inflammation that could result in tissue damage and disease. This is controlled by a variety of cell-extrinsic and -intrinsic mechanisms, including the expression of CBP/p300-interacting transactivator with glutamic acid/aspartic acid-rich carboxyl-terminal domain 2 (CITED2), a transcriptional coregulator that limits IFNγ-stimulated proinflammatory gene expression by inhibiting STAT1- and IRF1-regulated ISGs. In this study, we show that CITED1, another member of the CITED family of proteins, is itself an ISG and is expressed in a STAT1-dependent manner, and that IFNγ stimulates the nuclear accumulation of fluorescently tagged CITED1 proteins. In contrast to CITED2, ectopic expression of CITED1 enhanced the expression of a subset of ISGs, including Ccl2, Ifit3b, Isg15, and Oas2. This effect was reversed in a Cited1 null cell line produced by CRISPR-based genomic editing. Collectively, these data show that CITED1 helps to maintain proinflammatory gene expression during periods of prolonged IFNγ exposure and suggests a distinct and antagonistic relationship between CITED proteins in the regulation of macrophage inflammatory function.

overall_design: Macrophages were transduced with a CAS and either scramble guide RNA or guide RNA targeted to CITED1. Following selection, RNA was extracted.

first: David

last: Nelson

first: Erin

last: McClelland


title: Dynamic antagonism between key repressive pathways maintains the placental epigenome

overall_design: Profiling of DNA methylation, gene expression, genome-wide chromatin interactions and histone binding in trophoblast stem cells.

last: Kwong Yung

last: Navarro

last: Bolondi

first: Abhishek

last: Sampath Kumar

first: Chiara

last: Anania

first: Björn

last: Brändl

last: Meierhofer

first: Darío

first: Franz-Josef

first: Tugce


title: Regulation of Macrophage IFNγ-Stimulated Gene Expression by the 1 Transcriptional Coregulator CITED1

overall_design: Macrophages were transduced with an inducible CITED1 (p2080) or not (RAW). Cells of both types were treated with the inducer (doxycyline), gamma interferon, or both.  RNA was extracted following treatments.


title: In individuals with Williams syndrome, dysregulation of methylation in non-coding regions of neuronal and oligodendrocyte DNA is associated with pathology and cortical development

first: Asaf

last: Marco

address: Herzl 229

institute: The Hebrew University of Jerusalem

summary: Williams syndrome (WS) is a neurodevelopmental disorder caused by a heterozygous micro-deletion in the WS critical region (WSCR) and is characterized by hyper-sociability and neurocognitive abnormalities. Nonetheless, whether and to what extent WSCR deletion leads to epigenetic modifications in the brain and induces pathological outcomes remains largely unknown. By examining DNA methylation in frontal cortex, we revealed genome-wide disruption in the methylome of individuals with WS, as compared to typically developed (TD) controls. Surprisingly, differentially methylated sites were predominantly annotated as introns and intergenic loci and were found to be highly enriched around binding sites for transcription factors that regulate neuronal development, plasticity and cognition. Moreover, by utilizing enhancer–promoter interactome data, we confirmed that most of these loci function as active enhancers in the human brain or as target genes of transcriptional networks associated with myelination, oligodendrocyte (OL) differentiation, cognition and social behavior. Cell type–specific methylation analysis revealed aberrant patterns in the methylation of active enhancers in neurons and OLs, and important neuron-glia interactions that might be impaired in individuals with WS. Finally, comparison of methylation profiles from blood samples of individuals with WS and healthy controls, along with other data collected in this study, identified putative targets of endophenotypes associated with WS, which can be used to define brain-risk loci for WS outside the WSCR locus, as well as for other associated pathologies. In conclusion, our study illuminates the brain methylome landscape of individuals with WS and sheds light on how these aberrations might be involved in social behavior and physiological abnormalities. By extension, these results may lead to better diagnostics and more refined therapeutic targets for WS.

overall_design: To better understand the potential impact of the 7q11.23 deletion on the brain epigenome, we addressed DNA methylation in BA9 in frontal cortex sections taken from three individuals with WS and five matching controls. Reduced representation bisulfite followed by sequencing (RRBS) libraries were prepared from these brain tissues.

first: Sari

last: Trangle

first: Tali

last: Rosenberg

first: Hadar

last: Parnas

first: Gilad

last: Levy

first: Ela

last: Bar

first: Boaz

last: Barak


title: mt-Ty 5´tiRNA regulates skeletal muscle cell proliferation and differentiation

first: Xin

last: Wang

address: 300 Longwood Ave,

institute: Boston children's hospital

summary: Understanding the molecular mechanisms underlying the regulation of gene expression during skeletal muscle cell proliferation and differentiation is critical to the development of novel therapeutic targets to treat muscle disorders. In this study, we report the identification and characterization of mt-Ty 5’tiRNAs, tRNA-derived fragments (tiRNAs) we identified in skeletal muscle. mt-Ty 5’tiRNAs are found in the nucleus and mitochondria of skeletal muscle cells and its expression is induced by angiogenin. Gapmer-mediated inhibition of tiRNAs in skeletal muscle C2C12 myoblasts resulted in mitochondrial fragmentation. We also observed a decrease in cell proliferation and myogenic differentiation upon mt-Ty 5’tiRNA inhibition; consistent with this observation, RNA-seq and transcriptome analyses revealed that skeletal muscle cell differentiation and cell proliferation pathways were also downregulated. Conversely, overexpression of mt-Ty 5’tiRNAs in C2C12 cells led to a reversal of the transcriptional trends noted for these pathway genes. These data reveal mt-Ty 5’tiRNAs as a skeletal muscle enriched tiRNA that plays important roles in myoblast proliferation and differentiation. Our study also highlights the potential for the development of tiRNAs as novel therapeutic targets for muscle related diseases

overall_design: Comparative transcripts profiling analysis of RNA-seq data for mt-Ty 5’tiRNAs gapmer, and control with two biological replicates ;  the mt-Ty 5’tiRNAs overexpression and control with four biological replicates.

first: Jun

last: Cao

first: Vivek

last: Advani

first: Yao

middle: W

last: Lu

first: Andrea

last: Malizia

first: Gurinder

last: Bir Singh

first: Zhan-Peng

last: Huang

first: Jianming

last: Liu

first: Chunbo

first: John

last: Mably

first: Kaifu

last: Chen

first: Da-Zhi


title: RNA-sequencing analysis of hand cartilage tissue from osteoarthritis patients with/without common polymorphic variants in ALDH1A2 gene

city: Oxford

first: Linyi

last: Zhu

address: Roosevelt drive

country: United Kingdom

institute: Kennedy institute of Rheumatology, University of Oxford

summary:
Polymorphic variants (rs3204689 and rs4238326) in aldehyde dehydrogenase 1 family member A2 (ALDH1A2) are associated with severe hand OA.
Trapezium tissue from patients with severe base of thumb OA was collected at the time of surgery. To investigate the transcriptional effects of the ALDH1A2 variants, we compared articular cartilage samples from patients homozygous for low-risk and high-risk ALDH1A2 alleles.

overall_design: We collected trapezium samples from 19  hand osteoarthritis patients for bulk RNA-sequencing analysis. 12 patients were homozygous for the high risk variants (rs3204689 and rs4238326; genotype recorded as “HO”) and 7 were homozygous for low-risk variants (genotype recorded as “WT”).

first: Tonia

last: Vincent

first: Lada

last: Koneva

first: Stephen

middle: N

last: Sansom


title: Increased length-dependent activation of human engineered heart tissue after chronic α1A-adrenergic agonist treatment: testing a novel heart failure therapy

city: San Francisco

first: Anthony

middle: J.

last: Baker

address: 4150 Clement St

department: VA Medical Center and Department of Medicine

institute: University of California San Francisco

summary: Chronic stimulation of cardiac a1A-adrenergic receptors (a1A-ARs) improves symptoms in multiple preclinical models of heart failure. However, the translational significance remains unclear. This study tested the effects of chronic a1A-AR stimulation on human engineered heart tissue (EHT). EHTs were created from thin slices of decellularized pig myocardium seeded with human induced pluripotent stem cell (iPSC)-derived cardiomyocytes and fibroblasts. Using a paired experimental design, EHTs were cultured for 3 wk., mechanically tested, cultured again for 2 wk. with a1A-AR agonist A61603 (10 nM) or vehicle control, and retested after drug washout for 24-hr. Separate control experiments determined the effects of EHT age (3-5 wk.) or repeat mechanical testing. We found that chronic A61603 treatment caused a 25% increase of length-dependent activation (LDA) of contraction compared to vehicle treatment (n=7/group, P=0.035). EHT force was not increased after chronic A61603 treatment. However, after vehicle treatment, EHT force was increased 35% relative to baseline testing (n=7/group, P=0.022), suggesting EHT maturation. Control experiments showed increased EHT force resulted from repeat mechanical testing, not from EHT aging. RNA-seq analysis confirmed the a1A-AR is expressed in human EHTs and found chronic A61603 treatment affected gene expression in biological pathways which are known to be activated by a1A-ARs, including the MAP kinase signaling pathway. In conclusion, chronic stimulation of the a1A-AR, a promising preclinical target for treating heart failure, gave mixed results in human EHTs. Chronic a1A-AR stimulation had a positive effect (increased LDA) but potentially, a negative effect (lower EHT force). The translational significance of these findings requires further study.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for EHT cells under either chronic A61603 treatment or vehicle treatment (control).

first: Rupert

last: Cassady

first: Javier

middle: E

last: Lopez

first: Elizabeth

middle: J

last: Cortez-Toledo

first: Omar

last: De la Cruz Cabrera

first: Naomi

middle: C

last: Chesler

first: Paul

last: Simpson

first: Stuart

last: Campbell


title: Staphylococcus aureus stimulates neutrophil itaconate production that suppresses the oxidative burst

city: New York

first: Kira

middle: Leigh

last: Tomlinson

state: New York

address: 650 W 168th Street, Room 4416

department: Pediatrics

institute: Columbia University

summary: Neutrophils are critical in the host defense against Staphylococcus aureus, a major human pathogen. However, even in the setting of a robust neutrophil response, S. aureus can cause persistent infection. Here we demonstrate that S. aureus impairs neutrophil function by triggering the production of the anti-inflammatory metabolite, itaconate. The enzyme that synthesizes itaconate, Irg1, is selectively expressed in neutrophils during S. aureus pneumonia. Itaconate inhibits neutrophil glycolysis and oxidative burst, which impairs survival and bacterial killing. In a murine pneumonia model, neutrophil Irg1 expression protects critical lung cell populations from oxidative stress but compromises bacterial clearance. S. aureus is thus able to evade innate immune clearance by targeting neutrophil metabolism and inducing the production of the antiinflammatory metabolite itaconate.

overall_design: 8-week old WT and Irg1-/- C57BL/6NJ mice were intranasally infected with 2 x 10^7 USA300 LAC in 50 mL PBS or treated with PBS alone; 24 hours after infection, the lungs were harvested and subjected to scRNA-seq on the 10X Genomics platform.

first: Alice

last: Prince


title: Single-copy locus proteomics of early- and late-firing DNA replication origins identifies a role of Ask1/DASH complex in replication timing control

city: Munich

first: Tamas

last: Schauer

address: Feodor-Lynen-Straße 21

department: Institute of Epigenetics and Stem Cells

institute: Helmholtz Zentrum München

summary: The chromatin at origins of replication is thought to influence DNA replication initiation in eukaryotic genomes. However, it remains unclear how the chromatin composition controls the firing of early-efficient (EE) or late-inefficient (LI) origins. Here, we used site-specific recombination and single-locus chromatin isolation to purify EE and LI replication origins in Saccharomyces cerevisiae. Using mass spectrometry, we define the histone modification landscape and identify the protein composition of native chromatin regions surrounding the EE and LI replication start sites. In addition to the known origin interactors, we find novel origin-associated factors, such as the kinetochore-associated Ask1/DASH complex. Strikingly, we show that Ask1 regulates the replication timing control of specific origins in yeast. Thus, our unbiased approach identifies functionally-relevant proteomes at single-copy loci and would be widely applicable to provide an in-depth quantitative characterization of histone modification and protein interaction networks of chromatin at any genomic locus of interest.

overall_design: DNA sequencing based copy number analysis of origin activity in HU-arrested and synchronously released S. cerevisiae cells after targeted degradation of Ask1 or targeted recruitment of Ask1 at specific replication origins ARS305, ARS313 or ARS316.

first: Matthias

last: Weiß

first: Anna

last: Chanou

first: Stephan

last: Hamperl


title: Methylation profiling of dedifferentiated chondrosarcoma (DDCS) samples

first: Cristina

last: Antonescu

state: NY

address: 1275 York Avenue

institute: Memorial Sloan Kettering Cancer Center

summary:
Genome wide DNA methylation profiling of dedifferentiated chondrosarcoma samples. Genomic DNA was extracted from formalin-fixed paraffin-embedded (FFPE) tissue after manual macrodissection to ensure at least 10% tumor content, followed by bisulfite converstion.
All samples were processed on the Infinium 850k array and scanned using the Illumina iScan, according to the manufacturer's recommended protocol.

overall_design: For methylation analysis, we included a total of 18 in-house DDCS cases, of which 7 samples were macrodissected well-differentiated chondrosarcoma components and 11 were macrodissected dedifferentiated components

first: Josephine

middle: K

last: Dermawan

first: Meera

last: Hameed


title: TCR sequencing analysis of islets from NOD mice on hydrolyzed casein and hydrolyzed casein + 4% gluten diets

city: Chicago

first: Alexander

last: Chervonsky

institute: University of Chicago

summary: Protein source in diet greatly influences the incidence of type-1 diabetes (T1D) in non-obese diabetic (NOD) mouse colonies.  NOD mice fed a diet containing hydrolyzed casein (HC) as the sole protein source are protected from T1D. Replacing 1/5th of the HC with gluten restores high T1D to NOD mice. We hypothesized that gluten might promote inflammation in the islets. We used TCR sequencing  to characterize the clonal frequency of infiltrating T cells from the pancreatic islets of Langerhans of NOD mice fed a 20% HC diet or a 16% HC + 4% gluten diet.

overall_design: TCR-seq was performed on single cell suspensions from isolated pancreatic islets of Langerhans from 16 week old NOD/ShiLtJ mice fed either 20% HC or 16% HC + 4% gluten diets. Islets from 3 HC fed and 3 HC+4%gluten fed mice were labeled with unique Totalseq-C antibodies (Biolegend), pooled, and sequenced in two seperate runs (Run 1 and Run 2).

first: Matthew

last: Funsten

middle: V

first: Aly

middle: A

last: Khan

first: Derek

last: Reiman


title: Single cell analysis on the HIV-infected kidney cells

first: Zeguo

last: Sun

address: 1468 Madison Avenue

institute: Ichan School of Medicine at Mount Sinai

summary: HIV-vpr expression was mainly detected in tubule cells after infection. By comparing the HIV-infected and wild type kidney cells on single cell level, a strong shift was observed in the PT S3 segment cell population. Transcription factor analysis revealed Trp53 playing important regulation role in HIV infected cells.

overall_design: VPR;Pax8-rtTa mice were fed DOX grain-based rodent diet at 7 week of age and continued for 11 weeks induction until mice were euthanized. 3 VPR;Pax8-rtTa mice and 2 Wild type mice were sequenced.

first: Jia

last: Fu

first: Ya

last: Chen


title: H2AK119ub1 and H3K27me3 ChIP-seq in conditional knockout of Bap1

city: Madison

first: John

last: Svaren

state: WI

address: 1500 Highland Ave.

department: Comparative Biosciences

institute: University of Wisconsin

summary: ChIP-seq of H2AK119ub1  and H3K27me3 in a conditional knockout of Bap1

overall_design: H2AK119ub1 and H3K27me3 ChIP samples were prepared from mouse sciatic nerve, prepared from control (Bap1 fl/fl) or a Schwann cell-specific knockout of Bap1 (Bap1 fl/fl/Mpz-cre).

first: Raghu

last: Ramesh


title: Expression data of JMJD6-KO, scramble and wild-type MCF7 cells

city: MILAN

first: BIANCA

last: CIONI

state: MI

address: Via Amadeo 41

department: Molecular Oncology

institute: Istituto Nazionale Tumori

summary: JMJD6 expression was shown to have different roles in estrogen receptor (ER)-positive and -negative breast cancer cells

overall_design: In this dataset, JMJD6 gene was genetically deleted in MCF7 cells via CRISPR-Cas9 technology and submitted with scramble and wild-type controls for ClariomS assay

first: Bianca

last: Cioni

first: Silvia

last: Ratti

first: Annamaria

last: Piva

first: Irene

last: Tripodi

first: Matteo

last: Milani

first: Francesca

last: Menichetti

first: Tiziana

last: Langella

first: Laura

last: Botti

first: Loris

last: De Cecco

first: Claudia

last: Chiodoni

first: Daniele

middle: P

last: Lecis

first: Mario

last: Colombo


title: Expression data of JMJD6-KO, scramble and wild-type MDA-MB231 cells

overall_design: In this dataset, JMJD6 gene was genetically deleted in MDA-MB231 cells via CRISPR-Cas9 technology and submitted with scramble and wild-type controls for ClariomS assay


title: Gene expression analyses of HUVECs treated with conditioned medium derived from bone marrow mesenchymal stromal cells (BMSC-CM)

city: Rotterdam

first: Jeroen

last: van de Peppel

address: 's Gravendijkwal 230

country: Netherlands

institute: ErasmusMC

summary: A functional vascular system is a prerequisite for bone repair as disturbed angiogenesis often causes non-union. Paracrine factors released from human bone marrow mesenchymal stromal cells (BMSCs) have angiogenic effects on endothelial cells. However, whether these paracrine factors participate in blood flow dynamics within bone capillaries remains poorly understood. Here, we used two different microfluidic designs to investigate critical steps during angiogenesis and found pronounced effects of endothelial cell proliferation as well as chemotactic and mechanotactic migration induced by BMSC conditioned medium (CM). The application of BMSC-CM in dynamic cultures demonstrates that bioactive factors in combination with fluidic flow-induced biomechanical signals significantly enhanced endothelial cell migration. Transcriptional analyses of endothelial cells demonstrate the induction of a unique gene expression profile related to tricarboxylic acid cycle and energy metabolism by the combination of BMSC-CM factors and shear stress, which opens an interesting new avenue to explore during fracture healing. Our results stress the importance of in vivo - like microenvironments simultaneously including biochemical, biomechanical and oxygen levels when investigating key events during vessel repair.

overall_design: HUVECs were incubated with BMSC-CM in static culture and dynamic culture for 24 hours and gene expression profiles were generated.

first: Shuang

last: Zhang

first: Bram

last: van der Eerden


title: Regulation of endogenous retrovirus-derived regulatory elements by GATA2/3 and MSX2 in human trophoblast stem cells

city: Bethesda

first: Mingan

last: Sun

state: Maryland

address: Building 6B, 6 Center Drive

department: Eunice Kennedy Shriver National Institute of Child Health and Human Development

institute: National Institute of Health

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

overall_design: Refer to individual Series


title: Gene expression in rat brain 6 weeks after blast  exposure

city: Bronx

first: Rita

last: De Gasperi

address: 130 West Kingsbridge Road

institute: JJ Peters VA Medical Center

summary: Analysis of gene expression changes in rat brain of rats exposed to blast overexpression 6 weeks after blast exposure

overall_design: Sequence  analysis of RNA isolated  from  amygdala, anterior cortex , hippocampus and cerebellum isolated from  sham and blast exposed rats 6 weeks post-exposure

first: Gregory

last: Elder


title: Stable methylation loci are associated with systolic blood pressure in a Croatian island population

city: Cincinnati

first: Scott

middle: M

last: Langevin

state: OH

address: 160 Panzeca Way, ML0056

department: Department of Environmental Health

institute: University of Cincinnati College of Medicine

summary: Adriatic islanders have a high prevalence of metabolic syndrome that is not fully explained by previous behavioral, dietary, and genetic studies. Some DNA methylation remains stable and may be established in early life, whereas others are dynamic and may vary over time in response to different conditions. The objective of the present study was to identify stable and dynamic DNA methylation loci associated with cardiometabolic traits among the populations from the island of Hvar, the largest island in the eastern Adriatic coast. An epigenome-wide association study (EWAS) was conducted using peripheral blood that was longitudinally collected at two time points, 10 years apart. DNA methylation was analyzed via Infinium MethylationEPIC BeadArray. Stable and dynamic loci were identified using linear mixed models. A total of 43,041 loci were classified as stable, while 45,769 loci were classified as dynamic. Associations between various cardiometabolic traits and stable and dynamic methylation loci were respectively assessed using linear mixed effects models adjusted for age, sex, and smoking status. After adjustment for false-discovery rate, 24 CpG loci were significantly associated with systolic blood pressure (Q < 0.1), of which 22 were stable loci and 2 were dynamic loci; while 12/22 (55%) of the systolic blood-pressure-associated stable loci resided in a gene promoter. Additionally, there was one stable locus associated with serum calcium and one with C-reactive protein. The results suggest that multiple genes involved in the determination of systolic blood pressure may be regulated via epigenetic programming established in early life.

overall_design: Longitudinal

first: Ranjan

last: Deka

first: Liang

last: Niu

first: Alexander

last: Sprague

first: Xiang

first: Sasa

last: Missoni

first: Pavao

last: Rudan


title: RNAseq from Mfn2-R707W knock-in mice

city: Cambridge

first: David

middle: B

last: Savage

address: Addenbrooke's Hospital

department: Institute of Metabolic Science

institute: University of Cambridge

summary: Mitochondrial dysfunction has been reported in obesity and insulin resistance, but primary genetic mitochondrial dysfunction is generally not associated with these, arguing against a straightforward causal relationship. A rare exception, recently identified in humans, is a syndrome of lower body adipose loss, leptin-deficient severe upper body adipose overgrowth, and insulin resistance caused by the p.Arg707Trp mutation in MFN2, encoding mitofusin-2. How this selective perturbation of mitochondrial function leads to tissue- and adipose depot-specific growth abnormalities and systemic biochemical perturbation is unknown. To address this, Mfn2R707W/R707W knock-in mice were generated and phenotyped on chow and high fat diets. Electron microscopy revealed adipose-specific mitochondrial morphological abnormalities. Oxidative phosphorylation by isolated mitochondria was unperturbed, but the cellular integrated stress response was activated in adipose tissue. Fat mass and distribution, body weight, and systemic glucose and lipid metabolism were unchanged, however serum leptin and adiponectin concentrations, and their secretion from adipose explants were reduced. Pharmacological induction of the integrated stress response in wild-type adipocytes also reduced secretion of leptin and adiponectin, suggesting an explanation for the in vivo findings. These data suggest that the p.Arg707Trp MFN2 mutation perturbs mitochondrial morphology and activates the integrated stress response selectively in adipose tissue. In mice, this does not disrupt most adipocyte functions or systemic metabolism, whereas in humans it is associated with pathological adipose remodelling and metabolic disease. In both species, disproportionate effects on leptin secretion may relate to cell autonomous induction of the integrated stress response.

overall_design: Wild-type C57BL/6J mice and knock-ins with homozygous p.Arg707Trp mutations in Mfn2. (1) inguinal WAT from chow fed animals (n=7 WT, n=6 KI, single technical replicates); (2) inguinal WAT from HFD-fed animals (n=8 WT, n=8 KI, two technical replicates per animal); and (3) BAT from HFD-fed animals (n=6 WT, n=6 KI, single technical replicate)

first: Jake

last: Mann

first: Robert

last: Semple


title: Gene expression  in rat brain 12 months after exposure

summary: RNA seq analysis of amygdala, anterior cortex , hippocampus  and cerebellum from sham and blast exposed rats 12 months post-exposure

overall_design: RNA seq analysis of amygdala, anterior cortes and hippocampus from sham and blast exposed rats 12 months post-exposure


title: The F-box cofactor UFO redirects the LEAFY floral regulator to novel cis-elements

city: Grenoble

first: Christel

last: Carles

address: 17 avenue des Martyrs

country: France

institute: CNRS

summary: In angiosperms, flower patterning requires the localized expression of the APETALA3 (AP3) floral homeotic gene involved in petal and stamen development. AP3 is synergistically induced by the master transcription factor LEAFY (LFY) and the F-box protein UNUSUAL FLORAL ORGANS (UFO), but the molecular mechanism underlying this synergy has remained unknown. Here we show that the connection to ubiquitination pathways suggested by the F-box domain of UFO is mostly dispensable and that UFO instead acts by forming a transcriptional complex with LFY on newly discovered regulatory elements. The cryo-electron microscopy structure of the UFO-LFY-DNA complex shows that UFO-DNA contacts allows relocating LFY to novel DNA sites. Finally, we show that this complex has a deep evolutionary origin, largely predating flowering plants. This work unravels an unsuspected role for a member of one of the largest protein families in plants as a modulator of the DNA binding specificity of a master TF.

overall_design: ampDAP-seq experiments with tagged LFY, LFYK249R and UFO proteins were done on 2-weeks old Arabidopsis thaliana seedlings. LFY and UFO tagged proteins were first mixed together and then with DNA prior to affinity purification. The same strategy was applied to the mutated version of LFY (LFYK249R), with and without UFO. Three replicates were done for LFYUFO and only consensus peaks in all three were retained. Two replicates were done for LFYK249R and LFYK249RUFO and common peaks in the two were retained.

first: Philippe

last: Rieu

first: Laura

last: Turchi

first: Jérémy

last: Lucas

first: Romain

last: Blanc-Mathieu

first: François

last: Parcy

first: Chloe

last: Zubieta


title: Age-Associated B Cells are Heterogenous and Dynamic in Autoimmunity

first: Mark

last: Shlomchik

address: The Assembly, 5051 Centre Avenue

department: Department of Immunology

institute: University of Pittsburgh

summary: Age-Associated B Cells (ABCs) are a population of CD11c+ and/or CD11b+ B cells expanded in autoimmune lupus.  To understand potential heterogeneity within ABCs and the relationship of ABCs to other splenic B cell subsets, we performed scRNA-seq with 5' V(D)J and CITE-seq cell surface features using the 10x Genomics Chromium platform on total splenic CD19+ B cells sorted from lupus-prone MRL/lpr.  We found that ABC are a distinct transcriptional lineage that resolved into two related clusters.  Analysis of BCR heavy chains identified clones present in both ABC and plasmablast compartments.  Inferred lineage trees based on V gene somatic hypermutation suggested that self-renewing ABC feed into the plasmablast compartment repeatedly while continuing to themselves undergo mutation and expansion.

overall_design: scRNAseq data were generated from 3 female MRL/lpr mice at 18 weeks old

first: Kevin

last: Nickerson

first: Shuchi

last: Smita

first: Kenneth

last: Hoehn

first: Anthony

last: Marinov

first: Kayla

last: Thomas

first: Yi

last: Yang

first: Sheldon

middle: I

last: Bastacky

first: Steven

middle: H

last: Kleinstein


title: A Parathyroid Hormone/Salt Inducible Kinase Signaling Axis Controls Renal Vitamin D Activation and Organismal Calcium Homeostasis

city: Boston

first: Sung-Hee

middle: Seanna

last: Yoon

address: 50 Blossom St

institute: Massachusetts General Hospital

summary: The renal actions of parathyroid hormone (PTH) promote 1,25-vitamin D generation; however, the signaling mechanisms in renal epithelial cells downstream of the PTH receptor that control vitamin D metabolism remain unknown. Here we demonstrate that Salt Inducible Kinases (SIKs) control renal 1,25-vit D production downstream of PTH signaling via regulating Cyp27b1 expression. As PTH inhibits the cellular activity of SIKs by cAMP-dependent PKA phosphorylation in bone, we hypothesized that SIKs would also work as an essential mediator of PTH doownstream signaling in kidney, thus regulate vitamin D metabolism. Whole tissue and single cell transcriptomics in kidney demonstrates that both PTH and pharmacologic SIK inhibitors regulate a vitamin D gene module in specific proximal tubule cells. Moreover, small molecule SIK inhibitors directly increase 1,25-vit D production and renal Cyp27b1 mRNA expression in mice and in human embryonic stem cell-derived kidney organoids. Global- and kidney-specific Sik2/Sik3 mutant mice show Cyp27b1 upregulation, elevated serum levels of 1,25-vit D, and PTH-independent hypercalcemia. The SIK substrate CRTC2 shows PTH and SIK inhibitor-inducible binding to key Cyp27b1 regulatory enhancers in the kidney, which are also required for SIK inhibitors to increase Cyp27b1 in vivo. Lastly, in a podocyte injury mouse model of chronic kidney disease (CKD) characterized by low 1,25-vit D levels, SIK inhibitor treatment stimulates both renal Cyp27b1 expression and 1,25-vit D production.

overall_design: C57BL6 male mice were treated with a single IP injection of either the vehicle, hPTH(1-34) or YKL-05-099 (IP) and total kindney mRNA profiles were generated by deep sequencing (n= 6 per treatment).

first: Ruslan

last: Sadreyev

first: Marc

last: Wein


title: Single-Cell Gene Expression Atlas of Human Follicular Aspirates: Impacts of Leukocytes on the Ovulatory Process

city: LEXINGTON

first: Yohan

last: Choi

state: KY

address: 800 Rose St, UKMC MS341

institute: University of Kentucky

summary: Leukocytes are in situ regulators for ovarian functions. However, little is known about leukocyte subpopulations and functions in ovulatory follicles, especially in humans. This study performed single-cell RNA sequencing and bioinformatics analyses using follicular aspirates obtained from four IVF patients. Nineteen clusters were identified: six follicular cells, ten subsets of leukocytes, and one RBC/platelet. Using dominant follicle tissues collected throughout the periovulatory period from normally cycling women and cultured primary human granulosa/lutein cells, we determined the cell type-specific expression of PTPRC, CD68, IL1B, and IL1R1. RNA velocity analyses revealed developmental dynamics across five granulosa cell groups and two projections of granulosa cell differentiation. To understand how leukocytes promote periovulatory changes, we sought leukocyte-derived factors potentially involved in 1) granulosa cell function by activating their receptors, 2) tissue remodeling, and 3) angiogenesis in ovulatory follicles. Collectively, these findings identified diverse leukocyte-secreted factors that facilitate ovulation and luteinization in the human ovary.

overall_design: Ovarian follicular aspirates obtained from four women undergoing the standardized IVF procedure

first: Hayce

last: Jeon

first: Mats

last: Brännström

first: James

last: Akin

first: Thomas

last: Curry Jr

first: Misung

last: Jo


title: Early-life bacterial exposure affects lung surfactant gene-expression, surfactant function and respiratory rate compared to germ-free mice

city: Rockville

first: Julian

last: Lui

address: 605 WATTS BRANCH PKWY, 10 Center Drive

institute: NICHD

summary: Mouse lung tissue transcriptome at postnatal day 10. Samples include 9 germ-free, 9 monocultured Bifidobacterium innoculated, and 9 full caecium microbiota innoculated. Plus 5 adult lung tissue as reference.

overall_design: Pregnant BALB/c germ-free mice were kept in separate germ-free isolators until either (a) no exposure (GF), (b) exposure to either Bifidobacterium animalis ssp. Lactis (BI04) or to (c) full cecum content harvested from female SPF mice (Cecum). Subsequently, perinatally exposed offspring were used for the analyses. Lung tissue transcriptomics analysis was done at postnatal day 10 (PNday10) at first phase of lung alveolar development. RNA were extracted for bulk RNA-sequencing.

last: Klingenberg Barfod

last: Iben


title: Dynamic antagonism between key repressive pathways maintains the placental epigenome (MINUTE-ChIP)

city: Berlin

first: Sara

last: Hetzel

address: Ihnestraße 63

institute: Max Planck Institute for Molecular Genetics

summary: DNA and Histone-3 Lysine 27 methylation typically function as repressive modifications and operate within distinct genomic compartments. In mammals, the majority of the genome is kept in a DNA methylated state, whereas the Polycomb Repressive Complexes regulate the CpG-rich promoters of developmental genes. In contrast to this general framework, the extraembryonic lineages display noncanonical, globally intermediate DNA methylation levels that includes disruption of local Polycomb domains. To better understand this unusual landscape’s molecular properties, we genetically and chemically perturbed major epigenetic pathways in mouse Trophoblast Stem Cells (TSCs). We find that the extraembryonic epigenome reflects ongoing and dynamic de novo methyltransferase recruitment, which is continuously antagonized by Polycomb to maintain intermediate, locally disordered methylation. Despite its disorganized appearance, our data point to a highly controlled equilibrium between counteracting repressors within extraembryonic cells, one that can seemingly persist indefinitely without bistable features typically seen for embryonic forms of epigenetic regulation.

overall_design: Profiling of H3K4me3, H3K27me3 and H2AK119ub1 in embryonic and trophoblast stem cells.

first: Raha

last: Weigert

first: Nina

last: Bailly

first: Chuck

last: Haggerty

first: Ibrahim

last: Ilik

first: Philip

middle: Y

last: Kwong Yung

first: Carmen

last: Navarro

first: Adriano

last: Bolondi

first: Abhishek

last: Sampath Kumar

first: Chiara

last: Anania

first: Björn

last: Brändl

first: David

last: Meierhofer

first: Darío

middle: G

last: Lupiáñez

first: Franz-Josef

last: Müller

first: Tugce

last: Aktas

first: Simon

last: Elsässer

first: Helene

last: Kretzmer

first: Zachary

middle: D

last: Smith

first: Alexander

last: Meissner


title: Embryo Cryopreservation leads to sex-specific DNA methylation perturbations in both human and mouse placentas

city: Philadelphia

first: Yemin

last: Lan

address: 3400 Civic Center Blvd

institute: University of Pennsylvania

summary:
Transfer of frozen-thawed embryos leads to sex-specific DNA hypermethylation in both human and mouse placentas. : In humans, frozen ET led to placentas with significant alterations in DNA methylation with most genes demonstrating hypermethylation compared to placentas from fresh ET (4402 CpGs; 1600 genes; p-value < 0.05 in two–tailed unpaired t tests, and a mean methylation difference > 0.05). When compared to control samples, both frozen and fresh samples showed significant differences in methylation.(Frozen vs Control: 5600 CpGs, 2775 genes; Fresh vs Control: 4096 CpGS; 1914 genes; p-value < 0.05, mean methylation difference > 0.05). Paired analysis showed similar trends, despite controlling for maternal environment. Sex specific analysis revealed that these changes were primarily driven by male placentas, as seen by a larger number of CpGs that were differentially methylated, and a larger proportion of genes with at least 2 differentially methylated CpG sites in male placentas as compared to female placentas. (Males: 15572 CpGs, 4399 genes; Females: 7441 CpGs, 3070 genes; p-value < 0.05, mean methylation difference > 0.05). In order to isolate the effects of vitrification we utilized the mouse model, controlling for the effect of the maternal hormonal milieu. In the first genome-wide profiling of DNA methylation in a mouse IVF model, we found that trends in DNA methylation differences paralleled those seen in human placentas. Using similar significance cutoffs as for human samples, and BumpHunter analysis that identifies differentially methylated regions to compensate for a lower number of samples, placentas derived from frozen embryos were predominantly hypermethylated compared to placentas after fresh ET (589 DMRs, 445 genes). Both frozen and fresh embryo transfer samples demonstrated perturbations compared to control samples (Frozen vs Control: 1787 DMRs, 1319 genes; Fresh vs Control: 1119 DMRs, 808 genes). Consistent with the sex-specific trends we observed in human placentas, differences between samples derived from frozen compared to fresh embryo transfer were driven by changes in male placentas (Males: 1069 DMRs, 798 genes; Females:14 DMRs, 11 genes) .  When considering the effect of IVF as a whole, murine placentas were predominantly hypomethylated, replicating existing human genome-wide methylation data.  As seen previously, IVF led to changes in placental weight, placental morphology and microvessel density in mice, but no persistent changes were seen after embryo vitrification alone.
Sexually dimorphic epigenetic changes could indicate differential susceptibility of male and female embryos to IVF-associated perturbations. This observation highlights the importance of sex-specific evaluation of the incidence of adverse outcomes. Similarities between changes seen in mouse and human samples underscore the suitability of the mouse model in evaluating the effect of ART on the epigenetic landscape. This convergence is especially valuable, given limited access to human tissues and the ability to isolate specific interventions in the mouse model.

overall_design: Mouse (Mus musculus) term placentas from 8 term mouse placentas derived from blastocysts after frozen ET (4 female,4 male); 8 from fresh ET (4 female, 4 male) and 8 from unassisted conceptions were obtained for profiling of genome-wide DNA methylation changes.

first: Sneha

last: Mani

first: Jayashri

last: Ghosh

first: Eric

last: Rhon-Calderon

first: Teri

last: Ord

first: Charikleia

last: Kalliora

first: Joe

last: Chan

first: Bryant

last: Schultz

first: Elaine

last: Vaughan-Williams

first: Christos

last: Coutifaris

last: Sapienza

first: Suneeta

last: Senapati

first: Marisa

middle: S

last: Bartolomei

first: Monica

last: Mainigi