title: Single-cell analysis of CX3CR1+ cells reveal a pathogenic role for BIRC5+ myeloid proliferating cells driven by Staphylococcus aureus leukotoxins

city: New York City

first: Ze

last: Chen

address: 430 East 29th Street

institute: NYU Grossman School of Medicine

summary: Our previous studies identified a population of stem cell-like proliferating myeloid cells within inflamed tissues that could serve as a reservoir for tissue macrophages to adopt different activation states depending on the microenvironment. By lineage tracing cells derived from CX3CR1+ precursors in mice during infection and profiling by scRNA-seq, here we identify a cluster of BIRC5+ myeloid cells that expanded in the liver during either chronic infection with the parasite Schistosoma mansoni or the bacterial pathogen Staphylococcus aureus. In the absence of tissue damaging toxins, S. aureus infection does not elicit these BIRC5+ cells. Moreover, deletion of BIRC5 from CX3CR1 expressing cells results in improved survival during S. aureus infection. Hence, the combination of scRNA-Seq and genetic fate mapping CX3CR1+ cells revealed a toxin dependent pathogenic role for BIRC5 in myeloid cells during S. aureus infection.

overall_design: Single cell suspensions were obtained from livers . For each data set, cells isolated from 2-4 mice were pooled together, with equal numbers of cells pooled from each animal used. 12,000 cells from each group were loaded on a 10X Genomics Next GEM chip and single-cell GEMs were generated on a 10X Chromium Controller

first: Denis

middle: G

last: Loredan

first: Joseph

last: Devlin

first: Keenan

middle: A

last: Lacey

first: Erin

last: Zwack

first: Jian-Da

last: Lin

first: Kelly

middle: V

last: Ruggles

first: Kamal

last: Khanna

first: Victor

last: Torres

first: P’ng

last: Loke


title: DNA methylation profiling to determine the primary sites of metastatic cancers using formalin-fixed paraffin-embedded tissues [Test Set]

status: Public on Jun 11 2023

city: Shanghai

first: Leiqin

last: Liu

address: 320 Yueyang Road

country: China

institute: CAS Center for Excellence in Molecular Cell Science (CEMCS)

type: Methylation profiling by high throughput sequencing

summary: Accurate identification of the primary site of metastatic cancer is critical to guide the subsequent treatments. There is a significant portion of patients whose primary sites are initially classified as uncertain, and 3-9% of cancer patients are diagnosed with cancer of unknown primary (CUP) even after comprehensive diagnostic workups. Yet, a widely accepted molecular test is still not available. Here, we presented the combination of a novel DNA methylation sequencing-based method and an algorithm to predict the tissues of origin for metastatic cancers. The assay applied degraded DNA from formalin-fixed, paraffin-embedded (FFPE) tissues to generate reduced represent bisulfite sequencing libraries (FFPE-RRBS). Comparable DNA methylation metrics were obtained for the paired fresh frozen (FF) RRBS and FFPE-RRBS libraries and the FFPE-RRBS libraries of matched primary and metastatic cancer tissues. We generated and systemically evaluated 28 molecular classifiers built on four methylation evaluation methods and seven machine-learning approaches from a training data set of 498 primary cancer patients. Of those classifiers, the beta values-based (mean methylation) linear support vector (BELIVE) performed the best, achieving overall accuracies of 81-95% for identifying the primary sites of 215 metastatic cancer patients by utilizing the top-k predictions (k=1, 2, 3). The prediction accuracies ranged from 92% to 98% for 4702 patients with primary tumors in a cross-validation cohort. Lastly, BELIVE successfully identified the tissues of origin for approximately 81-93% of cases in a cohort of 68 patients initially diagnosed with CUP.

overall_design:
We sequenced RRBS libraries from 503 samples of primary tumor, 218 samples of metastatic tumor, and 68 samples of CUP patients.The assay was conducted using genomic DNA purified either from 10-20 mg of FF tissues or five to eight 5-10 µm FFPE tissue sections. Board-certified pathologists checked one H&E stained slide to ensure that tumor cells accounted for 10% or more cell population and that the necrosis area was less than 50%.
NOTE FROM SUBMITTER: Raw data (FASTQ files) are controlled to protect patient privacy.

first: Shutao

last: He


title: Effect of Inflammatory cytokines primed hUC-MSCs therapy on gene expression of macrophages in IUA-like mice model

status: Public on Jun 12 2023

city: Nanjing

first: Simin

last: Yao

address: 321 Zhongshan Road

institute: The Affiliated Drum Tower Hosipital, Medical School, Nanjing University

type: Expression profiling by high throughput sequencing

summary:
Intrauterine adhesion (IUA), characterized by endometrial fibrosis, is a common cause of female reproductive disorders. IUA-like mice model was created with curettage plus LPS stimulation to mimic endometrial fibrosis in IUA patients. Mesenchymal stem cells (MSCs) based therapy is applied to some refractory immune diseases to control inflammation and their immunomodulatory function can be enhanced via pre-treatment with inflammatory cytokines. In this work, we examined the effect of untreated and cytokines primed MSCs therapy on macrophage in IUA-like mice model.
The uteri samples were collected from sham, IUA modeling, IUA+control MSCs treatment and IUA+primed MSCs treatment groups of mice. Endometrial macrophages were sorted through F4/80 magnetic beads and performed RNA-seq.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for macrophages sorted from mice uteri with 3 sham operation (Sham), 3 IUA modeling (Matrigel), 3 IUA+control MSCs treatment (Ctrl) and 3 IUA+primed MSCs treatment (Primed).

first: Yali

last: Hu


title: DNA methylation profiling to determine the primary sites of metastatic cancers using formalin-fixed paraffin-embedded tissues [TrainingSet]


title: DNA methylation profiling to determine the primary sites of metastatic cancers using formalin-fixed paraffin-embedded tissues [ValidationSet]


title: DNA methylation profiling to determine the primary sites of metastatic cancers using formalin-fixed paraffin-embedded tissues

overall_design: We sequenced RRBS libraries from 503 samples of primary tumor, 218 samples of metastatic tumor, and 68 samples of CUP patients.The assay was conducted using genomic DNA purified either from 10-20 mg of FF tissues or five to eight 5-10 µm FFPE tissue sections. Board-certified pathologists checked one H&E stained slide to ensure that tumor cells accounted for 10% or more cell population and that the necrosis area was less than 50%.


title: Transcriptional response to standard AML drugs identifies synergistic combinations

city: Mainz

first: Piyush

last: More

address: Langenbeckstrasse 1

department: Department of Pharmacology

country: Germany

institute: University Medical Center, Johannes Gutenberg University of Mainz

summary: Unlike genomic alterations, gene expression profiles have not been widely used to refine cancer therapies. We analyzed transcriptional changes in acute myeloid leukemia (AML) cell lines in response to standard first-line AML drugs cytarabine and daunorubicin by means of RNA sequencing. Those changes were highly cell- and treatment-specific. By comparing the changes unique to treatment-sensitive and treatment-resistant AML cells, we enriched for treatment-relevant genes. Those genes were associated with drug response-specific pathways, including calcium ion-dependent exocytosis and chromatin remodeling. Pharmacological mimicking of those changes using EGFR and MEK inhibitors enhanced the response to daunorubicin with minimum standalone cytotoxicity. The synergistic response was observed even in the cell lines beyond those used for the discovery, including a primary AML sample. Additionally, publicly available cytotoxicity data confirmed the synergistic effect of EGFR inhibitors in combination with daunorubicin in all 60 investigated cancer cell lines. In conclusion, we demonstrate the utility of treatment-evoked gene expression changes to formulate rational drug combinations. This approach could improve the standard AML therapy, especially in older patients.

overall_design: RNA-Seq-based gene expression profiling of 3 AML cell lines (HL60, MOLM13, and OCIAML3) after treatment with DMSO, Cytarabine (0.1 µM), Daunorubicin (0.02 µM), and the combination of Cytarabine and Daunorubicin using 3 biological replicates.


title: Inflammatory cytokines pre-treatment enhances the immunomodulatory property of hUC-MSCs

summary: Mesenchymal stem cells (MSCs) exist in almost all tissues and participate in tissue regeneration and homeostasis. MSCs based therapy is applied to some refractory immune diseases to control inflammation, such as lupus nephritis, Crohn’s disease and rheumatoid arthritis. However, accumulating studies showed that the immunomodulatory capacity of naïve MSCs is mild and limited. To enhance MSCs immunomodulatory function, researchers innovated a new method to reprogram MSCs via pre-treatment with inflammatory cytokines. In this work, we firstly used a cocktail of three cytokines, IL-1β, TNF-a and IFN-γ, to treat hUC-MSCs (human MSCs from umbilical cord). We then performed gene expression profiling analysis using data obtained from RNA-seq of 3 untreated hUC-MSCs (ctrl) and 3 cytokines-treated hUC-MSCs (primed).

overall_design: Comparative gene expression profiling analysis of RNA-seq data for 3 untreated hUC-MSCs (ctrl) and 3 inflammatory cytokines treated hUC-MSCs (primed).


title: Optimized bisulfite sequencing analysis reveals the lack of 5-methylcytosine in mammalian mitochondrial DNA

city: shanghai

first: Zhenyu

last: Shao

state: --- Select a state ---

address: Yueyang Road 320

department: Center for Excellence in Molecular Cell Science

institute: Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences

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

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Refer to individual Series


title: MOF-mediated histone H4 lysine 16 acetylation governs mitochondrial and ciliary functions by controlling promoter interactions (ChIP-Seq)

city: Chicago

first: Rui

last: Yi

state: IL

address: 303 E. Chicago Avenue

country: USA

institute: Northwestern University

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Histone H4 lysine 16 acetylation (H4K16Ac), governed by the histone acetyltransferase (HAT) MOF, orchestrates critical functions in gene expression regulation and chromatin interaction. In this study, we show that conditional genetic deletion of Mof but not Kansl1, the essential component of the NSL complex, causes severe defects during murine skin development. Single-cell and bulk RNA-seq, in combination with MOF ChIP-seq, reveal that numerous MOF targeted and downregulated genes are highly enriched in mitochondria and cilia. Genetic deletion of Uqcrq, an essential subunit for electron transport chain Complex III, recapitulates the defects observed in MOF cKO. Single-cell ATAC-seq reveals that MOF targeted genes are controlled prominently through promoter interactions.

overall_design: ChIP-Seq

first: Dongmei

last: Wang

first: Haimin

last: Li

first: Navdeep

last: Chandel

last: Dou


title: Optimized bisulfite sequencing reveals the lack of 5-methylcytosine in mammalian mitochondrial DNA [TAPS]

summary: Whether 5mC exists in mitochondrial DNA is controversial, as data ranging from lack of 5mC to very extensive 5mC have been reported. By comprehensive bioinformatic analyses of published and our own data, we reveal that the observation of extensive and strand-biased mtDNA-5mC is artifact due to combinatorial effects of inefficient bisulfite conversion, extremely low sequencing reads in the L strand and interfering signals from nuclear mitochondrial DNA sequences (NUMTs).

overall_design: DNA methylation analysis of TAPS data for listed cells and tissues.


title: MOF-mediated histone H4 lysine 16 acetylation governs mitochondrial and ciliary functions by controlling promoter interactions


title: Optimized bisulfite sequencing reveals the lack of 5-methylcytosine in mammalian mitochondrial DNA [WGBS]

overall_design: DNA methylation analysis of WGBS data for listed cells and tissues.


title: PD-L1 checkpoint blockade promotes regulatory T-cell suppressive capacity underlying immunotherapy resistance

city: Rotterdam

first: Anneloes

last: van Krimpen

state: Zuid-Holland

address: Doctor Molewaterplein 40

department: Pulmonary Medicine

country: Netherlands

institute: Erasmus MC

summary: Transcriptome analyses of isotype-treated and aPD-L1-treated CD4+ T-cells, PD1- Treg cells and PD1+ Treg cells from a mouse model for mesothelioma.

overall_design: Biological replicated (n= 2-3) RNA-seq experiments

first: Ralph

last: Stadhouders

first: Mandy

middle: L

last: van Gulijk


title: Cellular profiling of a recently-evolved social behavior [snRNA-seq]

city: Atlanta

first: Larry

last: Young

state: GA

address: 954 Gatewood Rd NE

department: Department of Psychiatry

institute: Emory University

summary: Social behaviors are essential for survival and reproduction and vary strongly among individuals, species, and heritable brain diseases. The molecular and cellular bases of this variation are poorly resolved, and discovering them is necessary to understand how neural circuit and behavioral functions—and dysfunctions—vary in social contexts. Here we integrate single nucleus RNA-sequencing (snRNA-seq) with comparative genomics and automated behavior analysis to investigate the neurobiology of castle-building, a recently-evolved social, spatial, goal-directed, and repetitive construction behavior in Lake Malawi cichlid fishes. We simultaneously control for and analyze two biological variables correlated with castle-building behavior: quivering, a courtship “dance” behavior, and relative gonadal mass. We find signatures of building-, quivering-, and gonadal-associated neuronal excitation, gene expression, and neurogenesis in distinct cell populations. Converging lines of evidence support the involvement of estrogen, TrkB, and CCK signaling systems, and specific pallial excitatory neuronal subpopulations, in castle-building behavior. We show additional evidence that castle-building has evolved in part through genomic divergence in a gene module that is selectively expressed in stem-like quiescent radial glial cells (RGCs) lining the ventricular zone of the pallium. This RGC subpopulation exhibits signatures of a building-associated departure from quiescence, which in turn is associated with neuronal rebalancing in the putative fish homologue of the hippocampus. Our work supports an unexpected role for glia and neurogenesis in the evolution of social behavior, and more broadly shows how snRNA-seq can be used to systematically profile the cellular bases of previously unstudied social behaviors in new species systems.

overall_design: Single nuclei RNA seqeuncing was performed on nuclei isolated by Fluorescence-activated cell sorting (FACS) from the telencephalon of Mchenga conophoros during bower-building behavior and ‘control’ fish that were not building.

first: Zachary

last: Johnson

first: Brianna

last: Hegarty

first: George

last: Gruenhagen

first: Tucker

last: Lancaster

first: Patrick

last: McGrath

first: Jeffrey

last: Streelman


title: Spatial transcriptomics reveals a conserved segment polarity program that governs muscle patterning in Nematostella vectensis

city: Kansas City

first: Shiyuan

middle: (Cynthia)

last: Chen

state: MO

department: Computational Biology

institute: Stowers Institute for Medical Research

summary: During early animal evolution, the emergence of axially-polarized segments was central to the diversification of complex bilaterian body plans.  Nevertheless, precisely how and when segment polarity pathways arose remains obscure.  Here we demonstrate the molecular basis for segment polarization in developing larvae of the pre-bilaterian sea anemone Nematostella vectensis.  Utilizing spatial transcriptomics, we first constructed a 3-D gene expression atlas of developing larval segments.  Capitalizing on accurate in silico predictions, we identified Lbx and Uncx, conserved homeodomain-containing genes that occupy opposing subsegmental domains under the control of both BMP signaling and the Hox-Gbx cascade. Functionally, Lbx mutagenesis eliminated all molecular evidence of segment polarization at larval stage and resulted in an aberrant mirror-symmetric pattern of retractor muscles in primary polyps. These results demonstrate the molecular basis for segment polarity in a pre-bilaterian animal, suggesting that polarized metameric structures were present in the Cnidaria-Bilateria common ancestor over 600 million years ago.

overall_design:
A total of 24 samples were analyzed to identify DEGs under different Hox mutant background. 3 mutant strains (Anthox1a, Anthox8 and Anthox6a) were collected at three different developmental time points (48hpf, 60hpf and 72hpf) with 2 replicates each. 2 replicates of wildtype animals that were spawned on the same date as mutants were collected in parallel as controls.
A total of 6 samples were analyzed to identify DEGs after knocking down Pbx using shRNA. 2 independent shRNAs targeting Pbx were injected into wildtype oocytes. 1 scrambled control shRNA was injected into wildtype oocytes as control. 2 replicates per shRNA were collected at 72hpf.
1 single cell RNA-seq sample was analyzed to generate the Endo-atlas.

first: Shuonan

last: He

first: Wanqing

first: Ting

first: Matthew

middle: C

last: Gibson


title: Analysis of the organization and expression patterns of the convergent lasR/rsaL gene pair uncovers mutual influence

city: Braunschweig

first: Matthias

last: Preusse

address: Inhoffenstr. 7

institute: Helmholtz Centre for Infection Research

summary: The two adjacent genes encoding the major quorum sensing regulator, LasR, and its opponent, RsaL, overlap in their coding 3´ends and produce mRNA transcripts with long untranslated 3´ends that overlap with the sense transcripts of the gene on the opposing DNA strand. In this study, we evaluated whether the overlapping genes are involved in mutual regulatory events, such as transcriptional interference and interference by natural antisense transcripts. We introduced various gene expression constructs into a P. aeruginosa PA14 lasR/rsaL double deletion mutant, and found that although complementary RNA is produced, this does not interfere with the sense gene expression levels of lasR and rsaL and does not have functional consequences on down-stream gene regulation. Nevertheless, expression of lasR, but not of rsaL, was shown to be further enhanced if transcription was terminated at the end of the respective gene and no overlapping transcription with the gene on the opposite strand was allowed. Our data indicate that the natural organization with a partial overlap at the 3´ends of the lasR/rsaL genes gives rise to a system of checks and balances to prevent dominant and unilateral control by LasR over the RsaL transcriptional regulator of opposing function.

overall_design: RNASeq of different Pseudomonas aeruginosa PA14 lasR and rsaL mutants - planctonic growth in LB medium.

first: Silvia

last: Schinner

first: Susanne

last: Häussler


title: b1 integrin regulates epithelial proliferation and cell shape during alveolar repair

status: Public on Jun 10 2023

city: Nashville

first: Jennifer

last: Sucre

state: TN

address: 2215 B Garland Ave, 1125 Light Hall

department: Division of Neonatology, Department of Pediatrics

institute: Vanderbilt University Medical Center

summary: Integrins are extracellular matrix receptors comprised of an a and b subunit that connect and mediate signaling between cells and the surrounding matrix. In organogenesis of epithelial tissues, the b1 integrin subunit regulates essential epithelial cell functions, but the role of b1 integrin in epithelial repair is poorly understood. To define the role of b1 integrin during alveolar repair, we challenged b1 integrin deficient mice with intratracheal lipopolysaccharide, resulting in increased mortality with emphysematous lungs 21 days following injury. The alveolar barrier was repopulated with an overabundance of type 2 alveolar epithelial cells, with reduced numbers of elongated alveolar type 1 cells, suggesting b1 integrin is required for type 2 to type 1 epithelial transition. Consistent with this finding, b1 deficient type 2 epithelial cells proliferated at increased rates throughout repair, lacked actin-rich cellular protrusions necessary for lateral cellular extension, and exhibited transcriptomic dysregulation of adherens junction and actin polymerization pathways. Finally, we show that b1 integrin balances actin polymerization versus stabilization through GTPase activation. Taken together, these data support a novel role for b1 integrin in re-establishing the alveolar niche after injury through modulation of type 2 epithelial cell proliferation and cytoskeletal-dependent cell shape change.

overall_design: Single Cell RNAseq of dissociated adult mouse lungs from either Control (Itgb1 fl/fl) or epithelial iKO Itgb1 mice (Itgb1 iKO). Mice were treated with PBS or LPS, then sequenced 7 or 21 days later. Suspensions were enriched for cells that were CD45 negative, Ter119 negative, and viable. Lungs at 21 days post treatment were enriched for epithelial cells by collecting EPCAM positive cells.

middle: M

first: Fabian

last: Bock

first: Nicholas

last: Negretti

first: Xinyu

last: Dong

first: John

middle: T

last: Benjamin

first: Peter

last: Gulleman

first: Kimberly

last: Ferguson

first: Christopher

middle: S

last: Jetter

first: Wei

last: Han

first: Yang

last: Liu

first: David

last: Nichols

first: Seunghyi

last: Kook

first: Jonathan

last: Kropski

first: Timothy

last: Blackwell

first: Roy

last: Zent

first: Erin

middle: J

last: Plosa


title: An intact S-layer is advantageous to Clostridioides difficile within the host.

city: Glasgow

middle: Joseph

last: Cole

address: University Avenue

country: United Kingdom

institute: University of Glasgow

summary: The S-layer of C. difficile is a paracrystalline array that covers the bacterial cell but its contribution to overall disease remains unclear. A previously described spontaneous slpA-null mutant, FM2.5, with a point mutation in slpA offers the opportunity to study the role of the S-layer in vivo. Here, we confirm our previous observation that this strain is less virulent in vivo despite effectively colonising the host and producing toxin.  We also show a strong in vivo selection pressure for strains that express the intact S-layer array through sequence modifications that restore slpA expression. While such modifications do not affect the overall S-layer structure, RNA-Seq analysis in vitro showed large differences in gene expression between FM2.5, the revertant strains and R20291. Despite these differences, an intact S-layer clearly provides a selective advantage of C. difficile in the host, and, for one of the revertants tested, helped restore disease severity.

overall_design: Bulk RNA-seq and DNA-Seq of Clostridioides difficile strains FM2-5, R20291, Rv9, Rv189 and RW2-5.

first: Gillian

last: Douce

first: Michael

last: Ormsby


title: MacroH2A restricts melanoma progression via inhibition of chemokine expression in cancer-associated fibroblasts [spatial transcriptomics]

status: Public on Jun 09 2023

city: New York

first: Dan

last: Filipescu

state: New York

address: 1470 Madison Ave

department: Oncological Sciences

institute: Icahn School of Medicine at Mount Sinai

summary: MacroH2A has tumor suppressive functions in melanoma and other cancers, yet its role in the tumor microenvironment remains unappreciated. Using an autochthonous, immunocompetent mouse model of melanoma, we demonstrate that mice devoid of macroH2A variants exhibit increased tumor burden compared to wild type counterparts. MacroH2A-deficient tumors accumulate immunosuppressive monocytes and decreased functional cytotoxic T cells, and consistent with this compromised anti-tumor response, exhibit upregulation of pro-inflammatory cytokines. Single cell transcriptomics not only identifies cancer-associated fibroblasts (CAFs), whose frequency and activation increase in the absence of macroH2A, as the source of myeloid chemoattractants but also demonstrates dedifferentiation along the neural crest lineage of the tumor compartment. CAFs lacking macroH2A display hyper-inducible inflammatory gene expression and promoters of these genes present increased chromatin looping to enhancers that gain H3K27ac. In sum, we reveal a tumor suppressive role for macroH2A variants through regulation of chromatin architecture of inflammatory genes in the tumor stroma with potential implications for human melanoma.

overall_design: Spatial transcriptomics was carried out in WT and macroH2A deficient melanomas to map the localization of cell types previously identified by scRNA-seq.

first: Emily

last: Bernstein


title: Transcriptome analysis of different tissues in different materials

city: Guiyang

first: kai

last: pi

state: Guizhou

department: Key Laboratory for Tobacco Quality Research Guizhou Province

institute: Guizhou University

summary: We used two hybrid combinations and their 4 parents as materials. One week after topping the tobacco plant, samples were taken, including three tissues: root, stalk, and leaf. The RNA of these samples was extracted, and transcriptome sequencing analysis was carried out to explore the transcriptional profile differences between hybrids and their parents, and to analyze the gene differential expression between different tissues.

overall_design: Fifty-four samples were analyzed, including six plant materials,three plant tissues, each with three replicates.

first: Kai

last: Pi


title: MacroH2A restricts melanoma progression via inhibition of chemokine expression in cancer-associated fibroblasts [scRNA-seq]

summary: The histone variant macroH2A has been implicated as a tumor suppressor in melanoma and other cancers, yet its role in the tumor microenvironment remains unappreciated. Here we show that mice constitutively lacking macroH2A variants exhibit increased melanoma tumor burden compared to their wild-type counterparts, which is associated with an altered intratumoral immune cell compartment. MacroH2A-deficient tumors display an accumulation of immunosuppressive monocytes and decreased functional cytotoxic T cells. Consistent with this compromised anti-tumor response, macroH2A-deficient tumors displayed upregulation of chemokines such as Ccl2, Cxcl1 and Il6. Through single cell transcriptomics of the entire melanoma microenvironment, we identified the source of these pro-tumor myeloid chemoattractants as cancer-associated fibroblasts, whose frequency and activation were increased in the absence of macroH2A. Chemokine gene expression was hyper-inducible in the absence of macroH2A and accompanied by an altered epigenetic landscape. In sum, we uncovered a tumor suppressive role for macroH2A through repression of chemokine induction in the tumor stroma.

overall_design: Single cell profiling of gene expression was carried out in WT and macroH2A deficient melanomas was carried out to identify cell types involved in increased tumor growth and chemokine overexpression in the absence of macroH2A.


title: Genome-wide maps of ARID1A binding genes in H9 human embryonic stem cells

city: Indianapolis

first: Lei

last: Yang

state: Indiana

address: 1044 W. Walnut St.

department: Department of Pediatrics

institute: Indiana University School of Medicine

summary: We reported loss of ARID1A promoted neurogenesis and inhibited cardiogenesis. Here we used specific ARID1A antibody to pull down ARID1A binding genomic DNA in human embryonic stem cells, which let us know the potential genes regulated by ARID1A during neurogenesis and inhibited cardiogenesis. IgG was used as the control.

overall_design: Examination of ARID1A binding genes in hESCs.

first: Juli


title: Targeting PGLYRP1 promotes anti-tumor immunity while inhibiting autoimmunity

city: Boston

first: Linglin

last: Huang

state: MA

address: 60 Fenwood Road, HBTM 10016F

department: Neurology

institute: Brigham and Women’s Hospital


title: Mapping H3K4me3 in in vitro differentiated resident memory CD8+ T cells

city: Tampa

first: Dorina

last: Avram

state: Fl

address: 12902 Magnolia Drive, MRC4

department: Department of Immunology

institute: Moffitt Cancer Center

summary: The networks of transcription factors (TFs) that control multipotency versus effector programs in intestinal resident memory T (TRM) cells are poorly understood. Mice with post-activation, conditional deletion of the TF Bcl11b in CD8+ T cells, infected with a food-born pathogen, had increased numbers of intestinal TRM cells, and their precursors and decreased splenic effector cells and circulating memory cells and precursors. Loss of circulating memory cells was in part due to increased intestinal homing of Bcl11b-/- circulating precursors with no major alterations in their programs. Bcl11b-/- memory CD8+ T cells had an impaired recall response despite their accumulation in the gut. Intestinal Bcl11b-/- TRM cells and their precursors manifested major alterations in the residency program, with diminished expression of multipotency program genes and upregulation of the effector program genes. Integration of transcriptomics with chromatin accessibility, activating histone marks and Bcl11b genome binding showed a link between the reduction in the multipotent program genes with regions of decreased chromatin accessibility and activating histone marks in Bcl11b-/- cells. In contrast, the effector program genes displayed increased activating epigenetic status. We propose that Bcl11b regulates tissue resident TRM program genes and is positioned upstream of Tcf1 and Blimp1 in regulation of multipotency versus effector TRM program, respectively. Rescuing experiments normalized the increased numbers of intestinal Bcl11b-/- TRM cells. Thus, Bcl11b is a frontrunner in the memory tissue residency program and acts early in lineage decision, promoting TRM cell multipotency and restricting effector function.

overall_design: Examination of H3K4me3 deposition in wild type and Bcl11b-deficient CD8+ Trm-like cells

first: Tomas

last: Zelenka

first: Valeriu

middle: B

last: Cismasiu

middle: I

last: Shaw


title: Mapping Tcf1 in in vitro differentiated resident memory CD8+ T cells

overall_design: Examination of Tcf1 binding in wild type and Bcl11b-deficient CD8+ Trm-like cells


title: Mapping H3K27ac in in vitro differentiated resident memory CD8+ T cells

overall_design: Examination of H3K27ac deposition in wild type and Bcl11b-deficient CD8+ Trm-like cells

first: Eric

last: Helm

first: Xianhong

last: Chen


title: Differential expression of lncRNA in peripheral blood lymphocytes of Xinjiang Kazakh patients with essential hypertension

city: Changchun

first: Tongjian

last: Zhao

address: 1266 Fujin Road

institute: School of Pharmaceutical Sciences, Jilin University

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

summary: Investigate long non-coding RNA (lncRNA) expression characteristics in the peripheral blood lymphocytes of Xinjiang Kazakh people with essential hypertension.

overall_design: Six Kazakh patients with essential hypertension and six Kazakh healthy participants were randomly selected. After detecting the expression levels of lncRNA in the peripheral blood lymphocytes using gene chip technology, their levels in the hypertensive group were compared with those in the control group.

first: Yan

last: Wang

first: Ketao

last: Ma


title: MAVS signaling is required for preventing persistent chikungunya heart infection and chronic vascular tissue inflammation

first: Franco

last: Izzo

state: NY

address: 1470 Madison Avenue

summary: Chikungunya virus (CHIKV) infection has been associated with severe cardiac manifestations. However, how CHIKV infection leads to heart disease remains unknown. Here, using C57BL/6 mice we show that cardiac fibroblasts are the main target of CHIKV infection in the heart, and that cardiac tissue infection induces activation of the type I interferon (IFN-I) pathway in both infected and non-infected cardiac cells. The loss of IFN-I signaling results in increased CHIKV infection, virus spreading, and apoptosis in cardiac tissue. Importantly, signaling through the mitochondrial antiviral-signaling protein (MAVS) is essential for efficient CHIKV clearance from the heart. Indeed, persistent infection in cardiac tissue of MAVS-deficient mice leads to cardiac damage characterized by focal myocarditis and major vessel vasculitis. This study provides mechanistic insight on how CHIKV can lead to cardiac damage, underscoring the importance of monitoring cardiac function in patients with CHIKV infections.

overall_design: For bulk RNA-sequencing male mice were infected intravenously with 10^5 PFU of CHIKV Indian Ocean lineage (IOL) or HI control, and heart were collected at 2 and 5 dpi (days post-infection). Hearts were collected in 500 μl of TRIzolTM (Invitrogen) with one 5 mm stainless steel bead and processed as described above. RNA was extracted using Zymo direct-zol RNA purification kit (Zymo research, #R2072) according to the manufacturer’s instructions. A total of 500 ng per sample was sent to the NYU Genome Technology Center for quality control assessment, automated TruSeq stranded total RNA, with RiboZero Gold library preparation and Illumina Novaseq6000 sequencing (single pair 100).

first: Maria

last: Noval

first: Kenneth

last: Stapleford


title: Mapping Ahr in in vitro differentiated resident memory CD8+ T cells

overall_design: Examination of Ahr binding in wild type and Bcl11b-deficient CD8+ Trm-like cells

first: Timothy


title: Cas9-Mediated Knockout of Ndrg2 Enhances the Regenerative Potential of Dendritic Cells for Wound Healing

city: Tucson

first: Kellen

state: AZ

address: 1501 N Campbell Ave

department: Surgery

institute: University of Arizona

summary: We performed single-cell RNA sequencing on bone marrow derived, cultured dendritic cells after CRISPR-knockout of Ndrg2, after vitamin D3 treatment or no treatment. In addition we performed single-cell RNA sequencing of wounds that had been treated with hydrogels seeded with Ndrg2-KO dendritic cells, control (untreated) dendritic cells or blank hydrogels.

overall_design: scRNA-seq was performed on murine cells collected from parabiome excisional wounds. Cells labeled as GFP+ were isolated and sequenced to analyze cells in the circulation that hone to the wound upon treatment.

first: Dominic

last: Henn

first: Dehua

first: Dharshan

last: Sivaraj

first: Stanley

last: Qi

first: Geoffrey

last: Gurtner


title: SELF-PATTERNING OF HUMAN STEM CELLS INTO POST-IMPLANTATION LINEAGES

city: New Haven

first: Berna

last: Sozen

state: CT

address: 333 Cedar St

department: Department of Genetics

institute: Yale School of Medicine

summary: Investigating human development is a significant scientific challenge due to the technical and ethical limitations of working with bona fide embryo samples. In the face of these difficulties, stem cells have been applied to experimentally model inaccessible stages of human development in vitro. Here we show that human pluripotent stem cells can be triggered to self-organise into three-dimensional structures that recapitulate the spatiotemporal events of early human post-implantation embryonic development. We report single-cell transcriptomics that confirms differentiation into diverse cell states of the native gastrulating human embryo, including signatures of post-implantation epiblast, amniotic ectoderm, primitive streak, mesoderm, and hypoblast, as well as pre-patterning of non-neural ectoderm and yolk sac. Collectively, our system captures key features of human embryonic development spanning from Carnegie-stage13 (CS) 4 to CS7, providing a reproducible, tractable, and flexible experimental platform to understand the basic cellular and molecular mechanisms that underlie human development, and a high-throughput platform to dissect developmental disorders and congenital pathologies.

overall_design: Human pluripotent cells spontaneously differentiated in 3D culture platform into structures called "human extraembryoid (hEE)". The hEEs collected for single cell RNA sequencing either at Day 4 or Day 6 in vitro

first: Nicolas

last: Dias


title: Globally reduced N6-methyladenosine (m6A) in C9ORF72-ALS/FTD dysregulates RNA metabolism and contributes to neurodegeneration

city: Shanghai

first: Xiaoyang

last: Dou

address: 320 Yue Yang Road

institute: CAS Center for Excellence in Molecular Cell Science, CAS

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

overall_design: Refer to individual Series


title: MacroH2A restricts melanoma progression via inhibition of chemokine expression in cancer-associated fibroblasts [3D chromatin]

first: Dan

last: Filipescu

overall_design: 3D chromatin conformation analysis by Micro-C followed by promoter capture (pcMicro-C), in 2 replicates each in WT and dKO cultured CAFs

first: Emily

last: Bernstein


title: MacroH2A restricts melanoma progression via inhibition of chemokine expression in cancer-associated fibroblasts [epigenomics]

overall_design: Epigenomic analysis of sorted or cultured CAFs from WT and macroH2A melanomas was carried out to understand how macroH2A regulates inflammatory gene expression in this cell type. iDFs were used as a related cell type, developed individually, to validate these findings.


title: MacroH2A restricts melanoma progression via inhibition of chemokine expression in cancer-associated fibroblasts.


title: Context-defined cancer co-dependency mapping identifies a functional interplay between PRC2 and MLL-MEN1 complex in lymphoma

first: Chao

last: Lu

address: 1130 Saint Nicholas Ave

department: Genetics & Development

institute: Columbia University Medical Center

summary: This work highlights the versatility of combining genetic screens with molecular profiling of cancer cells for high-throughput genetic interaction discovery that will not only offer critical insight into basic regulatory mechanisms of cancer epigenome progression, but also lead to new therapeutic targets for genetically defined and epigenetically perturbed cancers.

overall_design: 32 RNA-seq samples; 41 CUT&Tag samples; 4 CRISPR screen samples

first: Xiao


title: N6-methyladenosine regulates RNA metabolism and neurodegeneration in C9ORF72-ALS/FTD [seq_polyA_RNA-decay]

summary: N6-methyladenosine (m6A) is the most prevalent internal mRNA modification and regulates RNA metabolism. Repeat expansion in C9ORF72 is the most common genetic cause of amyotrophic lateral sclerosis (ALS) and frontotemporal degeneration (FTD). Here, we showed that m6A is downregulated in C9ORF72-ALS/FTD patient-derived iPSC-differentiated neurons and postmortem brain tissues. The global m6A hypomethylation leads to transcriptome-wide mRNA stabilization and upregulated gene expression, especially ones involved in synaptic activity and neuronal functions. The m6A modification in the C9ORF72 intron sequence preceding the expanded repeats enhances RNA degradation via the nuclear reader YTHDC1. The m6A reduction leads to increased accumulation of repeat RNA and poly-dipeptides. Moreover, the antisense RNA can also be regulated by m6A. Elevating m6A level significantly reduces both sense and antisense repeat RNA and poly-dipeptides, rescues global mRNA homeostasis, and improves survival of C9ORF72-ALS/FTD patient neurons.

overall_design: The neurons at day 32 of differentiation were treated with actinomycin D to halt de novo transcription, and RNAs were collected 0, 3, and 6 hours after treatment for RNA-seq to quantify the transcriptome-wide mRNA half-life.

first: Yini

last: Li

first: Chuan

last: He

first: Shuying

last: Sun


title: Small Cell Lung Cancer Subtype Plasticity is Regulated by KDM6A/UTX [ChIP-Seq]

first: Leslie

last: Duplaquet

state: Massachusetts

address: 360 Longwood Ave

department: Medical Oncology

institute: DFCI

summary: Using a CRISPR-based autochthonous SCLC GEMM to study the consequences of KDM6A/UTX inactivation, we found that KDM6A inactivation induced plasticity from ASCL1 to NEUROD1 resulting in SCLC tumors with open chromatin at the NEUROD1 promoter that express both ASCL1 and NEUROD1. Mechanistically, KDM6A binds and maintains ASCL1 target genes in an active chromatin state with its loss decreasing H3K4me1 and increasing H3K27me3 at enhancers of neuroendocrine genes leading to a cell state that is primed for ASCL1 to NEUROD1 subtype switching. This work identifies KDM6A as an epigenetic regulator that controls ASCL1 to NEUROD1 subtype plasticity and provides a novel autochthonous SCLC GEMM to model ASCL1 and NEUROD1 subtype heterogeneity and plasticity, which is found in 35-40% of human SCLCs.

overall_design: ChIP-seq for Menin in a Kdm6a-WT mouse tumor derived cell lines (1014) CRISPR inactivated for KDM6A treated with VTP50469 or DMSO

first: Sarah

middle: N

last: Olsen

first: Matthew

last: Oser


title: Nr2f1 shapes mitochondria in the mouse brain unraveling novel insights into the neurodevelopmental disorder BBSOAS

city: Torino

first: Ivan

last: Molineris

state: TO

address: Via Nizza 52

department: Life Science and System Biology

country: Italy

institute: University of Turin

summary: The nuclear receptor Nr2f1 acts as a strong transcriptional regulator in embryonic and postnatal neural cells. In humans, its mutations cause the Bosch-Boonstra-Schaaf optic atrophy-intellectual syndrome (BBSOAS), a rare neurodevelopmental disorder characterized by multiple clinical features including optic nerve atrophy, intellectual disability, and autistic traits. In this study, by genome-wide and in silico analyses we identified a wide set of nuclear-encoded mitochondrial genes as potential genomic targets under direct Nr2f1 transcriptional control in neurons. By combining mouse genetics, neuroanatomical and imaging approaches we demonstrated that conditional Nr2f1 loss-of-function within the adult mouse hippocampal neurogenic niche results in a reduced mitochondrial mass associated with mitochondrial fragmentation and downregulation of key mitochondrial proteins in newborn neurons, whose functional integration and survival are impaired. Importantly, we also found dysregulation of several mitochondrial genes and downregulation in levels of key mitochondrial proteins in the brain of mice heterozygous for Nr2f1, a validated BBSOAS model. Our data point to an active role of Nr2f1 in the mitochondrial gene expression regulatory network in neurons and support the involvement of mitochondrial dysfunction in BBSOAS pathogenesis.

overall_design: 3 samples

first: Sara

last: Bonzano

first: Eleonora

last: Dallorto

first: Filippo

last: Michelon

first: Isabella

last: Crisci

first: Giovanna

last: Gambarotta

first: Francesco

last: Neri

first: Salvatore

last: Oliviero

first: Ruth

last: Beckervordersandforth

first: Dieter

last: Lie

first: Paolo

last: Peretto

first: Serena

last: Bovetti

first: Michèle

last: Studer

first: Silvia

last: De Marchis


title: RANK expression and Luminal BC response to CDK4/6 inhibitors

city: Lisbon

first: Sandra

last: Casimiro

address: Av. Prof. Egas Moniz

country: Portugal

institute: Instituto de Medicina Molecular

summary: The combination of endocrine therapy (ET) with CDK4/6 inhibitors (CDK4/6i) was the most recent life-changing hallmark in metastatic Luminal breast cancer (BC). However, intrinsic and acquired resistance affect long-term efficacy. Here, we studied the role of receptor activator of nuclear factor-kB (RANK) pathway in CDK4/6i resistance. We found that RANK overexpression in Luminal BC associates with intrinsic resistance to CDK4/6i, both in vitro and in mouse xenografts. Transcriptomic analysis of mouse tumors highlighted decreased proliferation rate and chronic interferon (IFN)-response as resistance drivers.

overall_design: The transcriptomic profiles of tumor tissue from MCF-7 and MCF-7 RANK OE xenografts implanted in NSG mice, collected after 21 days of treatment with Vehicle, Palbociclib+Fulvestrant or Palbociclib+Fulvestrant+OPG-Fc, were analyzed by RNASeq. 2-3 tumors per group were included.

first: Inês

last: Gomes

first: Luis

last: Costa


title: Mousemodel_Heptablastoma_ScRNAseq

city: Memphis

first: Xiang

last: Chen

address: 262 Danny Thomas Place

institute: St.Jude Children's Research Hospital

summary: Lack of relevant animal models and cell lines hampers the understanding of hepatoblastoma and identification of therapeutic targets. We report a hepatoblastoma murine model that faithfully recapitulates the pathological features of mixed fetal and embryonic hepatoblastoma, with transcriptomics resembling the high-risk gene signatures of human disease

overall_design: The Single nuclear RNAseq was performed using 10X genomics -Single cell gene expression kit V3.1.

first: Jun

last: Yang

first: John

last: Easton

first: Sivaraman

last: Natarajan

first: Jie

last: Fang


title: Loss of epigenetic suppression of retrotransposons with oncogenic potential in aging mammary luminal epithelial cells

city: Duarte

first: Dustin

middle: E

last: Schones

state: CA

address: 1500 E Duarte Rd.

institute: City of Hope

summary: Genome wide DNA methylation profiling of luminal epithelial cells obtained from reduction mammoplasty tissue from younger (<30 year) and older (>55 year) individuals.

overall_design: We performed whole genome bisufite sequencing on mammary luminal epithelial cells from young and old individuals.

first: Parijat

last: Senapati

first: Mark

last: LaBarge


title: Germ-cell specific eIF4E1b regulates maternal mRNA translation to ensure zygotic genome activation [Ribo-seq]

status: Public on Jun 08 2023

city: Bethesda

first: Guanghui

state: Maryland

address: 50 South Drive

department: National Institute of Diabetes and Digestive and Kidney Diseases

institute: National Institutes of Health

summary: Translation of maternal mRNAs is detected before transcription of zygotic genes and is essential for mammalian embryo development. How certain maternal mRNAs are selected for translation instead of degradation and how this burst of translation affects zygotic genome activation remains unknown. Using gene-edited mice, we document that the oocyte-specific eukaryotic translation initiation factor 4E family member 1b (eIF4E1b) is the regulator of maternal mRNA expression that ensures subsequent reprogramming of the zygotic genome. In oocytes, eIF4E1b binds to transcripts encoding translation machinery proteins, chromatin remodelers and reprogramming factors to promote their translation in zygotes and protect them from degradation. The protein products are thought to establish an open chromatin landscape in one-cell zygotes to enable transcription of genes required for cleavage stage development. Our results define a program for rapid resetting of the zygotic epigenome that is regulated by maternal mRNA expression and provides new insights into the mammalian maternal-to-zygotic transition.

overall_design: Low input ribo-seq profiling using eggs or embryos from Eif4e1b knockout or control female mice containing HA tagged Rpl22.

first: Qiliang

last: Xin

first: Jurrien

last: Dean


title: KDM6A Epigenetically Regulates Subtype Plasticity in Small Cell Lung Cancer

first: Leslie

last: Duplaquet


title: TONSOKU is required for the maintenance of repressive epigenetic modifications in Arabidopsis [BS-seq]

city: Beijing

first: Danhua

last: Jiang

address: No. 1 West Beichen Road, Chaoyang District

institute: Institute of Genetics and Developmental Biology, Chinese Academy of Sciences

summary: Epigenetic inheritance requires the faithful maintenance of epigenetic marks such as certain chromatin modifications through DNA replication. Plant TONSOKU (TSK) and its animal orthologue TONSOKU-like (TONSL) act as readers for newly synthesized histones and assist in DNA replication via facilitating DNA damage repair at post-replicative chromatin. However, whether TSK/TONSL regulates chromatin replication and the maintenance of chromatin modifications remain elusive. In this study, we demonstrate that TSK is dispensable for global histone and nucleosome accumulation but necessary for maintaining repressive chromatin modifications including H3K9me2, H2A.W, H3K27me3 and DNA methylation. TSK physically interacts with H3K9 methyltransferases SUVH5 and SUVH6, and the Polycomb protein RING1a/b. Moreover, TSK mutation strongly enhances defects in Polycomb pathway mutants. TSK is intended to only associate with nascent chromatin until it starts to mature. We propose that TSK ensures proper epigenetic inheritance by supporting the recruitment of epigenetic modifiers to post-replicative chromatin in a critical short window of time following DNA replication.

overall_design: Transcriptome, MNase, H3k27me3, H2A.W and H3K9me2 enrichment, and DNA methylation were examined in Col or tsk-4 mutant

first: Lin

last: Wang

first: Mande

last: Xue

first: Huairen

last: Zhang


title: RNA-seq of primary tumors from Braf/Pten/Cre APOE2 and APOE4 mice

first: Nneoma

last: Adaku

address: 1230 York Ave

institute: The Rockefeller University

summary: Genetic variation in the APOE gene modulates melanoma progression. To identify tumor cell-intrinsic mechanisms underlying differences in tumor progression between APOE2 and APOE4 genotypes, we crossed the Braf/Pten/Cre (BPC) genetically engineered mouse model with APOE2 (BPCE2) and APOE4 (BPCE4) knock-in mice. We performed RNA-Seq on tumors derived from these mice. This analysis revealed protein synthesis as a process that is enhanced by APOE2.  **Note: Raw fastq sequencing files for these samples were irrevocably corrupted due to a server failure and therefore could not be provided. Kallisto transcript abundance estimates have been included as processed data.**

overall_design: RNA-seq was conducted on 4 tumors each from BrafV600E/+;Pten fl/fl;Tyr-CreERT2;APOE2 (BPCE2) and BrafV600E/+;Pten fl/fl;Tyr-CreERT2;APOE4 (BPCE4) mice.

first: Sohail

last: Tavazoie


title: The mRNA methyltransferase Mettl3 modulates cytokine mRNA stability and limits functional responses in mast cells [m6A-CLIP-Seq]

city: Bellinzona

first: Cristina

last: Leoni

state: Ticino

address: Via F. Chiesa 5

department: Molecular Immunology

country: Switzerland

institute: Institute for Research in Biomedicine

summary: Mast cells are central players in allergy and asthma, and their dysregulated responses lead to reduced quality of life and life-threatening conditions such as anaphylaxis. The RNA modification N6-methyladenosine (m6A) has a prominent impact on immune cell functions, but its role in mast cells remains unexplored. Here, by optimizing tools to genetically manipulate primary mast cells, we revealed that the m6A mRNA methyltransferase complex modulates mast cell proliferation and survival. Depletion of the catalytic component Mettl3 exacerbated effector functions in response to IgE and antigen complexes, both in vitro and in vivo. Mechanistically, deletion of Mettl3 or Mettl14 led to the enhanced expression of inflammatory cytokines. By focusing on one of the most affected mRNAs, namely the one encoding the cytokine IL-13, we found that it was methylated in activated mast cells, and that Mettl3 affected its transcript stability in an enzymatic activity-dependent manner, requiring consensus m6A sites in the Il13 3’-untranslated region. Overall, we revealed that the m6A machinery is essential in mast cells to sustain growth and to restrain inflammatory responses.

overall_design: Bone marrow derived mast cells (BMMCs) in vitro differentiated from 4 mice were either left resting or stimulated with 2 nM PMA (Sigma) and 200 nM ionomycin for 2 h. Total RNA was isolated with Trizol.

first: Marian

last: Bataclan

first: Taku

last: Ito-Kurhea

first: Vigo

last: Heissmeyer

first: Silvia

last: Monticelli


title: Small Cell Lung Cancer Subtype Plasticity is Regulated by KDM6A/UTX [RNA-Seq]

summary: Using a CRISPR-based autochthonous SCLC GEMM to study the consequences of KDM6A/UTX inactivation, we found that KDM6A inactivation induced plasticity from ASCL1 to NEUROD1 resulting in SCLC tumors with open chromatin at the NEUROD1 promoter that express both ASCL1 and NEUROD1. Mechanistically, KDM6A binds and maintains ASCL1 target genes in an active chromatin state with its loss increasing H3K27me3 near their promoters leading to a cell state that is primed for ASCL1 to NEUROD1 subtype switching. This work identifies KDM6A as an epigenetic regulator that controls ASCL1 to NEUROD1 subtype plasticity and provides a novel autochthonous SCLC GEMM to model ASCL1 and NEUROD1 subtype heterogeneity and plasticity, which is found in 35-40% of human SCLCs.

overall_design: Comparative gene expression profiling analysis of RNA-seq data in mouse small cell lung cancer (SCLC) tumors that are KDM6A WT (sgControl RPP) or KDM6A-Mutant (sgKdm6a RPP).

first: Matthew

last: Oser


title: RNA-seq of B16F10 lung metastases derived from APOE2 and APOE3 mice

summary: Genetic variation in the APOE gene modulates melanoma progression. To identify tumor cell-intrinsic mechanisms underlying differences in metastasis between the APOE2 and APOE3 genotypes, we performed tail vein injections of luciferase and GFP-expressing (triple reporter) B16F10-shApoe melanoma cells into APOE2 and APOE3 knock-in mice. We performed RNA-Seq on lung metastatic nodules derived from these mice. This analysis revealed protein synthesis as a process that is enhanced by APOE2.

overall_design: RNA-seq conducted on purified lung metastatic nodules derived from tail vein injections of B16F10-TR-shApoe cells into APOE2 mice (n=8) and APOE3 mice (n=5).

first: Benjamin

last: Ostendorf


title: TONSOKU is required for the maintenance of repressive epigenetic modifications in Arabidopsis


title: The MOM1 complex recruits the RdDM machinery via MORC6 to establish de novo DNA methylation.

city: Los Angeles

first: Zhenhui

last: Zhong

address: 610 Charles E Young Dr East

department: Department of Molecular, Cell and Developmental Biology

institute: University of California, Los Angeles

summary: MOM1 is an Arabidopsis factor previously shown to mediate transcriptional silencing independent of major DNA methylation changes. Here we found that MOM1 localizes with sites of RNA-directed DNA methylation (RdDM). Tethering MOM1 with artificial zinc finger to unmethylated FWA promoter led to establishment of DNA methylation and FWA silencing. This process was blocked by mutations in components of the Pol V arm of the RdDM machinery, as well as by mutation of MORC6. We found that at some endogenous RdDM sites, MOM1 is required to maintain DNA methylation and a closed chromatin state.  In addition, efficient silencing of newly introduced FWA transgenes was impaired by mutation of MOM1 or mutation of genes encoding the MOM1 interacting PIAL1/2 proteins.  In addition to RdDM sites, we identified a group of MOM1 peaks at active chromatin near genes, again colocalized with MORC6.  These findings demonstrate a multifaceted role of MOM1 in genome regulation.

overall_design: This dataset includes ChIP-seq, RNA-seq, WGBS, ATAC-seq and BS-PCR to understand the role of MOM1 complex, especially the connection of MOM1 complex to the RdDM machinery and the MORC proteins.

first: Zheng

last: Li

first: Ming

first: Zhong

last: Zhenhui


title: N6-methyladenosine regulates RNA metabolism and neurodegeneration in C9ORF72-ALS/FTD [seq_total_RNA-MeRIP]

first: Xiaoyang

last: Dou

overall_design: MeRIP-seq was performed using control and C9-ALS patient iPSNs at day 32 of differentiation

first: Yini

first: Chuan

last: He

first: Shuying

last: Sun


title: Apilactobacillus kunkeei releases RNA-associated membrane vesicles and proteinaceous nanoparticles

city: Uppsala

first: Siv

last: Andersson

address: Husargatan 3

country: Sweden

institute: Uppsala University

summary: Extracellularly released particles, including membrane vesicles, have increasingly been recognized as important for bacterial community functions and host-interaction processes, but their compositions and functional roles differ between species and also between strains of the same species. In this study, we have determined the composition of membrane vesicles and protein particles identified in the cell-free pellets of two strains of Apilactobacillus kunkeei, a defensive symbiont of honeybees. The membrane vesicles were separated from the extracellular particles using density gradient ultracentrifugation. The peaks of the RNA and protein distributions were separated from each other and the highest concentration of RNA was observed in the fractions that contained the membrane vesicles while the highest protein concentration coincided with the fractions that contained extracellular particles. A comparative proteomics analysis by LC-MS/MS showed that 37 proteins with type-I signal peptides were consistently identified across the fractionated samples obtained from the cell-free pellets, of which 29 were orthologs detected in both strains. Functional predictions of the extracellular proteins revealed the presence of glycoside hydrolases, glycosyltransferases, giant proteins and peptidases. The extracellular transcriptomes mapped to a broad set of genes with a similar functional profile as the whole cell transcriptome. This study provides insights into the composition of membrane vesicles and extracellular proteins of a bee-associated symbiont.

overall_design: The RNA associated with cell-free pellets (CFP) from Apilactobacillus kunkeei strain A1401 was compared with RNA extracted from whole cell lysates (WCL). Biological replicates were prepared and paired CFP and WCL samples were obtained from each biological replicate.

first: Christian

last: Seeger

first: Karl

last: Dyrhage

first: Kristina

last: Näslund


title: N6-methyladenosine regulates RNA metabolism and neurodegeneration in C9ORF72-ALS/FTD [seq_polyA_RNA-RIP]

overall_design: YTHDF2 RIP was performed upon 5 individual human iPSN lines at day 22 of differentiation.


title: Pharmacological inhibition of lipolysis prevents adverse metabolic outcomes during glucocorticoid administration

city: Baton Rouge

first: Jaroslaw

last: Staszkiewicz

state: LA

address: 6400 Perkins Rd

department: Genomics Core Facility

institute: Pennington Biomedical Research Center

overall_design: Refer to individual Series


title: Skeletal muscle adaptations in response to continuous corticosterone exposure and pharmacological inhibition of lipolysis

summary: Purpose: to identify adaptations in skeletal muscle gene expression following two weeks of corticosterone treatment with and without ATGL inhibition.

overall_design: mRNA profiles from mixed gastrocnemius skeletal muscle of mice given ethanol (EtOH) or corticosterone (Cort) for two weeks. Subsets of each group were fed either a control diet (CON) or a diet that had Atglistatin (ATGLi).

first: Melissa

last: Linden

first: Heidi

last: Batdorf

first: Humza

last: Pirzadah

first: Jasleen

last: Kaur

first: Sara

last: Webb

first: Richard

last: Carmouche

first: Sujoy

last: Ghosh

first: Susan

last: Burke

first: J

last: Collier

first: Robert

last: Noland


title: MacroH2A restricts melanoma progression via inhibition of chemokine expression in cancer-associated fibroblasts [RNA-seq]

city: New York

first: Dan

last: Filipescu

address: 1470 Madison Ave

institute: Icahn School of Medicine at Mount Sinai

summary: The histone variant macroH2A has been implicated as a tumor suppressor in melanoma and other cancers, yet its role in the tumor microenvironment remains unappreciated. Here we show that mice constitutively lacking macroH2A variants exhibit increased melanoma tumor burden compared to their wild-type counterparts, which is associated with an altered intratumoral immune cell compartment. MacroH2A-deficient tumors display an accumulation of immunosuppressive monocytes and decreased functional cytotoxic T cells. Consistent with this compromised anti-tumor response, macroH2A-deficient tumors displayed upregulation of chemokines such as Ccl2, Cxcl1 and Il6. Through single cell transcriptomics of the entire melanoma microenvironment, we identified the source of these pro-tumor myeloid chemoattractants as cancer-associated fibroblasts, whose frequency and activation were increased in the absence of macroH2A. Chemokine gene expression was hyper-inducible in the absence of macroH2A and accompanied by an altered epigenetic landscape. In sum, we uncovered a tumor suppressive role for macroH2A through repression of chemokine induction in the tumor stroma.

overall_design: Gene expression analysis of bulk melanomas, melanoma-infiltrating CD8+ T cells and CAFs from WT and macroH2A deficient mice 50 days post induction was carried out to understand the molecular impact of macroH2A loss in the melanoma microenvironment.

first: Emily

last: Bernstein


title: Liver adaptations in response to continuous corticosterone exposure and pharmacological inhibition of lipolysis

summary: Purpose: to identify adaptations in hepatic  gene expression following two weeks of corticosterone treatment with and without ATGL inhibition.

overall_design: mRNA profiles from liver of mice given ethanol (EtOH) or corticosterone (Cort) for two weeks. Subsets of each group were fed either a control diet (CON) or a diet that had Atglistatin (ATGLi).


title: Small RNA sequencing analysis of peptide-affinity isolated plasma extracellular vesicles distinguishes pancreatic cancer patients from non-affected individuals

city: Moncton

first: Eric

middle: P

last: Allain

state: New Brunswick

address: 330 University ave

department: Medical Genetics

country: Canada

institute: Vitalité Health Network

type: Non-coding RNA profiling by high throughput sequencing

summary: Pancreatic ductal adenocarcinoma (PDAC) has a high fatality rate, mainly due to its asymptomatic nature until late-stage disease and therefore delayed diagnosis that leads to a lack of timely treatment intervention. Consequently, there is a significant need for better methods to screen populations that are at high risk of developing PDAC. Such advances would result in earlier diagnosis, more treatment options, and ultimately better outcomes for patients. Several recent studies have applied the concept of liquid biopsy, which is the sampling of a biofluid (such as blood plasma) for the presence of disease biomarkers, to develop screening approaches for PDAC; several of these studies have focused on analysis of extracellular vesicles (EVs) and their cargoes. While these studies have identified many potential biomarkers for PDAC that are present within EVs, their application to clinical practice is hindered by the lack of a robust, reproducible method for EV isolation and analysis that is amenable to a clinical setting. Our previous research has shown that the Vn96 synthetic peptide is indeed a robust and reproducible method for EV isolation that has the potential to be used in a clinical setting. We have therefore chosen to investigate the utility of the Vn96 synthetic peptide for this isolation of EVs from human plasma and the subsequent detection of small RNA biomarkers of PDAC by Next-generation sequencing (NGS) analysis. We find that analysis of small RNA from Vn96-isolated EVs permits the robust discrimination of PDAC patients from non-affected individuals. Moreover, analyses of all small RNA species, miRNAs, and lncRNAs are most effective at segregating PDAC patients from non-affected individuals. We further identified 34 chromosomal regions that encode small RNAs whose differential expression in Vn96-isolated EVs strongly distinguishes PDAC patients from non-affected individuals. Several of the identified small RNA biomarkers have been previously associated with and/or characterized in PDAC, indicating the validity of our findings, whereas other identified small RNA biomarkers may have novel roles in PDAC or cancer in general. Overall, our results provide a basis for a clinically-amendable detection and/or screening strategy for PDAC using a liquid biopsy approach that relies on Vn96-mediated isolation of EVs from plasma.

overall_design: Small RNA sequencing was performed on extracellular vesicle RNA derived from plasma of PDAC patients (n = 16) and non-affected individuals (n = 13).

first: Jeremy

middle: W

last: Roy

first: Gabriel

last: Wajnberg

first: Alexie

last: Ouellette

first: Julie

last: Boucher

first: Jacynthe

last: Lacroix

first: Simi

last: Chacko

first: Anirban

first: Rodney

first: Stephen

last: Lewis


title: Nuclear translocation of an aminoacyl-tRNA synthetase may mediate a chronic "integrated stress response"

city: La Jolla

first: Xiang-Lei

last: Yang

address: 10550 North Torrey Pines Road BCC-110

department: Molecular Medicine

institute: Scripps Research

summary: Various stress conditions are signaled through phosphorylation of translation initiation factor eukaryotic initiation factor 2α (eIF2α) to inhibit global translation while selectively activating transcription factor ATF4 to aid cell survival and recovery. However, this integrated stress response is acute and cannot resolve lasting stress. Here, we report that tyrosyl-tRNA synthetase (TyrRS), a member of the aminoacyl-tRNA synthetase family that responds to diverse stress conditions through cytosol-nucleus translocation to activate stress-response genes, also inhibits global translation. However, it occurs at a later stage than eIF2α/ATF4 and mammalian target of rapamycin (mTOR) responses. Excluding TyrRS from the nucleus over-activates translation and increases apoptosis in cells under prolonged oxidative stress. Nuclear TyrRS transcriptionally represses translation genes by recruiting TRIM28 and/or NuRD complex. We propose that TyrRS, possibly along with other family members, can sense a variety of stress signals through intrinsic properties of this enzyme and strategically located nuclear localization signal and integrate them by nucleus translocation to effect protective responses against chronic stress.

overall_design: Tyrosyl-tRNA synthetase DNA binding sites (TF-ChIP)

last: Wei

first: Xianglei

first: Haissi

last: Cui


title: mousemodel_Heptablastoma spatial transcriptomics

first: Xiang

last: Chen

department: Computational Biology

summary: Lack of relevant animal models and cell lines hampers the understanding of hepatoblastoma and identification of therapeutic targets. We report a  hepatoblastoma murine model that faithfully recapitulates the pathological features of mixed fetal and embryonic hepatoblastoma, with transcriptomics resembling the high-risk gene signatures of human disease

overall_design: Flash Frozen Tumor was harvested  from Hepatocellular model. Spatial Tx data was generated using  Visium Spatial Gene expression kit from 10X genomics. The H&E images were acuqired using Zeiss- AxioScan slide scanner.

first: Jie

last: Fang

first: Shivendra

last: Singh

first: Sivaraman

last: Natarajan

first: Jun


title: KDM6A Epigenetically Regulates Subtype Plasticity in Small Cell Lung Cancer (atac batch2)

city: Boston

first: Yingtian

last: Xie

address: 450 BROOKLINE AVE

institute: DANA FARBER CANCER INSTITUTE INC

summary: Small cell lung cancer (SCLC) exists broadly in four molecular subtypes: ASCL1, NEUROD1,POU2F3, and Inflammatory. Initially SCLC subtypes were thought to be mutually exclusive, butrecent evidence shows intra-tumoral subtype heterogeneity and plasticity between subtypes.Using a CRISPR-based autochthonous SCLC GEMM to study the consequences of KDM6A/UTXinactivation, we found that KDM6A inactivation induced plasticity from ASCL1 to NEUROD1resulting in SCLC tumors with open chromatin at the NEUROD1 promoter that express bothASCL1 and NEUROD1. Mechanistically, KDM6A binds and maintains ASCL1 target genes in anactive chromatin state with its loss increasing H3K27me3 near their promoters leading to a cell state that is primed for ASCL1 to NEUROD1 subtype switching. This work identifies KDM6A as \an epigenetic regulator that controls ASCL1 to NEUROD1 subtype plasticity and provides a novel autochthonous SCLC GEMM to model ASCL1 and NEUROD1 subtype heterogeneity and plasticity, which is found in 35-40% of human SCLCs.

overall_design: Assay for Transposase-Accessible Chromatin using sequencing (ATAC-seq) in WT and KDM6A KO sclc cells

first: Matthew

middle: G

last: Oser


title: Extension of mRNA poly(A) tails and 3′UTRs during neuronal differentiation exhibits variable association with post-transcriptional dynamics

city: Callaghan

first: Dylan

last: Kiltschewskij

state: NSW

address: Room MS616, Level 6, Medical Sciences Building, University Drive

department: School of Biomedical Sciences and Pharmacy

country: Australia

institute: The University of Newcastle

summary: Differentiation of neural progenitor cells into mature neuronal phenotypes relies on extensive temporospatial coordination of mRNA expression to support the development of functional brain circuitry. Cleavage and polyadenylation of mRNA has tremendous regulatory capacity through the alteration of mRNA stability and modulation of microRNA (miRNA) function, however the extent of utilization in neuronal development is currently unclear. Here, we employed poly(A) tail sequencing, mRNA sequencing, ribosome profiling and small RNA sequencing to explore the functional relationship between mRNA abundance, translation, poly(A) tail length, alternative polyadenylation (APA) and miRNA expression in an in vitro model of neuronal differentiation. Differential analysis revealed a strong bias towards poly(A) tail and 3´UTR lengthening during differentiation, both of which were associated with changes in mRNA abundance but not translation. Globally, changes in miRNA expression were predominantly associated with mRNA abundance and translation, however several miRNA-mRNA pairings with potential to regulate poly(A) tail length were identified. Furthermore, 3´UTR lengthening was observed to significantly increase the inclusion of non-conserved miRNA binding sites, potentially enhancing the regulatory capacity of these molecules in mature neuronal cells. Together, our findings suggest poly(A) tail length and APA function as part of a rich post-transcriptional regulatory matrix during neuronal differentiation.

overall_design: Examination of changes in mRNA expression, translation, poly(A) tail length, alternative polyadenylation and miRNA expression during neuronal differentiation of SH-SY5Y cells.

first: Paul

last: Harrison

first: Chantel

last: Fitzsimmons

first: Traude

last: Beilharz

first: Murray

last: Cairns


title: Impact of Plasmodium falciparum Infection on DNA Methylation of Circulating Immune Cells

city: Abu dhabi

first: Youssef

last: Idaghdour

state: Abu dhabi

address: Saadiyat Marina District

department: Biology

country: United Arab Emirates

institute: NYUAD

summary: The regulation of immune cell responses to infection is a complex process that involves various molecular mechanisms, including epigenetic regulation. DNA methylation has been shown to play central roles in regulating gene expression and modulating cell response during infection. However, the nature and extent to which DNA methylation is involved in the host immune response in human malaria remains unknown. Here, we present a longitudinal study investigating the temporal dynamics of genome-wide in vivo DNA methylation profiles using 189 MethylationEPIC 850K profiles from 66 children in Burkina Faso, West Africa, sampled three times: before infection, during symptomatic parasitemia, and after malaria treatment. The results revealed major changes in the DNA methylation profiles of children in response to both Plasmodium falciparum infection and malaria treatment, with widespread hypomethylation of CpGs upon infection (82% of 6.8K differentially methylated regions). We document a remarkable reversal of CpG methylation profiles upon treatment to pre-infection states. These changes implicate divergence in core immune processes, including the regulation of lymphocyte, neutrophil, and myeloid leukocyte function. Integrative DNA methylation-mRNA analysis of a top differentially methylated region overlapping the pro-inflammatory gene TNF implicates DNA methylation of TNF cis regulatory elements in the molecular mechanisms of TNF regulation in human malaria. Our results highlight a central role of epigenetic regulation in mounting the host immune response to Plasmodium falciparum infection and in response to malaria treatment.

overall_design: 189 samples from 66 children in Burkina Faso of which 57 are fully matched sampled collected at three time points: before infection, during symptomatic parasitemia, and after malaria treatment.

first: Dareen

last: Almojil

first: Aïssatou

last: Diawara

first: Issiaka

last: Soulama

first: Mame

last: Massar Dieng

first: Vinu

last: Manikandan

first: Samuel

last: Sermé

first: Salif

last: Sombié

first: Amidou

last: Diarra

first: Aissata

last: Barry

first: Sam

last: Aboubacar Coulibaly

first: Sodiomon

last: Sirima


title: KDM6A Epigenetically Regulates Subtype Plasticity in Small Cell Lung Cancer (atac batch3)

first: Yingtian

last: Xie

first: Matthew

last: Oser


title: The mRNA methyltransferase Mettl3 modulates cytokine mRNA stability and limits functional responses in mast cells [RNA-seq]

first: Cristina

last: Leoni

overall_design: Bone marrow derived mast cells (BMMCs) were in vitro differentiated from 4 mice and transfected with Mettl3 RNPs or mock transfected (pulsed). Total RNA was extracted from each sample and used for mRNA sequencing

first: Marian

last: Bataclan

first: Silvia

last: Monticelli


title: The mRNA methyltransferase Mettl3 modulates cytokine mRNA stability and limits functional responses in mast cells [NanoString]

overall_design: 3 sets of in vitro bone marrow derived mast cells were CRISPR/Cas9 modified to knockout Mettl3 or cKit, as  a control, and insert a zsGreen reporter gene at the knockout target site. Cells were sorted for high zsGreen expression and were either left resting or were stimulated for 4h with IgE and antigen complexes.

last: Abigaile


title: KDM6A Epigenetically Regulates Subtype Plasticity in Small Cell Lung Cancer (atac batch1)

summary: Small cell lung cancer (SCLC) exists broadly in four molecular subtypes: ASCL1, NEUROD1,POU2F3, and Inflammatory. Initially SCLC subtypes were thought to be mutually exclusive, butrecent evidence shows intra-tumoral subtype heterogeneity and plasticity between subtypes.Using a CRISPR-based autochthonous SCLC GEMM to study the consequences of KDM6A/UTXinactivation, we found that KDM6A inactivation induced plasticity from ASCL1 to NEUROD1resulting in SCLC tumors with open chromatin at the NEUROD1 promoter that express bothASCL1 and NEUROD1. Mechanistically, KDM6A binds and maintains ASCL1 target genes in anactive chromatin state with its loss increasing H3K27me3 near their promoters leading to a cell state that is primed for ASCL1 to NEUROD1 subtype switching. This work identifies KDM6A as an epigenetic regulator that controls ASCL1 to NEUROD1 subtype plasticity and provides a novel autochthonous SCLC GEMM to model ASCL1 and NEUROD1 subtype heterogeneity and plasticity, which is found in 35-40% of human SCLCs.

overall_design: Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for control and KDM6A KO in sclc cells


title: Promising AAV.U7snRNAs vectors targeting DMPK improve DM1 hallmarks in patient-derived cell lines

city: Columbus

first: Camila

middle: F

last: Almeida

state: OH

address: 700 Children's Drive, RB2 WA3103

department: Center for Gene therapy

institute: Nationwide Children's Hospital

summary: To evaluate the effect of AAV.U7snRNAs in the gene expression and alternative expression profiles of myotonic dystrophy cell lines.

overall_design: Comparative RNAseq analysis of cell lines before and after AAV treatment

first: Robert

last: Weiss

first: Nicolas

last: Wein


title: Activation of the Aryl Hydrocarbon Receptor in Muscle Exacerbates Ischemic Pathology in Chronic Kidney Disease

city: Gainesville

first: Terence

middle: E.

last: Ryan

state: Florida

address: 1864 Stadium Rd

department: Applied Physiology and Kinesiology

institute: University of Florida

summary: This study examines the transcriptional profile at a single-nuclei resolution in mice with muscle-specific deletion of the aryl hydrocarbon receptor (AHR). Wildtype (AHRfl/fl, floxed Cre negative littermates) and AHRmKO (AHR muscle specific knockout) mice were given chronic kidney disease via adenine supplemented diet. Mice were then subjected to femoral artery ligation as an experimental model of peripheral artery disease. Tibialis anterior muscles were harvested at five days post-surgery and immediately snap frozen in liquid nitrogen and stored at -80C until nuclei isolation. In both libraries, muscle specimens were pooled from three males prior to nuclei isolations were performed.

overall_design: Nuclei were isolated from tibialis anterior muscle of male AHRfl/fl and AHRmKO mice with chronic kidney disease five days after femoral artery ligation surgery and analyzed using snRNAseq.

first: Nicholas

last: Balestrieri

first: Caroline

last: Pass


title: Lactate-dependent Transcriptional Regulation Controls Mammalian Eye Morphogenesis [H3K27ac ChIP-seq]

city: Chicago

first: NOZOMU

last: TAKATA

state: Illinois

address: 303 East Superior Street, 10-220

department: Center for Vascular and Developmental Biology

institute: Northwestern University Feinberg School of Medicine

summary: Mammalian retinal metabolism favors aerobic glycolysis. However, the role of glycolytic metabolism in retinal morphogenesis remains unknown. We report that aerobic glycolysis is necessary for the early stages of retinal development. Taking advantage of an unbiased approach that combines the use of eye organoids and single-cell RNA sequencing, we identify specific glucose transporters and glycolytic genes in retinal progenitors. Next, we determine that the optic vesicle territory of mouse embryos displays elevated levels of glycolytic activity. At the functional level, we show that removal of Glucose transporter 1 and Lactate dehydrogenase A gene activity from developing retinal progenitors arrests eye morphogenesis. Surprisingly, we uncover that lactate-mediated upregulation of key eye-field transcription factors is controlled by the epigenetic modification of histone H3 acetylation through histone deacetylase activity. Our results identify an unexpected bioenergetic independent role of lactate as a signaling molecule necessary for mammalian eye morphogenesis.

overall_design: Comparative histone h3 k27 acetylation (H3K27ac) profiling analysis of ChIP-seq data for the eye organoids and its treatments (control, GNE-140, and GNE-140 with lactate).

first: Nozomu

last: Takata

first: Marc

middle: A

last: Morgan

first: Zack

last: Rosenstadt

first: Ryan

last: Embry

last: Schipma


title: RNA-seq analysis of sorted duodenum epithelial cells and duodenum lamia propria macrophages from mice where Tsc2 is deleted in Lyz2-expressing cells

city: Vienna

first: Thomas

last: Weichhart

address: Waehringerstraße 10

department: Center for Pathobiochemistry and Genetics

country: Austria

institute: Medical University of Vienna

summary: Iron is an essential cellular and important for many physiological functions including erythropoiesis and host defense. It is taken up from the diet in the duodenum, the first part of the small intestine by enterocytes and loaded onto transferrin, the main iron transport molecules in the circulation. Macrophages are ancient sentinel cells that are implicated in all steps of systemic iron metabolism except duodenal iron import. By assessing mice that harbor a macrophage-specific deletion of the tuberous sclerosis complex 2 (Tsc2), a negative regulator of mTORC1, we found that these mice possessed various defects in iron metabolism including defective steady state erythropoiesis and a highly reduced saturation of transferrin with iron. This iron-deficiency phenotype was caused by a block of iron import from the duodenal epithelial cells into the circulation. Activation of mTORC1 in villous duodenal CD68+ macrophages induced expression of serine proteases and promoted local degradation of the major iron transport molecule transferrin, whereas depletion of macrophages in mice increased transferrin levels in the duodenal villi. Inhibition of serine protease activity restored transferrin levels in the Tsc2-deficient mice. Physiologically, transferrin levels were regulated in the villi during Citrobacter rodentium infection in mice. These data show that transferrin is controlled locally in the duodenum and establish macrophages as regulators of duodenal iron uptake.

overall_design: Duodenum lamina propria macrophages (Cd45+, Ly6G-, SiglecF-, Cd24-, Cd11c+/-, Cd103-, Cd11b+, Ly6C-, Cd64+, MHCII+) and EpCAM+ epithelial cells (Cd45-, EpCAM+) from 3 Tsc2fl/fl and 4 Tsc2Lyz2-cre mice were isolated as per manufacturer’s instructions (MACS Miltenyi Biotec, 130-097-410) and FACS sorted at the FACS Core Unit of the Children’s Cancer Research Institute (CCRI) of Vienna. Freshly sorted cells were harvested in RLT-Buffer (RNeasy RNA isolation kits, QIAGEN) containing ß-mercaptoethanol (Sigma) and stored at -80 °C. RNA was isolated using an RNeasy Micro Kit (QIAGEN, Cat No.ID: 74004).

first: Nyamdelger

last: Sukhbaatar

first: Stephanie

last: Fritsch

first: Michael

last: Schuster


title: scRNA-seq analysis of duodenum CD68+ macrophages from mice where Tsc2 is deleted in Lyz2-expressing cells

summary: Iron is an essential cellular trace element and important for many physiological functions including erythropoiesis and host defense. It is taken up from the diet in the duodenum, the first part of the small intestine by enterocytes and loaded onto transferrin, the main iron transport molecules in the circulation. Macrophages are ancient sentinel cells that are implicated in all steps of systemic iron metabolism except duodenal iron import. By assessing mice that harbor a macrophage-specific deletion of the tuberous sclerosis complex 2 (Tsc2), a negative regulator of mTORC1, we found that these mice possessed various defects in iron metabolism including defective steady state erythropoiesis and a highly reduced saturation of transferrin with iron. This iron-deficiency phenotype was caused by a block of iron import from the duodenal epithelial cells into the circulation. Activation of mTORC1 in villous duodenal CD68+ macrophages induced expression of serine proteases and promoted local degradation of the major iron transport molecule transferrin, whereas depletion of macrophages in mice increased transferrin levels in the duodenal villi. Inhibition of serine protease activity restored transferrin levels in the Tsc2-deficient mice. Physiologically, transferrin levels were regulated in the villi during Citrobacter rodentium infection in mice. These data show that transferrin is controlled locally in the duodenum and establish macrophages as regulators of duodenal iron uptake.

overall_design: Duodenum lamina propria CD45+ immune cells with CD68-tag macrophages (Cd45+,Cd68+)


title: KDM6A Epigenetically Regulates Subtype Plasticity in Small Cell Lung Cancer

last: Tolstorukov

state: MA

address: 360 Longwood Avenue

department: Informatics

institute: Dana Farber Cancer Institute

summary: Using a CRISPR-based autochthonous SCLC GEMM to study the consequences of KDM6A/UTX inactivation, we found that KDM6A inactivation induced plasticity from ASCL1 to NEUROD1 resulting in SCLC tumors with open chromatin at the NEUROD1 promoter that express both ASCL1 and NEUROD1. Mechanistically, KDM6A binds and maintains ASCL1 target genes in an active chromatin state with its loss increasing H3K27me3 near their promoters leading to a cell state that is primed for ASCL1 to NEUROD1 subtype switching. This work identifies KDM6A as an epigenetic regulator that controls ASCL1 to NEUROD1 subtype plasticity and provides a novel autochthonous SCLC GEMM to model ASCL1 and NEUROD1 subtype heterogeneity and plasticity, which is found in 35-40% of human SCLCs.

overall_design: Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for histone marks H3K4me2 and H3K27me3 and the histone demethylase KDM6A in primary mouse small cell lung cancer cells (SCLC) derived from SCLC genetically-engineered mouse models that are KDM6A WT (sgControl RPP) or KDM6A-Mutant (sgKdm6a RPP).

first: Leslie

last: Duplaquet

first: Matthew

last: Oser


title: KDM6A Epigenetically Regulates Subtype Plasticity in Small Cell Lung Cancer.

first: Yingtian

last: Xie

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

overall_design: Refer to individual Series


title: KDM6A Epigenetically Regulates Subtype Plasticity in Small Cell Lung Cancer (scatac)

overall_design: single cell ATAC sequencing for WT and KDM6A KO sclc cells


title: Germ-cell specific eIF4E1b regulates maternal mRNA translation to ensure zygotic genome activation [RIP-seq]

first: Guanghui

last: Yang

address: 50 South Drive

institute: National Institutes of Health

summary: Translation of maternal mRNAs is detected before transcription of zygotic genes and is essential for mammalian embryo development. How certain maternal mRNAs are selected for translation instead of degradation and how this burst of translation affects zygotic genome activation remains unknown. Using gene-edited mice, we document that the oocyte-specific eukaryotic translation initiation factor 4E family member 1b (eIF4E1b) is the regulator of maternal mRNA expression that ensures subsequent reprogramming of the zygotic genome. In oocytes, eIF4E1b binds to transcripts encoding translation machinery proteins, chromatin remodelers and reprogramming factors to promote their translation in zygotes and protect them from degradation. The protein products are thought to establish an open chromatin landscape in one-cell zygotes to enable transcription of genes required for cleavage stage development. Our results define a program for rapid resetting of the zygotic epigenome that is regulated by maternal mRNA expression and provides new insights into the mammalian maternal-to-zygotic transition.

overall_design: RNA profiles after IP with anti-HA beads using M2 eggs or early 2-cell embryos collected from Eif4e1b FLAGHA knock-in(KI) females or WT controls.

first: Qiliang

last: Xin

first: Jurrien

last: Dean


title: Loss of LCMT1 and biased protein phosphatase 2A heterotrimerization drive prostate cancer progression and therapy resistance

city: Philadelphia

first: Irfan

last: Asangani

state: PA

address: 421 Curie Blvd

department: Cancer Biology

institute: University of Pennsylvania

summary: The first line of therapy for advanced prostate cancer (PCa) is androgen-deprivation therapy (ADT) through surgical or chemical castration; however, in the majority of cases, tumors relapse in a hormone refractory or castration resistant prostate cancer (CRPC) form. Once the PCa has recurred in CRPC form, it progresses to a highly aggressive disease with frequent metastasis and poses an increased risk of morbidity and death. This study shows that the loss of PP2AcÎ± methylation in enzalutamide (Enza)-resistant CRPC cells plays a central role in imparting the resistant to the cancer cells by stabilizing the interaction of MED1, BRD4, and AR associated transcriptional complex, thereby amplifying the oncogenic AR transcriptional output through chromatin re-modulatory mechanism. Further, this study demonstrates that targeting the PP2ACÎ± regulatory mechanisms or its downstream epigenetic effectors/mechanisms abolish the enzalutamide-resistance phenotype, thus paving the way for the development of more effective therapeutics to curtail mCRPC. The below given experiments  validate the above findings.

overall_design: LNCaP enzalutamide-resistant (LNCaP-MDVR) cell line was generated by culturing the parental LNCaP cells in presence of enzalutamide for 4-6 months. The comparison of the two cell lines in terms of MED1 and AR chromatin recruitment was performed upon DMSO or ARD-69 or DT-061 treatment. LNCaP cells stably over-expressing LacZ and leucine carboxyl methyltransferase 1 (LCMT1) were prepared and used to study the differential chromatin binding of MED1 and AR.

first: Reyaz

middle: U

last: Rasool

first: Mohammed

last: Alhusayan


title: Lactate-dependent Transcriptional Regulation Controls Mammalian Eye Morphogenesis [scRNA-seq]

first: NOZOMU

last: TAKATA

overall_design: Comparative gene expression profiling analysis of single cell RNA-seq data for the eye organoids at two stages (day 4 and day 6 in culture).

first: Nozomu

last: Takata

first: Luciano

last: Fiore

first: Nikita

last: Joshi

first: Alexander

middle: V

last: Misharin


title: Multimodal sequencing (gene expression, chromatin accessibility, and mtDNA genotyping) of single cells of the RPE and choroid in human MELAS (m.3243A>G) and control samples

city: Iowa City

first: Nathaniel

middle: Kevin

last: Mullin

state: IA

address: 375 Newton Road

institute: University of Iowa

summary: The mitochondrial m.3243A>G variant is known to cause retinal dystrophy and vision loss. We used single cell multimodal sequencing to understand how the presence of this mutation affects cellular phenotype in a cell type-specific manner.

overall_design: Human retinas were dissected to isolate the RPE/choroid (RPE/choroid) or RPE was mechanically dissected away from Bruch's membrane (purified RPE). RPE/choroid samples were then dissociated to single cells using collagenase. Samples from a MELAS patient and age/sex-matched healthy control donor were prepared.

first: Mullin

middle: K

last: Nathaniel

first: Tucker

last: Budd

first: Mullins

last: Robert


title: Lactate-dependent Transcriptional Regulation Controls Mammalian Eye Morphogenesis [RNA-seq]

overall_design: Comparative gene expression profiling analysis of RNA-seq data for the eye organoids and its treatments (control, GNE-140, and GNE-140 with lactate).

first: Ryan

last: Embry

first: Matthew

last: Schipma


title: Gene expression from single cells of the retina and choroid in human MELAS (m.3243A>G) and control samples [scRNA-seq]

summary: The mitochondrial m.3243A>G variant is known to cause retinal dystrophy and vision loss. We used single cell RNA sequencing to understand how the presence of this mutation affects cellular phenotype in a cell type-specific manner.

overall_design: Human retinas were dissected into neural retina and RPE/choroid from two regions (macular and peripheral). Samples were then dissociated to single cells using papain or collagenase respectively. Samples from a MELAS patient and age/sex-matched healthy control donor were prepared.

first: Budd

last: Tucker

first: Robert

last: Mullins


title: A hybrid nitrogenase with regulatory elasticity in Azotobacter vinelandii

status: Public on Jun 07 2023

city: Madison

first: Betül

last: Kaçar

state: WI

address: 1550 Linden Dr.

department: Department of Bacteriology

institute: University of Wisconsin-Madison

summary:
Biological nitrogen fixation, the microbial reduction of atmospheric nitrogen to bioavailable ammonia, represents both a major limitation on biological productivity and a highly desirable engineering target for synthetic biology. However, the engineering of nitrogen fixation requires an integrated understanding of how the gene regulatory dynamics of host diazotrophs respond across sequence-function space of its central catalytic metalloenzyme, nitrogenase. Here, we interrogate this relationship by analyzing the transcriptome of Azotobacter vinelandii engineered with a phylogenetically inferred ancestral nitrogenase protein variant. The engineered strain exhibits reduced cellular nitrogenase activity but recovers wild-type growth rates following an extended lag period. We find that expression of genes within the immediate nitrogen fixation network is resilient to the introduced nitrogenase sequence-level perturbations. Rather the sustained physiological compatibility with the ancestral nitrogenase variant is accompanied by reduced expression of genes that support trace metal and electron resource allocation to nitrogenase. Our results spotlight gene expression changes in cellular processes adjacent to nitrogen fixation as productive engineering considerations to improve compatibility between remodeled nitrogenase proteins and engineered host diazotrophs.

IMPORTANCE
Azotobacter vinelandii is a key model bacterium for the study of biological nitrogen fixation, an important metabolic process catalyzed by nitrogenase enzymes. Here, we demonstrate that compatibilities between engineered A. vinelandii strains and nitrogenase variants can be modulated at the regulatory level. The engineered strain studied here responds by adjusting the expression of proteins involved in cellular processes adjacent to nitrogen fixation, rather than that of nitrogenase proteins themselves. These insights can inform future strategies to transfer nitrogenase variants to non-native hosts.

overall_design: Comparative RNA-seq gene expression analysis for Azotobacter vinelandii WT and  strain ancNif (ΔnifD) grown diaztrophically. Three biological replicates were sequenced and analyzed for each strain.

first: Alex

last: Rivier

first: Kevin

middle: S

last: Myers

first: Amanda

last: Garcia

first: Morgan

last: Sobol

first: Betul

last: Kacar


first: Yixiang

last: Li

address: 450 Brookline Ave

institute: Dana-Farber Cancer Institute

summary: Using a CRISPR-based autochthonous SCLC GEMM to study the consequences of KDM6A/UTX inactivation, we found that KDM6A inactivation induced plasticity from ASCL1 to NEUROD1 resulting in SCLC tumors with open chromatin at the NEUROD1 promoter that express both ASCL1 and NEUROD1. Mechanistically, KDM6A binds and maintains ASCL1 target genes in an active chromatin state with its loss increasing H3K27me3 near their promoters leading to a cell state that is primed for ASCL1 to NEUROD1 subtype switching. This work identifies KDM6A as an epigenetic regulator that controls ASCL1 to NEUROD1 subtype plasticity and provides a novel autochthonous SCLC GEMM to model ASCL1 and NEUROD1 subtype heterogeneity and plasticity, which is found in 35-40% of human SCLCs

overall_design: Single cell RNA sequencing in primary mouse small cell lung cancer cells (SCLC) derived from SCLC genetically-engineered mouse models that are KDM6A WT (sgControl RPP) or KDM6A-Mutant (sgKdm6a RPP).

last: Oser


title: Visium Spatial Transcriptomics of a murine model of IPMN

first: Anirban

last: Maitra

state: NY

address: Smilow 910, 522 1st Ave

department: Perlmutter Cancer Center

institute: New York University Grossman School of Medicine

summary: Pancreata were collected from two Kras;Gnas mice and analyzed by Visium Spatial Gene Expression to evaluate the transcriptional profile associated with NKX6-2 that was observed in the human IPMN samples.

overall_design: Samples were collected from mice at 15 and 25 weeks post doxycycline treatment and were flash frozen. The samples were then cryosectioned and analyzed with the Visium Spatial Gene Expression platform from 10x genomics.

first: Marta

last: Sans

first: Kimal

last: Rajapakshe

first: Yuki

last: Makino

first: Jimin

last: Min

first: Paola

last: Guerrero


title: A gut commensal protist protects against virus-mediated loss of oral tolerance_2

city: PITTSBURGH

first: Reinhard

last: Hinterleitner

address: 5051 Centre Avenue

department: Immunology

institute: University of Pittsburgh

summary:
To better understand how Tritrichomonas arnold colonization impacts the reovirus-mediated proinflammatory response in mesenteric lymph nodes, we examined the transcriptional profile of mesenteric lymph nodes
For RNA-sequencing single cell suspension of mesenteric lymph nodes  were lysed in RLT buffer (Qiagen) and RNA was isolated

overall_design: Wild-type mice were inoculated perorally with 10^6 Tritrichomonas arnold or PBS control for 12 days followed by peroral inoculation with 10^9 PFU of reovirus strain T1L or PBS control for 48 hours. Bulk RNA-sequencing  mesenteric lymph nodes  (PBS, n =4), (T1L, n = 4), (T1L+T. arnold, n =4), (T. arnold, n = 4).

first: Das

last: Jishnu

first: Mehta

middle: D

last: Isha


title: A gut commensal protist protects against virus-mediated loss of oral tolerance

summary:
To better understand how Tritrichomonas arnold colonization impacts the reovirus-mediated proinflammatory response in dietary antigen-presenting  dendtric cells (cDC1), we examined the transcriptional profile of mesenteric lymph node-derived cDC1.
For RNA-sequencing on CD103+ CD11b- CD8a+ DCs, 1,000 CD103+ CD11b- CD8a+ DCs were FACS sorted into a 96-well plate

overall_design: Wild-type mice were inoculated perorally with 10^6 Tritrichomonas arnold or PBS control for 12 days followed by peroral inoculation with 10^9 PFU of reovirus strain T1L or PBS control for 48 hours. Smart-seq RNA-sequencing of MHC-II+ CD11c+ CD103+ CD11b- CD8a+ mLN DCs (PBS, n =4), (T1L, n = 3), (T1L+T. arnold, n =4), (T. arnold, n = 3).


title: Germ-cell specific eIF4E1b regulates maternal mRNA translation to ensure zygotic genome activation [NOMe-seq]

city: Bethesda

first: Guanghui

last: Yang

department: National Institute of Diabetes and Digestive and Kidney Diseases

overall_design: NOMe-seq of single Eif4e1b+/- or Eif4e1b-/- embryos collected at different developmental stages for joint profiling of chromatin accessibility and DNA methylation.

first: Qiliang

last: Xin

first: Jurrien

last: Dean