title: Mucosal immune responses to live attenuated influenza vaccine in young adults

city: London

first: Ryan

last: Thwaites

state: London

address: St Mary's Hospital

institute: Imperial College London

summary: Compared to intramuscular vaccines, nasally administered vaccines have the advantage of inducing local mucosal immune responses that may block infection and interrupt transmission of respiratory pathogens1. Live attenuated influenza vaccine (LAIV) is commonly used in children2, but its effectiveness declines with age3. This may be attributed to the gradual accumulation of homo- or hetero-subtypic immunity that blocks vaccine replication necessary to induce protective responses3, 4. Despite its demonstrable efficacy against influenza in children, correlates of protection for LAIV remain elusive5. Studying young adult volunteers we found that LAIV induced distinct, compartmentalized, antibody responses in the mucosa and blood. LAIV also induced mucosal IL-33 release in the first 8 hours post-inoculation and distinct CD8+ and cTfh T cell activation profiles. Mucosal antibodies are induced separately from blood antibodies and may provide a simple and novel correlate of protection for mucosal vaccination.

overall_design: 40 adult volunteers were inoculted with live attenuated influenza vaccine, and nasal inferior turbinate samples collected by nasal curettage using RhinoPro curettes at baseline (up to 7 days prior to inoculation) and 72 hours post inoculation.


title: Expression of the CIC-DUX4 fusion oncoprotein mirrors human CIC-rearranged sarcoma in a transgenic mouse model (ChIP-Seq)

status: Public on Oct 19 2023

city: Durham

first: Peter

last: Hendrickson

state: North Carolina

address: 308 Research Drive

country: USA

institute: Duke University

type: Genome binding/occupancy profiling by high throughput sequencing

summary: CIC-DUX4 sarcoma (CDS) is a rare but highly aggressive undifferentiated small round cell sarcoma driven by a fusion between the tumor suppressor Capicua (CIC) and DUX4. Currently, there are no effective treatments and efforts to identify and translate better therapies is limited by the scarcity of tissues and patients. To minimize, we made three genetically engineered mouse models of CDS (Ch7CDS, Ai9CDS, and TOPCDS). Remarkably, chimeric mice from all three models developed spontaneous tumors and widespread metastasis in the absence of Cre-recombinase. The penetrance of spontaneous (Cre-independent) tumor formation was complete irrespective of bi-allelic CIC function and loxP site proximity. Characterization of primary and metastatic mouse tumors showed that they consistently expressed the CIC-DUX4 fusion protein as well as other downstream markers of the disease authenticating them as CDS. Lastly, tumor-derived cell lines were generated and ChIP-seq was preformed to map fusion-gene specific binding using an N-terminal HA epitope tag. These datasets, along with paired H3K27ac ChIP-seq maps, validate CIC-DUX4 as a neomorphic transcriptional activator and point to ETS family transcription factors as cooperative and redundant drivers of the core regulatory circuitry.

overall_design: To authenticate the tumors and tumor-derived cell lines obtained from our genetically-engineered mouse models (Ch7CDS, Ai9CDS, TOPCDS), RNA-sequencing and ChIP-sequencing was performed. Comparative gene expression profiling analysis was carried out using KP (KrasG12D, p53fl/fl) tumors/cells and normal hindlimb skeletal muscle as controls.

middle: G

first: Kristianne

middle: M

last: Oristian

first: MaKenna

middle: R

last: Browne

first: Lixia

last: Lou

first: Yan

last: Ma

first: Dianna

last: Cardona

first: Corinne

last: Linardic

first: David

last: Kirsch


title: IL-27 produced during acute malaria infection regulates Plasmodium-specific memory CD4+ T cells

city: Nagasaki

first: Katsuyuki

last: Yui

state: Nagasaki

address: 長崎市坂本１丁目12番4号

country: Japan

institute: Nagasaki University

type: Expression profiling by high throughput sequencing

summary: Malaria infection elicits both protective and pathogenic immune responses, and IL-27 is a critical cytokine that regulate effector responses during infection. Here, we identified a critical window of CD4+ T cell responses that is targeted by IL-27. Neutralization of IL-27 during acute infection with Plasmodium chabaudi expanded specific CD4+ T cells, which were maintained at high levels thereafter. In the chronic phase, Plasmodium-specific CD4+ T cells in IL-27-neutralized mice consisted mainly of CD127+KLRG1- and CD127-KLRG1+ subpopulations that displayed distinct cytokine production, proliferative capacity and are maintained in a manner independent of active infection. Single cell RNA-seq analysis revealed that these CD4+ T cell subsets formed independent clusters that express unique Th1-type genes. These IL-27-neutralized mice exhibited enhanced cellular and humoral immune responses and protection. These findings demonstrate that IL-27, which is produced during the acute phase of malaria infection, inhibits the development of unique Th1 memory precursor CD4+ T cells, suggesting potential implications for the development of vaccines and other strategic interventions.

overall_design: scRNA-seq and CITE-Seq analysis were performed on splenic Plasmodium-specific TCR transgenic CD4+ T cells from Plasmodium chabaudi chabaudi (Pcc)-infected C57BL/6 mice that were treated with either anti-IL-27 mAb or IgG control. Single cell data from IgG- and  anti-IL-27 mAb-treated mice were collected and analyzed for days 7 and 28 post-infection (pi) time point, as well as day 14 pi timepoint from anti-IL-27 mAb-treated mouse, with 1 replicate each (total of 5 samples). CITE-Seq analysis made use of IgG2a control, CD49d, CD127 and KLRG1 as target markers.

first: Maria-Lourdes

last: Macalinao

first: Shin-Ichi

last: Inoue

first: Sanjaadorj

last: Tsogtsaikhan

first: Hirotaka

last: Matsumoto

first: Ganchimeg

last: Bayarsaikhan

first: Jiun-Yu

last: Jian

first: Kazumi

last: Kimura

first: Yoshiaki

last: Yasumizu

first: Tsuyoshi

first: Hiroki

last: Yoshida

first: Julius

last: Hafalla

first: Daisuke


title: SMARCB1 loss creates patient-specific MYC topologies that drive malignant rhabdoid tumor growth

city: Utrecht

first: Irene

middle: Evelyn Karla

last: Paassen

address: Heidelberglaan 25

country: Netherlands

institute: Prinses Maxima Centrum

summary: Malignant rhabdoid tumor (MRT) is a highly malignant and often lethal childhood cancer. MRTs are genetically defined by bi-allelic inactivating mutations in SMARCB1, a member of the BRG1/BRM-associated factors (BAF) chromatin remodeling complex. Here, we studied derailed regulatory landscapes as a consequence of SMARCB1 loss in the context of MRT. Our multi-omics approach on patient-derived MRT organoids revealed a dramatic reshaping of the regulatory landscape upon SMARCB1 reconstitution. Chromosome conformation capture experiments subsequently revealed patient-specific looping of distal enhancer regions with the promoter of the MYC oncogene and validation of this data in patients with combined single-cell RNA-seq and ATAC-seq.

overall_design:
Paired scRNA and scATAC-seq on the same cell of primary malignant rhabdoid tumor samples (viably frozen or snapfrozen)

>>>Submitter states: raw data are unavailable due to patient privacy concerns.<<<

first: Ning

middle: Q

last: Liu

first: Lars

last: Custers

first: Hans

last: Teunissen

first: Dilara

last: Ayyildiz

first: Jiayou

last: He

first: Eelco

last: W Hoving

first: Elzo

last: de Wit

first: Jarno

last: Drost


title: Nonfucosylation of an anti-TIGIT antibody enhances FcyR engagement, driving innate immune activation and antitumor activity

status: Public on Oct 18 2023

city: Bothell

first: Alyson

middle: J

last: Smith

state: WA

address: 21717 30th Drive S.E., Building 3

department: Research

institute: Seagen Inc.

summary: TIGIT is an immune checkpoint receptor expressed on activated and memory T cells, immunosuppressive T regulatory cells, and natural killer (NK) cells. TIGIT has emerged as an attractive target for antitumor therapies, due to its proposed immunosuppressive effects on lymphocyte function and T cell activation. We generated an anti-TIGIT monoclonal antibody (mAb) that binds with high affinity to human, non-human primate, and murine TIGIT and through multiple experimental methodologies demonstrated that checkpoint blockade alone is insufficient for antitumor activity. Generating anti-TIGIT mAbs with various Fc backbones we show that muting the Fc-Fcγ receptor (FcγR) interaction failed to drive antitumor activity, while mAbs with Fc functional backbones demonstrate substantial antitumor activity, mediated through activation of antigen-presenting cells (APCs), T cell priming, and NK-mediated depletion of suppressive Tregs and exhausted T cells. Further, nonfucosylation of the Fc backbone resulted in enhanced immune responses and antitumor activity relative to the intact IgG1 backbone. The improved activity correlated with the biased FcγR interaction profile of the nonfucosylated anti-TIGIT mAb, which supports that FcγRIIIa binding with decreased FcγRIIb binding favorably activates APCs and enhances tumor-specific CD8+ T cell responses. The anti-TIGIT mAbs with intact FcγR interacting backbones also demonstrated synergistic enhancement of other standard antitumor treatments, including anti-PD-1 treatment and a model monomethyl auristatin E antibody–drug conjugate. These findings highlight the importance of the anti-TIGIT mAb’s Fc backbone to its antitumor activity and the extent to which this activity can be enhanced through nonfucosylation of the backbone

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Bulk RNA-seq was performed on tumors from syngeneic mouse models, and gene-level transcript levels were subsequently used to compute immune-related gene signature scores that were correlated with the activity of SEA-TGT in those models.

first: Robert

middle: E

last: Thurman

first: Lupe

last: Gutierrez


title: Single-cell transcriptomics illuminates anterior-posterior patterning of the mesoderm in Drosophila embryos at gastrulation

city: Pasadena

first: Angelike

last: Stathopoulos

state: CA

address: 1200 E Calif Blvd

institute: Caltech

summary: Single-cell technology was used to investigate dynamic transcriptional changes in the blastula and gastrula of Drosophila embryos. Cells from embryo of stages 5 to 9 were isolated and subjected to library construction. Mesoderm cells were labeled with mCD8-GFP utilizing the GAL/UAS system. PCA identified 12 unique cell clusters that represent the blastoderm, mesoderm, endoderm and ectoderm anlage, primordia, and a sampling of differentiating cell types. In the cellular blastoderm, we found that the major wave of zygotic gene activation consists of two steps, which is supported by sub-clustering analysis. Trajectory analysis of stage 6-8 mesoderm cells suggested that transcription factors, previously linked to ectodermal segment polarity, also control mesoderm segmentation, patterning and differentiation. With the cells originating from the posterior ventral region of the blastoderm, trajectory analysis identified a group of cells that are multipotent progenitors giving rise to visceral muscles, Malpighian tubules as well as a new cell type. In summary, this single-cell study of gastrulation demonstrates early patterning involves trunk- versus pole-centered programs, the mesoderm is segmentally patterned, and a new cell type arises from the caudal mesoderm.

overall_design: Embryos with mesoderm cells marked with twi(2xPE)>mCD8-GFP were collected for 1.5 hours and aged for 3 hours 15min at 22°C. A single-cell suspension was prepared through manual dissociation and immediately subjected to library construction without fixation.

first: Jingjing

last: Sun

first: Fan

last: Gao


title: Small Molecule Inducers of Neuroprotective miR-132 Identified by HTS-HTS in Human iPSC-derived Neurons

city: Boston

first: Zhiyun

last: Wei

state: Massachusetts

address: 60 Fenwood Road

department: Neurology

institute: Brigham and Women's Hospital

type: Non-coding RNA profiling by high throughput sequencing

summary: MicroRNAs (miRNAs) are short RNAs that regulate fundamental biological processes. miR-132, a key miRNA with established functions in Tau homeostasis and neuroprotection, is consistently downregulated in Alzheimer’s disease (AD) and other tauopathies. miR-132 overexpression rescues neurodegenerative phenotypes in several AD models. To complement research on miRNA-mimicking oligonucleotides targeting the central nervous system, we developed a high-throughput-screen coupled high-throughput-sequencing (HTS-HTS) in human induced pluripotent stem cell (iPSC)-derived neurons to identify small molecule inducers of miR-132. We discovered that cardiac glycosides, which are canonical sodium-potassium ATPase inhibitors, selectively upregulated miR-132 in the sub-μM range. Coordinately, cardiac glycoside treatment downregulated total and phosphorylated Tau in rodent and human neurons and protected against toxicity by glutamate, N-methyl-D-aspartate, rotenone, and Aβ oligomers. In conclusion, we identified small-molecule drugs that upregulated the neuroprotective miR-132 and ameliorated neurodegenerative phenotypes. Our dataset also represents a comprehensive resource for discovering small molecules that modulate specific miRNAs for therapeutic purposes.

overall_design: Human NGN2-driven iPSC-derived neurons (NGN2-iNs) were cultured in 96-well plates. On day 19, the Selleckchem library (N=1,902 compounds), a diverse library of bioactive molecules, was pin-transferred into plates to achieve 10 μM final concentration. DMSO (0.1% final concentration) and forskolin (10 μM) were used as the negative and positive controls, respectively. NGN2-iNs were imaged to monitor neuronal health 24h later, followed by direct lysis to release RNA and RealSeq small RNA library preparation. Illumina sequencing was performed in six batches, each indexed with 384 multiplex barcodes. Exclusion criteria included cell death, neurite degeneration, loss of cells during washes, enrichment of astrocytes, or less than 1,000 total annotated read counts.

first: Sergio

last: Barberan-Soler

first: Colin

last: Hortman

first: Lien

middle: D

last: Nguyen

first: Anna

last: Krichevsky


title: RAAS-Deficient Organoids Reveal that Delayed Angiogenesis Is The Pathomechanism Underlying Autosomal Recessive Renal Tubular Dysplasia

city: Cincinnati

first: Konrad

last: Thorner

state: OH

address: 3333 Burnet Ave

institute: Cincinnati Children's Hospital Medical Center

summary: Autosomal Recessive Renal Tubular Dysgenesis (AR-RTD) is a fatal genetic disorder affecting proximal tubule (PT) development in patients harboring mutations in genes comprising the Renin–Angiotensin–Aldosterone System (RAAS). To uncover the pathomechanism of AR-RTD, we differentiated ACE and AGTR1 deficient pluripotent stem cells and reprogrammed AR-RTD patient cells into kidney organoids. Marker analyses confirmed that all mutant and control organoids generated PT in room air (21% O2) or under hypoxic conditions (2% O2). Mature (d24) AGTR1-/- and control organoids transplanted under the kidney capsule of immunodeficient mice engrafted and differentiated well, as did renal vesicle stage (d14) control organoids. By contrast, d14 AGTR1-/- organoids failed to engraft due to insufficient pro-angiogenic VEGF-A expression. When grown under hypoxic conditions VEGF-A expression was stimulated and organoids engrafted. Thus, PT dysgenesis in AR-RTD is a non-autonomous consequence of a developmental delay in VEGF-A induction linking ANGII pro angiogenic role to PT dysgenesis.

overall_design: Bulk RNA-seq was performed comparing unmodified iPSCs to those with CRISPR/Cas9 deletion of either ACE or ATGR1 genes.

first: Naomi

last: Pode-Shaked

first: Megan

last: Slack

first: Nambirajan

last: Sundaram

first: Ruth

last: Schreiber

first: Benjamin

last: Dekel

first: Michael

last: Helmrath

first: Raphael

last: Kopan


title: Dissecting long non-coding RNAs derived from microRNA genes in hematopoiesis (Illumina short-read sequencing)

city: Beijing

first: Yue

last: Huo

state: Beijing

address: Dongcheng District, Dongdan 3 Tiao 5 Hao

department: State Key Laboratory of Medical Molecular Biology, Department of Biochemistry and Molecular Biology, Institute of Basic Medical Sciences

country: China

institute: Chinese Academy of Medical Sciences, School of Basic Medicine Peking Union Medical College

summary: Genome-wide transcription produces a complex population of transcripts and a wide variety of functional lncRNAs occasionally evolve from them. Here, we identify and characterize miRNA gene originated lncRNAs (molncRNAs) by employing Pacific Biosciences long-read sequencing. Combining with RNA-sequencing in differentiating hematopoietic erythrocytes, we construct a global molncRNA landscape during human erythropoiesis. Among them, miR-301b and miR-130b originated lncRNA, molnc-301b, orchestrates erythropoiesis independent of miRNAs. Mechanistically, molnc-301b attenuates the transcription of a series of erythropoietic and translation-associated genes by antagonizing SMARCA5's chromatin binding. Furthermore, molnc-301b perturbs translation of key erythroid mRNAs selectively, as GATA1 and FOS. Our findings identify a novel class of lncRNA and reveal the essential role of molnc-301b in erythroid differentiation. Of note, molnc-301b orchestrates erythropoiesis via jointly controlling the transcription and translation of key erythroid regulators. In addition, we finally establish functional molncRNA screen platform by using large-scale pooled CRISPR-based single-cell RNA-seq, and uncover 36 functional molncRNAs in erythrocytes. These results thus unearth a previously unannotated class of lncRNAs and provide a useful platform for the discovery and functional study of molncRNAs.

overall_design: smallRNA-seq, Poly(A)+ RNA-seq, ChIRP-seq, Ribo-seq and RNA-seq datasets.

first: Weiqian

last: Li

last: Ren

first: Fang

last: Wang

first: Jia

last: Yu


title: Loss of H3.1K27me1 in Arabidopsis confers resistance to Geminivirus by sequestering DNA repair proteins onto host genome [RNA-seq]

status: Public on Oct 17 2023

city: college station

first: Zhen

state: Texas

address: 300 Olsen Blvd,

department: Department of Biochemistry & Biophysics

institute: Texas A & M University

summary: Transcriptome-wide association studies (TWAS) to pinpoint enhanced expression of genes involved in DNA repair attributed to viral resistance of atxr5 atxr6.

overall_design: To investigate how the transcriptome is reprogramed upon virus infection, we performed comprehensive TWAS with high quality reads from mock-treated and virus-treated samples. Among genes showed enhanced expression upon virus inoculation, we observed that 365 genes that were expressed at higher levels in atxr5 atxr6 compared to Col-0, brca1 atxr5 atxr6, mbd9 atxr5 atxr6 and sac3b atxr5 atxr6, in both mock-treated and virus-inoculated samples. GO analysis classified the top three enriched biological processes into response to DNA damage, DNA repair, and DNA recombination.


title: Loss of H3.1K27me1 in Arabidopsis confers resistance to Geminivirus by sequestering DNA repair proteins onto host genome

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

overall_design: Refer to individual Series


title: Dissecting long non-coding RNAs derived from microRNA genes in hematopoiesis (scRNA sequencing)

summary: Albeit majority of eukaryotic genome can be pervasively transcribed to a diverse population of lncRNAs and various subtypes of lncRNA have been discovered. However, the genome-wide study of miRNA-derived lncRNAs is still lacking. Here, we report that over 800 miRNA gene-originated lncRNAs (molncRNAs) generated from miRNA loci. One of them, molnc-301b from miR-301b and miR-130b, functions as an "RNA decoy" to facilitate dissociation of the chromatin remodeling protein SMARCA5 from chromatin and thereby sequester transcription and mRNA translation. Specifically, molnc-301b attenuates erythropoiesis via mitigating the transcription of erythropoietic and translation-associated genes, such as GATA1 and FOS. In addition, we have established a useful and powerful CRISPR screen platform to characterize the biological functions of molncRNAs at large-scale and single-cell level, and identified 26 functional molncRNAs in hematopoietic cells. Collectively, we focused on miRNA-derived lncRNAs, deciphered their landscape during normal hematopoiesis, and comprehensively evaluated their potential roles.

sample_organism:
Homo sapiens
synthetic construct

platform_organism:
Homo sapiens
synthetic construct

overall_design: We designed 3 pooled gRNA libraries: library 1 (molncRNA(-) and internal control) contained more than 2 pairs of gRNAs for each of 12 internal controls (ICs) and the 361 miRNA loci, which were targeted by D1 and D2 gRNAs recognizing the 1-3 kb or 0.5-1 kb downstream region of miRNA precursor to specifically perturb molncRNAs' expression. Library 2 (molncRNA(-/-) and internal control) contained gRNAs for the same target genes as library 1, but recognized the 1-2 kb upstream or 1-3 kb downstream region of the miRNA precursors (U1 and D1 gRNAs) to knock out both molncRNAs and the cognate miRNAs). Library 3 (non-targeting control library) consisted of 2,456 pairs of non-targeting control (NTC) gRNAs. We have applied three replicates for library1 and library2. K562 cells were infected with a mix of lentiviruses expressing Cas9-mCherry at a multiplicity of infection (MOI) ~50 and gRNA-EGFP at a MOI ~4. Infected EGFP+/mCherry+ cells were sorted for 10x Genomics 3' scRNA-seq poly(A)-primed platform, and their corresponding gRNAs were further amplified using a specific amplification protocol(Hill et al., 2018).


title: Bulk and single-nucleus RNA sequencing highlight immune pathways induced in individuals during an Ixodes scapularis tick bite [RNA-Seq]

city: Hangzhou

first: Xiaotian

last: Tang

state: Zhejiang

address: Agro-Bio-Environ Building, Zhejiang University

department: Entomology

institute: Zhejiang University

summary: We performed RNA sequencing of skin and blood of persons presenting with naturally acquired, attached Ixodes scapularis ticks.

overall_design: We leveraged RNA sequencing to define the global immune events from skin biopsies and peripheral blood in individuals with actively attached naturally acquired I. scapularis.

first: Erol

last: Fikrig


title: Bulk and single-nucleus RNA sequencing highlight immune pathways induced in individuals during an Ixodes scapularis tick bite [snRNA-Seq]

summary: We performed single-nucleus RNA sequencing (snRNA-seq) of skin and blood of persons presenting with naturally acquired, attached Ixodes scapularis ticks.

overall_design: We leveraged single-nucleus RNA sequencing (snRNA-seq) to define the global immune events from skin biopsies and peripheral blood in individuals with actively attached naturally acquired I. scapularis.

first: Yingjun

last: Cui


title: Transcriptome profiling of human endothelial cells undergoing endothelial-to-mesenchymal transition

status: Public on Oct 16 2023

city: St. Petersburg

first: JIANHUA

last: XIONG

state: FL

address: 501 6th Ave South, REB Bldg, 3rd Floor, Rm 3111, REB/Rm 3111

institute: Johns Hopkins University/Johns Hopkins All Children's Hospital

summary: To investigate the transcriptome of endothelial cells undergoing endothelial-to-mesenchymal transition, transcription profiling was performed on primary human endothelial cells in the presence or absence of 40mM acetate following control or cytokine treatment for 4 days. We then performed gene expression profiling analysis using data obtained from RNA-seq of primary human endothelial cells.

overall_design: RNA sequencing for transcription profiling was performed on primary human endothelial cells in the presence or absence of 40mM acetate following control or cytokine treatment for 4 days.

first: Jianhua

last: Xiong

first: Junfang

last: Lyu


title: RNA-seq profiling of Drosophila larval ring glands of kdm5 null mutants

city: Bronx

middle: Francis

last: Rogers

state: NY

address: 1300 Morris Park Ave, Ullmann 809

department: Genetics

institute: Albert Einstein College of Medicine

summary: kdm5 is an essential gene in Drosophila that has critical developmental roles in the prothoracic gland cells of the larval ring gland.  We performed a bulk transcriptome analysis of the larval ring gland in w[1118] (wild type) and kdm5[140] (null mutant) in order to identify  genes in the prothoracic gland involved in the lethality of kdm5 null mutants.  We found that the absence of kdm5 causes dysregulation of genes involved in various metabolic pathways.  In particular, genes both bound by KDM5 and differentially expressed in this cell type are involved in regulation of mitochondrial biology and autophagy.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for Drosophila larval ring glands of wild type and kdm5 null animals

middle: F

first: Owen

last: Marshall

first: Julie

last: Secombe


title: Genome-winding binding profile of KDM5 in Drosophila larval prothoracic gland cells via Targeted DamID analysis

summary: kdm5 is an essential gene in Drosophila that has critical developmental roles in the prothoracic gland cells of the larval ring gland.  To profile KDM5 binding within these cells and this developmental stage, we performed Targeted DamID (TaDa). By profiling the nearest genes to significant TaDa peaks (FDR < 0.01), we identified 5815 candidate KDM5 target genes.  Interestingly, 42% of these candidate KDM5 target genes appear to be conserved across multiple cellular contexts in Drosophila and span many cellular processes for future investigation.

overall_design: Targeted DamID (TaDa) for profiling KDM5 binding in Drosophila prothoracic gland cells


title: Bulk and single-nucleus RNA sequencing highlight immune pathways induced in individuals during an Ixodes scapularis tick bite


title: KDM5 binding and kdm5 null mutant differential expression profiling of Drosophila prothoracic gland cells


title: Pro-inflammatory macrophages impair skeletal muscle regeneration in ischemic-damaged limbs by inducing precocious differentiation of satellite cells [RNA-seq]

first: Yarui

last: Diao

address: 307 Research Drive

department: Cell Biology

summary:
To validate the findings from scRNA-seq, we purified CD11b+/F480+ macrophages by FACS from the hind limb muscle of C57BL/6 (n=3) and BALB/c (n=2) mice at 3-days post HLI for bulk RNA-seq analysis
We then performed gene expression profiling analysis using data obtained from RNA-seq of isolated macrophages from C57BL/6 and BALB/c mice at day 3 after HLI surgery.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for macrophages of C57BL/6 and BALB/c mice

first: Kevin

last: Southerland

first: Yueyuan

last: Xu

first: Derek

middle: T

last: Peters

first: YaruI


title: MicroRNA-218 instructs proper assembly of hippocampal networks

status: Public on Oct 15 2023

city: Modena, Italy

first: Giordano

last: Lippi

address: 10550 North Torrey Pines, DNC-202

institute: The Scripps Research Institute

summary: The assembly of the mammalian brain is orchestrated by temporally coordinated waves of gene expression. Post-transcriptional regulation of gene expression by microRNAs (miRNAs) is a key aspect of this program. Indeed, deletion of neuron-enriched miRNAs induces strong developmental phenotypes, and miRNA levels are altered in patients with neurodevelopmental disorders. However, the mechanisms used by miRNAs to instruct brain development remain largely unexplored. Here, we identified miR-218 as a critical regulator of hippocampal assembly. MiR-218 is highly expressed in the hippocampus and enriched in both excitatory principal neurons (PNs) and GABAergic inhibitory interneurons (INs). Early life inhibition of miR-218 results in an adult brain with a predisposition to seizures. Changes in gene expression in the absence of miR-218 suggest that network assembly is impaired. Indeed, we find that miR-218 inhibition results in the disruption of early depolarizing GABAergic signaling, structural defects in dendritic spines, and altered intrinsic membrane excitability. Conditional knockout of miR-218 in INs, but not PNs, is sufficient to recapitulate long-term instability. Finally, de-repressing Kif21b and Syt13, two miR-218 targets, phenocopies the effects on early synchronous network activity induced by miR-218 inhibition. Taken together, the data suggest that miR-218 orchestrates formative events in PNs and INs to produce stable networks.

overall_design:
To identify targets of miR-218, we conducted three different RNA-sequencing experiments
1) Locked nuclei acid miR antagomiRs targeting either miR-218 or a scrambled sequence were injected into the dorsal hippocampus of P2 mice (3/condition). The dorsal hippocampus was dissected at P8 for RNA extraction. 2) The dorsal hippocampi of P15 miR-218-1-/-; miR-218-2+/- mice (n=5) and miR-218-1+/+; miR-218-2+/+ (n=3) mice were dissected and RNA extracted for RNA-sequencing
3) We used FACS to isolate VGluT2+ neurons from the dorsal hippocampus of 4 P8 Ai14; VGluT2-Cre mice and to isolate VGAT+ neurons from the dorsal hippocampus of 3 Ai14; VGAT-IRES-Cre mice.

first: Seth

last: Taylor

first: Mariko

last: Kobayashi


title: LSM14B is essential for oocyte meiotic maturation by regulating maternal mRNA storage and clearance

city: Jinan

first: Yanling

last: Wan

address: Medicine west wenhua road

institute: Center for Reproductive Medicine, Shandong University

summary: Fully grown oocytes remain transcriptionally quiescent, yet many maternal mRNAs are synthesized and retained in growing oocytes. We now know that maternal mRNAs are stored in a structure called the mitochondria associated ribonucleoprotein domain (MARDO). But the components and functions of MARDO remain elusive. Here, we found that LSM14B knockout prevents the proper storage and timely clearance of mRNAs (including Cyclin B1, Btg4, and other mRNAs that are translationally activated during meiotic maturation), specifically by disrupting MARDO assembly during oocyte growth and meiotic maturation. With decreased levels of storage and clearance, the LSM14B knockout oocytes failed to enter meiosis II, ultimately resulting in female infertility. Our results demonstrate the function of LSM14B in MARDO assembly, couple the MARDO with mRNA clearance and oocyte meiotic maturation.

overall_design: WT and LSM14B-none oocytes [GV,MI and 14h after hCG (denoted as MII)stages] for three replicates are performed RNA sequencing.

first: Hongbin

last: Liu

first: Shuang

last: Yang


title: Pro-inflammatory macrophages impair skeletal muscle regeneration in ischemic-damaged limbs by inducing precocious differentiation of satellite cells [scRNA-seq]

summary: To investigate the molecular mechanisms responsible for failed skeletal muscle repair in the Chronic limb threatening ischemia (CLTI) limb. We used single cell RNA sequencing (scRNA-seq) to profile the transcriptomes of the skeletal muscle specimens.

overall_design: Hindlimb ischemia (HLI) or sham surgery was performed on young adult C57BL/6 and BALBc mice. Hindlimb skeletal muscle tissue was collected on post-op day 1 (sham and HLI) and post-op days 3 and 7 (HLI only). Skeletal muscle was dissociated into single cells, which were analyzed by scRNA-seq.


title: Effect of Notch2 in chondrocytes

city: Farmington

first: Ernesto

last: Canalis

state: CT

address: 263 Farmington Avenue

department: Orthopaedic Surgery

institute: UConn Health

summary:
To investigate the role of Notch2 in the inflammatory response in chondrocytes, we cultured primary chondrocytes derived from wild type and Notch2 mutant mice and treated TNFalpha or vehicle.
We then performed gene expression profiling analysis using data obtained from RNA-seq of wild type and Notch2 mutant chondrocytes.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for wild type and Notch2 mutant chondrocytes with/without TNFalpha

first: Jungeun

last: Yu


title: Effects of Id1 depletion in TAMs on the genes expression of MC38 cells.

first: Yang

last: Chen

address: 1 Xian Nong Tan Street

institute: Peking Union Medical Collage

summary: We report the application of high-throughput mRNA sequencing to explore the effect of Id1 depletion in TAMs on its  gene expression . Different groups of TAMs were isolated from the subcutaneous  tumors in Id1f/f or myeloid cell lineage speciﬁc Id1 deficient mice (Id1-Lyz-KO) .

overall_design: Examination of two groups of TAMs with 3 replicates separately.

last: Shang

first: Chen

first: Fang

last: Hua


address: 2# NanWei Road XiCheng District

department: Pharmacology

institute: Institute of Materia Medica

summary: We report the application of high-throughput mRNA sequencing to explore the effect of Id1 depletion in TAMs on gene expression of MC38 cells. Different groups of TAMs were isolated from the subcutaneous  tumors in Id1f/f or myeloid cell lineage speciﬁc Id1 deficient mice (Id1-Lyz-KO) . MC38 cells were then mixed with two groups of TAMs and be implanted in C57BL/6J mice. CD45- cells were then isolated from the subcutaneous tumors described above to do the transcriptome sequencing.

overall_design: Examination of two groups of MC38 cells with 3 replicates separately.


title: Single-cell RNA Sequencing (scRNA−seq) Identifies L1CAM as a Key Mediator between Epithelial Tuft Cell and Innate  Lymphoid Cell in the Colon of Hnrnp I Knockout Mice

status: Public on Oct 14 2023

city: Urbana

first: Guanying

middle: Bianca

state: Illinois

address: 905 S Goodwin Ave Rm #472B1

department: Food Science and Human Nutrition

institute: University of Illinois at Urbana-Champaign

summary: (1) Background: Knockout (KO) of heterogeneous nuclear ribonucleoprotein I (Hnrnp I) in mouse intestinal epithelial cells (IECs) induced a severe inflammatory response in the colon, followed by hyperproliferation. This study aimed to investigate the epithelial lineage dynamics and cell–cell communications that underlie inflammation and colitis. (2) Methods: Single cells were isolated from the colons of wildtype (WT) and KO mice and used in scRNA−seq. Whole colons were collected for immunofluorescence staining and cytokine assays. (3) Results: from scRNA−seq, the number of DCLK1 + colonic tuft cells was significantly higher in the Hnrnp I KO mice compared to the WT mice. This was confirmed by immunofluorescent staining of DCLK1. The DCLK1 + colonic tuft cells in KO mice developed unique communications with lymphocytes via interactions between surface L1 cell adhesion molecule (L1CAM) and integrins. In the KO mice colons, a significantly elevated level of inflammatory cytokines IL4, IL6, and IL13 were observed, which marks type−2 immune responses directed by group 2 innate lymphoid cells (ILC2s). (4) Conclusions: This study demonstrates one critical cellular function of colonic tuft cells, which facilitates type−2 immune responses by com-municating with ILC2s via the L1CAM–integrins interaction. This communication promotes pro−inflammatory signaling pathways in ILC2, leading to the increased secretion of inflammatory cytokines.

overall_design:
Only male mice were used in this study and were from the cross of Hnrnp Iflox/flox; VillinCre/+ males with Hnrnp Iflox/flox females. The wild−type (WT) group included the Hnrnp Iflox/flox mice, and the knockout (KO) group were the Hnrnp Iflox/flox VillinCre/+ mice. The experimental unit for this study was a single animal.
For ScRNA−seq, littermates were selected to minimize potential confounding factors introduced by genetic or environmental variability. This was an a priori criterion set to ensure consistency and reliability in our findings. For other experiments, the primary inclusion criterion was the age of the animals. Animals with an age difference of no more than one week were selected. Other than age and the littermate criteria for the sequencing experiment, no additional inclusion or exclusion criteria were established. All data points from each experimental group were included in the analysis. There were no exclusions
Immediately after euthanasia and necropsy, whole colon tissues from WT and KO male mice (littermates, 3−month−old, male, n = 3) were selected for cell isolation and ScRNA-seq.

middle: B

first: Yuan-Xiang

last: Pan

first: Wenyan

last: Mei

first: Hong


title: Mucosal-associated invariant T cells contribute to suppression of inflammatory myeloid cells in immune-mediated kidney disease

status: Public on Oct 13 2023

city: Hamburg

first: Jan-Eric

last: Turner

address: Martinistraße 52

country: Germany

institute: University Medical Center Hamburg-Eppendorf

summary: Mucosal-associated invariant T (MAIT) cells have been implicated in various inflammatory diseases of barrier organs, but so far, their role in kidney disease is unclear. Here we report that MAIT cells that recognize their prototypical ligand, the vitamin B2 intermediate 5-OP-RU presented by MR1, reside in human and mouse kidneys. Single cell RNAseq analysis reveals several intrarenal MAIT subsets, and one, carrying the genetic fingerprint of tissue-resident MAIT17 cells, is activated and expanded in a murine model of crescentic glomerulonephritis (cGN). An equivalent subset is also present in kidney biopsies of patients with anti-neutrophil cytoplasmatic antibody (ANCA)-associated cGN. MAIT cell-deficient MR1 mice show aggravated disease, whereas B6-MAITCAST mice, harboring higher MAIT cell numbers, are protected from cGN. The expanded MAIT17 cells express anti-inflammatory mediators known to suppress cGN, such as CTLA-4, PD-1, and TGF-β. Interactome analysis predicts CXCR6 – CXCL16-mediated cross-talk with renal mononuclear phagocytes, known to drive cGN progression. In line, we find that cGN is aggravated upon CXCL16 blockade. Finally, we present an optimized 5-OP-RU synthesis method which we apply to attenuating cGN in mice. In summary, we propose that CXCR6+ MAIT cells might play a protective role in cGN, implicating them as a potential target for anti-inflammatory therapies.

overall_design: To investigate transcriptional programs of murine MAIT cells in the crescentic glomerulonephritis (cGN) model, we sorted 5-OP-RU+ MAIT cells from the pooled kidneys of CAST mice on day 1 and day 9 after the induction of cGN, as well as of naive CAST mice. Additionally, we sorted CD45+ non-MAIT cells on day 9. Each libraries were created with the 10x Genomics Chromium Singel Cell 5'v1.1 reagent kit and sequenced on Illumina NovaSeq600 with 150 base pairs and paired-end configurations.

first: Ann-Christin

last: Gnirck

first: Marie-Sophie

last: Philipp

first: Alex

last: Waterhölter

first: Malte

last: Wunderlich

first: Nikhat

last: Shaikh

first: Virginia

last: Adamiak

first: Lena

last: Henneken

first: Tobias

last: Kautz

first: Tingting

last: Xiong

first: Daniela

last: Klaus

first: Pascal

last: Tomczyk

first: Mohamad

last: Al-Bahra

first: Dirk

last: Menche

first: Mark

last: Walkenhorst

first: Olivier

last: Lantz

first: Anne

last: Willing

first: Manuel

middle: A

last: Friese

last: Huber

first: Christian

last: Krebs

first: Ulf

last: Panzer

last: Kurts


title: Hi-C profiling of SynVIII strain

city: New York

first: Jitong

last: Cai

address: 435 E 30th street

department: NYU Langone Health

institute: NYU Langone Health

summary: Building synthetic yeast chromosomes with thousands of designer modifications empowers probing of genome flexibility on an unprecedented scale. Here, we describe the assembly and characterization of synthetic chromosome VIII (synVIII), which served as a platform for further genome manipulation of an essential functional element, the centromere. Recent findings suggest that centromeres may have distinct characteristics and thus relocating the centromere to various positions along a single chromosome (VIII) enables us to directly test the suitability of several ectopic destinations for centromere function and stability. We used a novel CRISPR/Cas9-mediated approach, Single-Step Intrachromosomal Centromere Transplantation (SSICT), to relocate the 118 bp point centromere sequence of CEN8. Successful transplantation of CEN8 invariably resulted in chromosome VIII aneuploidy regardless of ectopic centromere position in wild-type or synVIII strains and euploid structural variants hinted that including left-flanking pericentromeric sequences of CEN8 allowed ectopic function. Plasmid loss rate assays revealed that specific left-flanking sequences increased stability of a CEN8-containing circular minichromosome and reduced incidence of aneuploidy during SSICT. Together, these results show that the minimal point centromere of budding yeast is insufficient to confer chromosome stability when transplanted ectopically, and is rescued by left-flanking pericentromeric sequence. This is an important consideration for future genome engineering projects involving transplantation of functional elements.

overall_design: We perform Hi-C to assess whether two centromere copies in the SynVIII strain was structurally functional (Hi-C).

first: Stephanie

last: Lauer

first: Jef

last: Boeke


title: Loss of H3.1K27me1 in Arabidopsis confers resistance to Geminivirus by sequestering DNA repair proteins onto host genome [ChIP-seq]

first: Zhen

last: Wang

summary: We identified RAD51 and RPA1A as partners of virus-encoded Rep protein. The two DNA repair proteins showed increased binding to heterochromatic regions and defense-related genes in atxr5 atxr6 vs wild-type plants. Consequently, the proteins had reduced binding to viral DNA in the mutant, thus hampering viral amplification. Additionally, RAD51 recruitment to the host genome arose via BRCA1, HOP2, and CYCB1;1, and this recruitment was essential for viral resistance in atxr5 atxr6. Thus, Geminiviruses adapt to healthy plants by hijacking DNA repairing pathways, whereas the unstable genome, triggered by lack of H3.1K27me1, could retain DNA repairing proteins to suppress viral infection in atxr5 atxr6.

overall_design: We first validated the specificity of anti-RAD51 and RPA1A antibodies (Extended Data Fig. 8. a-f), and then performed genome-wide chromatin immunoprecipitation-sequencing (ChIP-seq) for the two proteins in Col-0 and atxr5 atxr6.

first: Chun-Yip

last: Tong


title: Rational design of a JAK1-selective siRNA inhibitor for the modulation of autoimmunity in the skin

city: Worcester

first: Samuel

last: Hilebrand

state: MA

address: 368 Plantation Street AS4.1016

department: RNA Therapeutics Institute

institute: University of Massachusetts Chan Medical School

summary: Inhibition of Janus kinase (JAK) family enzymes has surged as a popular strategy for treating inflammatory and autoimmune skin diseases. In the clinic, current available small molecule JAK inhibitors show distinct efficacy and safety profiles, likely reflecting their variable selectivity for JAK subtypes. Nonselective inhibition of multiple JAK enzymes is associated with increased side effects and has resulted in FDA-mandated black box warnings. Developing therapeutics to enable absolute JAK subtype selectivity is challenging and has not yet been achieved. Here, we harness RNA interference (RNAi) for rationally designing small interfering RNA (siRNA) therapeutics that offer sequence-specific gene silencing of JAK1, thus narrowing the spectrum of action on JAK-dependent cytokine signaling pathways to maintain efficacy and improve safety. We developed a fully chemically modified siRNA that supports efficient silencing of JAK1 expression in human skin explant and functional modulation of JAK1-dependent inflammatory signaling. A single injection of the JAK1 siRNA into mouse skin enables JAK1 silencing for 5 weeks in vivo. In a mouse model of vitiligo, local administration of the JAK1 siRNA significantly reduces skin infiltration of autoreactive CD8+ T cells and prevents epidermal depigmentation. This work outlines a robust framework for designing targeted RNAi-based therapies and establishes a path toward novel siRNA treatments for inflammatory and autoimmune skin diseases.

overall_design: PolyA RNA sequencing from human skin biopsies treated with PBS, non-targeting siRNA, or JAK1-targeting siRNA with and without stimulation with 10 ng/mL of recombinant IFN-γ and 10 ng/mL of TNF-α

first: Qi

last: Tang

first: Hassan

middle: H

last: Fakih

first: Mohammad

middle: Z

last: UI Abideen

last: Hildebrand

first: Khashayar

last: Afshari

first: Katherine

last: Gross

first: Jacquelyn

last: Sousa

first: Allison

middle: S

last: Maebius

first: Christina

last: Bartholdy

first: Pia

last: Pernille Søgaard

first: Malene

last: Jackerott

first: Vignesh

last: Hariharan

first: Ashley

last: Summers

first: Xueli

last: Fan

first: Ken

last: Okamura

first: Kathryn

last: Monopoli

first: David

last: Cooper

first: Dimas

last: Echeverria

first: Brianna

last: Bramato

first: Nicholas

last: McHugh

first: Raymond

middle: C

last: Furgal

first: Annabelle

last: Biscans

first: Jane

last: Chuprin

first: Nazgol-Sadat

last: Haddadi

first: Shany

last: Sherman

first: Ümmügülsüm

last: Yıldız-Altay

first: Claire

last: Bouix-Peter

first: Carine

last: Blanchard

first: Adam

last: Clauss

first: Julia

last: Alterman

first: Anastasia

last: Khvorova

first: John

last: Harris


title: Transcription factor EBF1 non-cell autonomously regulates cardiac growth and differentiation

status: Public on Oct 12 2023

city: New York City

first: Alireza

last: Khodadadi-Jamayran

address: 550 1st Ave, MSB 304

department: Division of Advanced Research Technologies (DART)

institute: New York University, NYU Langone Medical Center

summary: Reciprocal interactions between non-myocytes and cardiomyocytes are implicated in the control of cardiac growth and differentiation.  Here, we report the identification of early B-cell factor 1 (Ebf1) as a transcription factor highly enriched in non-myocytes that potently regulates heart development.  Postnatal Ebf1 deficient hearts are characterized by marked myocardial hypercellularity and reduced cardiomyocyte size, as well as ventricular conduction system hypoplasia and conduction system disease.  Growth abnormalities in Ebf1 knockout hearts are observed as early as embryonic day 13.5, with dysregulated cell cycling, myocardial hyperplasia, and immature trabecular morphology.  Transcriptional profiling of Ebf1-deficient non-myocytes isolated from embryonic hearts demonstrates dysregulation of Polycomb repressive complex 2 (PRC2) targets, while ATAC-Seq reveals differences in chromatin accessibility near many of these same genes. Gene set enrichment analysis of differentially expressed genes in cardiomyocytes isolated from E13.5 hearts of wildtype and Ebf1 deficientmice reveals significant enrichment of MYC targets, and consistent with this finding, we observe increased abundance of MYC in mutant hearts.  Mechanistically, we show that EBF1-deficient non-myocytes, but not wildtype non-myocytes, are sufficient to induce excessive accumulation of nuclear-localized MYC protein in co-cultured wildtype cardiomyocytes. Finally, we demonstrate that BMP signaling induces Ebf1 expression in embryonic heart cultures and controls a gene program enriched in EBF1 targets.  Taken together, these results reveal a novel non-cell autonomous pathway controlling cardiac growth and development.

overall_design: Gene transcirptional profiling was performed on embryonic day 13.5 mouse heart cells after dissociation and FACS sorting using PECAM1 antibody and TMRM dye to identify endocardial cells and cardiomyocytes respectively. ATAC-Seq was performed on non-myocyte cells from E13.5 mouse hearts isolated by preplating.

first: Eugene

last: Kim

first: Glenn

last: Fishman


title: Trim-away depletion of Zona Pellucida 3 (ZP3) protein in mouse zygotes (analysis after 10h)

city: Muenster

first: Michele

last: Boiani

address: Roentgenstrasse 20

institute: Max Planck Institute Muenster

summary: Mutation studies always defined the functions of the zona pellucida (ZP) as extracellular, namely: to encase the oocytes in ovarian follicles, to ensure species-specific sperm binding, and to dampen shear stress on the embryo surface. Therefore, mutations in the three ZP mouse genes ZP1, ZP2 or ZP3 cause primary infertility due to empty follicles, polyspermic fertilization or harmful contact between embryos and oviductal epithelium. However, the concepti of ZP2-null and ZP3-null oocytes were still unviable also when the defects were obviated by monospermic fertilization in vitro and blastocyst transfer to uterus (PMID 11245577). This suggests that the tasks of ZPs don’t end in the extracellular space as previously assumed, but there may be also intracellular functions yet to be discovered. The present study tested if experimentally induced degradation of intracellular ZP3 impacted on the development and transcriptome of mouse embryos. To this end we degraded ZP3 using its antibody in conjunction with the ubiquitin-protein ligase TRIM21. This method is known as 'Trim-away' (PMID 29153837). Briefly, in this method a cell (e.g. oocyte) expressing TRIM21 is supplied e.g. injected with a specific antibody to a protein of interest, in this case ZP3. As a result, the ternary complex (target protein-antibody-TRIM21) is destroyed in the proteasome. TRIM21 is here always to be understood as translation product of microinjected mCherry-Trim21 mRNA. We compared two experimental groups, as follows. Pronuclear-stage oocytes (B6C3F1 x CD1) were microinjected with approx. 100 picoliters of mix comprised of mCherry-Trim21 mRNA 0.2 mg/mL + anti-ZP3 antibody (Proteintech 21279-1-AP) 1 mg/mL + dextran beads 0.02 mg/mL, forming a group named 'Trim-away ZP3' group, in triplicate. As a reference, pronuclear-stage oocytes were microinjected with the same mixture as above, except that the antibody buffer was used in lieu of the antibody itself, in triplicate, forming a group named ‚no Trim'. To identify differently expressed genes we compared group 'Trim-away ZP3' with group ‘no Trim’. Ten hours after microinjection, embryos were collected and lysed for transcriptome analysis. Transcriptome analysis revealed that embryos of group 'Trim-away ZP3' and group ‘no Trim' differed in gene expression and were resolved in principal component analysis. The data support a conclusion that ZP3 found inside the embryo was not merely a remnant from oogenesis, but served an intracellular, post-fertilization role during mouse preimplantation development.

overall_design: Biological replicates were analyzed for each of two experimental conditions: 'Trim-away of ZP3' (protein degradation) in triplicate (r1, r2, r3) and ‚no Trim’ (Trim-away cocktail containing the buffer of the antibody in place of the antibody) in triplicate (r1, r2, r3). Trim-away followed to the coinjection of antibody anti ZP3 (Proteintech 21279-1-AP; 1 mg/mL), mCherry-Trim21 mRNA (0.2 mg/mL) and oregon green dextran beads (0.02 mg/mL), resulting in protein degradation in the proteasome. No Trim-away reaction took place after coinjection of mCherry-Trim21 mRNA (0.2 mg/mL), ZP3 antibody buffer, and oregon green dextran beads (0.02 mg/mL). RNA was extracted and analyzed at 10 hours past injection, when the embryos were at the pronuclear stage.

first: Steffen

last: Israel


title: High neighborhood deprivation impacts DNA methylation and gene expression in cancer-related genes (RNA-Seq)

city: Bethesda

first: Stefan

last: Ambs

state: MD

address: Building 37, Room 3050B

department: CCR

institute: NCI

summary: Background: Low socioeconomic status (SES) has been linked to chronic stress and poorer health outcomes for marginalized communities in the U.S. The biological processes mediating the impact of SES on health to promote chronic diseases, such as cancer, remain poorly understood. Here, we aim to determine how changes in DNA methylation are linked to neighborhood-level socioeconomic deprivation, thereby promoting cancer initiation and progression through cancer- and immune-related pathways.     Methods: We extracted DNA from 402 fresh frozen tissues from 289 women in the NCI-Maryland Breast Cancer Cohort, including 185 tumor samples, 113 additional paired adjacent normal samples, and 104  normal tissues from reduction mammoplasty. Census-tract level socioeconomic deprivation was measured via a Neighborhood Deprivation Index (NDI) using the geocoded addresses from our study participants. DNA methylation values were acquired using the Illumina Infinium MethylationEPIC 850K Beadchip. We conducted a differentially methylated probe (DMP) analysis between tumor and adjacent normal tissue to characterize methylation profiles by NDI status. We also used methylCIBERSORT to estimate immune cell subpopulation differences by tissue type and NDI status.     Results: NDI was significantly associated with self-reported race in our cohort in both normal (p<0.0001, 95% CI -4.32, -2.58) and tumor (p<0.0001, 95% CI -4.22, -2.79) tissue. Our DMP analysis showed more significant methylation events (hypo- or hyper- methylation of the tumor tissue compared to adjacent normal tissue) in the NDI high group compared to the NDI low group. We also identified five CpG sites that were significantly hypomethylated in the tumors of the NDI high group, including two tumor suppressor genes, LRIG1 (p=1.99 x10-10) and WWOX (p=6.47 x 10-9). These methylation changes translated to an inverse relationship between gene expression for these tumor suppressor genes and NDI. We also identified significantly lower neutrophils in the NDI high tumors compared to NDI low tumors (p=0.001, 95% CI -0.112, -0.028).     Conclusions: Overall, our analysis provides evidence that high neighborhood deprivation can lead to pro-tumorigenic changes in DNA methylation and gene expression that may impact the immune microenvironment, breast cancer progression, and overall survival.

overall_design: RNA was isolated from frozen breast tumors using TRIzol method. The mRNA profiles of the above breast tumors were generated by deep sequencing using Illumina NovaSeq S2.

first: Brittany

last: Lord

first: Emily

last: Rossi

first: Tiffany

last: Dorsey

first: Anuoluwapo

last: Ajao

first: Catherine

last: Pichardo

first: William

last: Wooten

first: Margaret

first: Wei

last: Tang

first: Ruby

last: Hutchison

first: Maeve

last: Bailey-Whyte


title: Methylation Array Data From PBMC Cell Lineages Derived From Control, At-Risk and Early RA Patients

city: La Jolla

first: Gary

last: Firestein

address: 9500 Gilman Dr

department: Division of Rheumatology, Allergy & Immunology

institute: University of California, San Diego School of Medicine

type: Methylation profiling by genome tiling array

summary: Data is derived from samples from three groups using the ACPA assay ‘anti-CCP3’: anti-CCP3(+) without IA (termed ‘At-Risk’), Early RA, and anti-CCP(-) Controls. Peripheral B, memory and naïve T cells were separated and analyzed for methylation array analysis.

overall_design: Bisulphite converted DNA from the 382 samples were hybridised to the Illumina Infinium MethylationEPIC chip.

first: Eddie

last: James

first: V

middle: M

last: Holers

first: Radhika

last: Iyer

first: E

last: Prideaux

first: Navin

middle: L

last: Rao

first: Cliff

last: Rims

first: Virginia

last: Muir

first: Sylvia

last: Posso

first: Michelle

last: Bloom

first: Amin

last: Zia

first: Serra

last: Elliott

first: Julia

last: Adamska

first: Rizi

last: Ai

first: Camille

last: Brewer

first: Jennifer

last: Seifert

first: LauraKay

last: Moss

first: Saman

last: Barzideh

first: M

middle: K

last: Demoruelle

first: Christopher

last: Striebich

first: Yuko

last: Okamoto

first: Enkhtsogt

last: Sainbayar

first: Alexandra

last: Crook

first: Ryan

last: Peterson

first: Lauren

last: Vanderlinden

last: Wang

first: David

last: Boyle

last: Robinson

first: Jane

last: Buckner

first: Kevin

last: Deane


title: Dual targeting of castration-resistant prostate cancer with a combined HDAC and PI3K inhibitor fimepinostat suppresses both adenocarcinoma and neuroendocrine tumor growth

city: Seattle

first: Ilsa

last: Coleman

address: 1100 Fairview Ave N, E2-112

department: Human Biology

institute: Fred Hutchinson Cancer Center

summary: Histone deacetylation is crucial for the regulation of chromatin structure, gene transcription and the epigenetic state of the cell.  With the emergence of the neuroendocrine (NE) phenotype in castration-resistant prostate cancer (CRPC), we hypothesized that the inhibition of HDAC1/2 would disrupt the epigenetic events involved in the conversion of prostate adenocarcinoma to a NE prostate cancer (NEPC) phenotype.  Additionally, monotherapies tend to have limited efficacy in cancer treatment and combination treatments show more promise.  Since the PI3K/AKT/mTOR pathway is a well-established tumor growth-promoting pathway and a promising target in CRPC and NEPC, we investigated the tumor suppressing activities of fimepinostat, a dual HDAC and PI3K inhibitor, on both prostate adenocarcinoma and NEPC. We evaluated multiple cell lines and patient-derived xenograft (PDX) models of prostate adenocarcinoma and NEPC to determine the roles of HDAC1/2 and AKT inhibition, both individually and in combination, in suppressing CRPC and NEPC tumor growth in vitro and in vivo.  HDAC1/2 inhibition in combination with PI3K/AKT inhibition was more efficacious than targeting each pathway alone, induced growth inhibitory effects (in some cases synergistic) through cell cycle inhibition and apoptosis across an array of prostate adenocarcinoma and NEPC patient-derived xenografts (PDX) and cell lines in vitro and in vivo. Molecular profiling revealed on-target effects of combined HDAC1/2 and PI3K/AKT inhibition as well as a common downregulation of Myc, E2F, DNA repair, and G2M checkpoint targets, independent of prostate cancer phenotype. Fimepinostat therapy was also associated with the suppression of lineage transcription factor expression including androgen receptor (AR) in prostate adenocarcinoma and Achaete-scute homolog 1 (ASCL1) in NEPC. Our findings demonstrate that dual HDAC1/2 and PI3K/AKT pathway inhibition leads to robust tumor growth inhibition in both adenocarcinoma and NEPC, suggesting that fimepinostat may represent a novel therapeutic for both prostate adenocarcinoma and NEPC phenotypes of CRPC.

overall_design: RNA sequencing of in vitro cultures of dissociated prostate tumor patient-derived xenograft (PDX) tumors and cell lines using Illumina TruSeq Library prep and sequenced on Illumina NovaSeq 6000.

first: Ailin

last: Zhang

first: Nathan

last: Lau

first: Alicia

last: Wong

first: Lisha

middle: G

last: Brown

first: Navonil

last: De Sarkar

first: Dapei

last: Li

first: Diana

last: DeLucia

first: Mark

middle: P

last: Labrecque

first: Holly

last: Nguyen

last: Conner

first: Daniel

middle: W

last: Lin

first: Eva

last: Corey

first: Joshi

last: Alumkal

first: Peter

last: Nelson

first: Colm

last: Morrissey

first: John

last: Lee


title: Release of Histone H3K4-reading transcription factors from chromosomes in mitosis is independent of adjacent H3 phosphorylation

city: Newcastle upon Tyne

first: Jonathan

last: Higgins

address: Framlington Place

department: Biosciences Institute

country: United Kingdom

institute: Newcastle University

summary: Histone modifications influence the recruitment of reader proteins to chromosomes to regulate events including transcription and cell division. The idea of a histone code, where particular combinations of modifications specify unique downstream functions, is widely accepted and can be demonstrated in vitro. For example, on synthetic peptides, phosphorylation of Histone H3 at threonine-3 (H3T3ph) prevents the binding of reader proteins that recognise trimethylation of the adjacent lysine-4 (H3K4me3), including the TAF3 component of TFIID. To study these combinatorial effects in cells, we analyzed the genome-wide distribution of H3T3ph and H3K4me3 during mitosis. We find that H3K4me3 hinders adjacent H3T3ph deposition in cells, and that the PHD domain of TAF3 can bind H3K4me3 in mitotic chromatin despite the presence of H3T3ph. Unlike in vitro, H3K4 readers are displaced from chromosomes in mitosis in Haspin-depleted cells lacking H3T3ph. H3T3ph is therefore unlikely to be responsible for transcriptional downregulation during cell division.

overall_design: ChIP-seq/CIDOP-seq was perfomed in asynchronous or mitotic enriched (nocodazole) HeLa S3 cells with 2-3 biological replicates each with corresponding input samples

first: Rebecca

middle: J

last: Harris

first: Maninder

last: Heer

first: Jennifer

last: Mitchell

last: Coxhead

last: Rico


title: Klf6 is required in thymic epithelial cells for normal thymus development [scATAC-seq]

first: Yongge

last: Zhao

address: 9000 Rockville Pike

summary:
Thymic epithelial cells (TEC) control T cell development and play essential roles in establishing self-tolerance. Transcription factors controlling TEC development are poorly characterized. We report that Klf6 plays a critical role in TEC development. Mice deficient for Klf6 in TEC had a hypoplastic thymus - evident from fetal stages into adulthood. Proliferation of TEC was not reduced, but a dramatic increase in the frequency of apoptotic TEC in fetal and adult thymus was observed. Among cortical TEC (cTEC), we found expansion of a previously unreported cTEC population expressing the transcription factor Sox10. Medullary TEC (mTEC) subsets were not equally impacted with Ccl21a+ mTEC I and Tuft-like mTEC IV being disproportionately affected. Consistent with these TEC defects, naïve conventional T cells and NKT cells were reduced in the spleen, and signs of autoimmunity were evident. Thus, Klf6 has a pro- survival role in TEC and is also required for differentiation of the mTEC I and mTEC IV populations of TEC in adult mice.
In this work, we report that mice with Foxn1-Cre mediated ablation of Klf6 in TEC demonstrate thymic hypoplasia beginning from prenatal life and extending through adulthood. Guided by single-cell transcriptional profiling, we determined that loss of Klf6 increased programmed cell death of TEC in prenatal and adult mice. In adult mice, thymic Klf6 deficiency severely impacted the mTEC I and mTEC IV populations. In addition, Klf6 deficiency led to the expansion of a previously uncharacterized cTEC population expressing Sox10 that is present in wild-type mice at very low frequencies. We observed concordant reductions of the naïve αβ T cell and iNKT pools in the periphery of young adult mice. Furthermore, we detected T cell infiltration in salivary and lacrimal glands, indicating defects in T cell tolerance.

overall_design: scATAC-seq of Adult_KO_TEC and Adult_WT_TEC

first: Justin

last: Malin

first: Gustavo

middle: U

last: Martinez Ruiz

first: Avinash

last: Bhandoola


title: LHP1-mediated conditional repression of subgenome-diversified defenses is responsible for the high plasticity of allopolyploid wheat

city: shanghai

first: yijing

last: zhang

address: 2005 Songhu Road

department: Biochemistry

institute: Fudan University

summary: High plasticity of common wheat is attributed to the captured and polyploidization-promoted diversity. However, uncontrolled subgenome diversification can lead to hybrid conflict and dysgenesis, resulting in decreased diversity. How genomic diversity is maintained and interpreted to increase plasticity is unclear. By data-mining from the binding of 193 genome-wide trans-factors and genetic perturbations in common wheat, we identified LHP1 as a major regulator of subgenome-diversified defense genes, enhancer RNAs, and metabolite synthesis-related gene clusters via H3K27me3. Stripe rust infection leads to a global decrease in LHP1-mediated H3K27me3, deprivation of which enhances common wheat stripe rust resistance. We also revealed the consistency between subgenome diversity and population diversity, potentially promoted by LHP1, implying the recent diversification preferentially occurred in the captured subgenome-diversified regions regulated by LHP1. Thus, common wheat benefitted from multi-faced role of LHP1 in promoting sequence diversity and repressing subgenome-diversified defenses; this constraint is eliminated by pathogen infections, enabling timely release and fixation of favorable variations, conferring the evolutionary advantage and high plasticity of common wheat.

overall_design: To elucidate how genomic diversity is maintained and interpreted to increase plasticity in common wheat, we obtained gene expression profiles, H3K27me3, LHP1 and eRNA  in wild type (JW1) and lhp1-abd mutant.

first: Yijing

last: Zhang


title: Subgenome-specific usage of transposable element-derived promoters and enhancers in bread wheat development

type: Methylation profiling by high throughput sequencing

summary: Transcription start site (TSS) is the hub integrating regulatory input from promoters and enhancers. Common wheat converged three subgenomes adapted to different environment and established as a major crop worldwide. Detection of more precise and new TSS of both genes and enhancers in common wheat is the basis in interpreting transcriptional regulatory networks as well as the subgenome divergent regulation in their entirety.

sample_organism:
Triticum aestivum
Triticum urartu

platform_organism:
Triticum aestivum
Triticum urartu

overall_design: In the present study, we profiled the genome wide TSS by CAGE-seq together with DNA methylation and histone modifications from four typical tissues in common wheat, the resulting catalogue of enhancers enabled more precise detection of ubiquitous and tissue specific TSS usage of coding genes and enhancers.


title: Differentially expressed genes after transfection of miRNA in a human cancer cell line (miR-139-5p and miR-139-3p)

status: Public on Oct 11 2023

city: Kagoshima

first: Yuya

last: Tomioka

institute: Kagoshima University

summary: To identify differentially expressed genes by miRNAs transfection in human cancer, lung adenocarcinoma cells were subjected to agilent whole genome microarrays

overall_design: Human lung adenocarcinoma cells (A549) were treated with miRNAs (miR-139-5p and miR-139-3p)

first: Naohiko

last: Seki


title: Sis2 regulates yeast replicative lifespan in a dose-dependent manner

city: Illkirch-Graffenstaden

first: David

last: Moreno Fortuno

state: Bas Rhin

address: 1 Rue Laurent Fries

country: France

institute: INSTITUT DE GÉNÉTIQUE ET DE BIOLOGIE MOLÉCULAIRE ET CELLULAIRE

summary: Application of microfluidic platforms facilitated high-precision measurements of yeast replicative lifespan (RLS); however, comparative quantification of lifespan across strain libraries has been missing. Here we microfluidically measured RLS of 307 yeast strains, each deleted for a single gene. Despite previous reports of extended lifespan in these strains, we found that 56% of them did not actually live longer than the wild-type; while the remaining 44% showed extended lifespans, the degree of extension was often different from what was previously reported. Deletion of SIS2 gene led to the largest RLS increase observed. Sis2 regulated yeast lifespan in a dose-dependent manner, implying a role for the coenzyme A biosynthesis pathway in lifespan regulation. Introduction of the human PPCDC gene in the sis2Δ background neutralized the lifespan extension. RNA-seq experiments revealed transcriptional increases in cell-cycle machinery components in sis2Δ background. High-precision lifespan measurement will be essential to elucidate the gene network governing lifespan.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for BY4741 wild type yeast cells and its derivative sis2Δ mutant, with two biological replicate per condition.

middle: F

last: Moreno

first: Tolga

last: Olmez

first: Ping

last: Liu

first: Zane

last: Johnson

first: Madeline

last: McGinnis

first: Benjamin

last: Tu

first: Mark

last: Hochstrasser

first: Murat

last: Acar


title: Synthetic Augmentation of Bilirubin Metabolism in Human Pluripotent Stem Cell-derived Liver Organoids

city: Cincinnati

first: Hasan Al

last: Reza

address: 3333 Burnet Ave

department: Developmental Biology

institute: CCHMC

summary: To investigate the efect of perturbation of glucocorticoid signaling on the difference between the biliurbin metabolism in HLOs after treatment with drugs.

overall_design: We performed RNAseq on control HLOs compared to HLOs treated with Mifepristone (1uM) after treatment with bilirubin (10 mg/L).

first: Reza

last: Hasan Al

first: Takebe

last: Takanori


title: Klf6 is required in thymic epithelial cells for normal thymus development [scRNA-seq]

first: Yongge

last: Zhao

overall_design: scRNA-seq of Adult_KO_TEC, Adult_WT_TEC, E15_KO_TEC, E15_WT_TEC.

first: Justin

last: Malin

first: Gustavo

last: Martinez Ruiz

last: Bhandoola


title: High neighborhood deprivation impacts DNA methylation and gene expression in cancer-related genes

first: Stefan

last: Ambs

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

overall_design: Refer to individual Series


title: The three-dimensional structure of the EBV genome plays a crucial role in regulating viral gene expression in EBVaGC [HiChIP]

city: Philadelphia

first: Italo

last: Tempera

address: 3601 Spruce Street

institute: The Wistar Institute

summary: Epstein-Barr virus (EBV) establishes lifelong asymptomatic infection by replication of its chromatinized episomes with the host genome. EBV exhibits different latency-associated transcriptional repertoires, each with distinct three-dimensional structures. CTCF, Cohesin and PARP1 are involved in maintaining viral latency and establishing episome architecture. Epstein-Barr virus-associated gastric cancer (EBVaGC) represents 1.3% to 30.9% of all gastric cancers globally. EBV-positive gastric cancers exhibit an intermediate viral transcription profile known as "Latency II", expressing specific viral genes and noncoding RNAs. In this study, we investigated the impact of PARP1 inhibition on CTCF/Cohesin binding in Type II latency. We observed destabilization of the binding of both factors, leading to a disrupted three-dimensional architecture of the episomes and an altered viral gene expression. Despite sharing the same CTCF binding profile, Type I, II, and III latencies exhibit different 3D structures that correlate with variations in viral gene expression. Additionally, our analysis of H3K27ac-enriched interactions revealed differences between Type II latency episomes and a link to cellular transformation through docking of the EBV genome at specific sites of the Human genome, thus promoting oncogene expression. Overall, this work provides insights into the role of PARP1 in maintaining active latency and novel mechanisms of EBV-induced cellular transformation.

sample_organism:
Homo sapiens
human gammaherpesvirus 4

platform_organism:
Homo sapiens
human gammaherpesvirus 4

overall_design: HiChIP sequencing of Human and EBV genomes in YCCEL1 and SNU719 cell lines using H3K27ac antibody

first: Davide

last: Maestri

first: Giorgia

last: Napoletani

first: Andrew

last: Kossenkov

first: Sarah

last: Preston-Alp

first: Lisa

last: Caruso


title: The three-dimensional structure of the EBV genome plays a crucial role in regulating viral gene expression in EBVaGC [HiC]

overall_design: HiC sequencing of EBV genome in YCCEL1 and SNU719 cell lines following Olaparib treatment


title: The three-dimensional structure of the EBV genome plays a crucial role in regulating viral gene expression in EBVaGC [ChIP-seq]

overall_design: Chromatin Immunoprecipitation DNA-sequencing of histones H3K4me1 and H3K27ac


title: The three-dimensional structure of the EBV genome plays a crucial role in regulating viral gene expression in EBVaGC


title: CDK4 phosphorylation status and rational use for combining CDK4/6 and BRAF/MEK inhibition in advanced thyroid carcinomas

status: Public on Oct 10 2023

city: Brussels

first: Pierre

last: Roger

address: 808 Route de Lennik, Campus Erasme Bat.C - C1.132

country: Belgium

institute: IRIBHM, Université Libre de Bruxelles

summary: Despite overall good prognosis associated to thyroid cancer, poorly differentiated carcinomas (PDTC) and anaplastic carcinomas (ATC) represent major clinical challenges. We have shown that the presence of active T172-phosphorylated CDK4 predicts sensitivity to CDK4/6 inhibitory drugs (CDK4/6i) including palbociclib. Here, CDK4 phosphorylation was detected in all well-differentiated thyoid carcinomas (n=29), 19/20 PDTC, 16/23 ATC, and 18/21 thyroid cancer cell lines including 11 ATC-derived ones. The cell lines lacking CDK4 phosphorylation were insensitive to CDK4/6i. RNA-sequencing and immunohistochemistry revealed that tumors and cell lines without phosphorylated CDK4 presented very high p16CDKN2A levels that were associated with proliferative activity. One of the main mechanisms of resistance to CDK4/6i is RB1 defects or inactivation. RB1 mutations were present in the 3 insensitive cell lines but were not found in 5 of the 7 tumors without phosphorylated CDK4. p16/KI67 immunohistochemistry and a previously developed 11-gene signature identified the likely insensitive tumors and cell lines lacking CDK4 phosphorylation. In cell lines, palbociclib synergized with dabrafenib/trametinib, completely and irreversibly arresting proliferation. The combined drugs prevented resistance mechanisms induced by palbociclib, most notably Cyclin E1-CDK2 activation and a paradoxical stabilization of phosphorylated CDK4 complexes. Our study supports the evaluation of CDK4/6i for ATC/PDTC treatment, including in combination with MEK/BRAF inhibitors.

overall_design: 16 thyroid cancer cell lines in basal conditions were profilled by RNA-seq.

first: Jaime

last: Pita

first: Eric

last: Raspé

first: Frederick

last: Libert

first: Carine

last: Maenhaut


title: Klf6 is required in thymic epithelial cells for normal thymus development


title: AhR promotes dedifferentiation in MYCN amplified neuroblastoma

city: Buffalo

first: Mark

middle: D

last: Long

address: Elm & Carlton Streets

department: Bioinformatics & Biostatistics

institute: Roswell Park Comprehensive Cancer Center


title: Trim-away depletion of Zona Pellucida 3 (ZP3) protein in mouse zygotes (analysis after 24h)

first: Michele

last: Boiani

summary: Mutation studies always defined the functions of the zona pellucida (ZP) as extracellular, namely: to encase the oocytes in ovarian follicles, to ensure species-specific sperm binding, and to dampen shear stress on the embryo surface. Therefore, mutations in the three ZP mouse genes ZP1, ZP2 or ZP3 cause primary infertility due to empty follicles, polyspermic fertilization or harmful contact between embryos and oviductal epithelium. However, the concepti of ZP2-null and ZP3-null oocytes were still unviable also when the defects were obviated by monospermic fertilization in vitro and blastocyst transfer to uterus (PMID 11245577). This suggests that the tasks of ZPs don’t end in the extracellular space as previously assumed, but there may be also intracellular functions yet to be discovered. The present study tested if experimentally induced degradation of intracellular ZP3 impacted on the development and transcriptome of mouse embryos. To this end we degraded ZP3 using its antibody in conjunction with the ubiquitin-protein ligase TRIM21. This method is known as 'Trim-away' (PMID 29153837). Briefly, in this method a cell (e.g. oocyte) expressing TRIM21 is supplied e.g. injected with a specific antibody to a protein of interest, in this case ZP3. As a result, the ternary complex (target protein-antibody-TRIM21) is destroyed in the proteasome. TRIM21 is here always to be understood as translation product of microinjected mCherry-Trim21 mRNA. We compared two experimental groups, as follows. Pronuclear-stage oocytes (B6C3F1 x CD1) were microinjected with approx. 100 picoliters of mix comprised of mCherry-Trim21 mRNA 0.2 mg/mL + buffer of ZP3 antibody + dextran beads 0.02 mg/mL as tracer, forming a group named 'Trim21 overexpression', in quadruplicate. As a reference, pronuclear-stage oocytes were microinjected with mCherry-Trim21 mRNA 0.2 mg/mL + anti-ZP3 antibody (Proteintech 21279-1-AP) 1 mg/mL + dextran beads, forming a group named 'Trim-away ZP3' group, in triplicate. To identify differently expressed genes we compared group 'Trim-away ZP3' with group 'Trim21 overexpression'. In addition, a single 'non-manipulated' sample was also incuded merely to confirm that Trim21 was detectable in the two experimental groups, but not in the non-microinjected embryos. On the day after microinjection, embryos were collected and lysed for transcriptome analysis. Those of  group 'Trim21 overexpression' were at the 2-cell stage so as the non-manipulated embryos, whereas  those of group 'Trim-away ZP3' were arrested at the 1-cell stage. Transcriptome analysis revealed that embryos of group 'Trim-away ZP3' and 'group 'Trim21 overexpression' differed in the expression of 197 of 11137 genes  (t test, FDR<0.05). The data support a conclusion that ZP3 found inside the embryo was not merely a remnant from oogenesis, but served an intracellular, post-fertilization role during mouse preimplantation development.

overall_design: Biological replicates were analyzed for each of two experimental conditions: 'Trim-away of ZP3' (protein degradation) in triplicate (r1, r2, r3) and 'Trim21 overexpression' (no protein degradation) in quadruplicate (r1, r2, r3, r4). Trim-away followed to the coinjection of antibody anti ZP3 (Proteintech 21279-1-AP; 1 mg/mL), mCherry-Trim21 mRNA (0.2 mg/mL) and oregon green dextran beads (0.02 mg/mL), resulting in protein degradation in the proteasome. 'Trim21 overexpression'  followed to the coinjection of mCherry-Trim21 mRNA (0.2 mg/mL), antibody buffer (but no antibody) and oregon green dextran beads (0.02 mg/mL), resulting in no protein degradation. RNA was extracted and analyzed at the late chronological 2-cell stage, when the embryos overexpressing Trim21 were at the 2-cell stage whereas those receiving mCherry-Trim21 and anti-ZP3 antibody were arrested at the 1-cell stage.

first: Steffen

last: Israel


title: Circulating extracellular vesicle-derived microRNAs as novel diagnostic and prognostic biomarkers for non-viral-related hepatocellular carcinoma

city: Bangkok

first: Bootsakorn

last: Boonkaew

state: Bangkok

address: 1873 Rama IV

country: Thailand

institute: Chulalongkorn University

summary: The potential significance of plasma extracellular vesicle-derived miRNAs in non-hepatitis B-, non-hepatitis C-related hepatocellular carcinoma as biomarker for the diseases was explored.

overall_design: In this study, we compared plasma extracellular vesicle-derived miRNA profiles between non-hepatitis B-, non-hepatitis C-related hepatocellular carcinoma (n=9) and control groups including non-alcoholic fatty liver disease (n=9) and healthy controls (n=9) using NanoString method. The differentially expressed extracellular vesicle-derived miRNA were validated in another set of plasma samples by qRT-PCR.

first: Nantawat

middle: N

last: Satthawiwat

first: Nutcha

last: Pinjaroen

first: Natthaya

last: Chuaypen

first: Pisit

last: Tangkijvanich


title: Transcriptional legacy in the armadillo

city: Kensington

first: Sara

last: Ballouz

state: NSW

address: High St

country: Australia

institute: UNSW

summary: Longitudinal study of armadillo transcription.

overall_design: We performed RNA-seq on PBMCs of five sets of quadruplets of the nine-banded armadillo across three time points.

first: Risa

last: Kawaguchi

first: Maria

middle: T

last: Pena

first: Stephan

last: Fischer

first: Megan

last: Crow

first: Leon

last: French

first: Frank

last: Knight

first: Linda

middle: B

last: Adams

first: Jesse

last: Gillis


title: FoxP3 Pulldown-seq (Nucleosomal DNA)

status: Public on Oct 09 2023

city: Boston

first: Wenxiang

last: Zhang

address: 300 Longwood Ave

institute: Boston Childrens Hospital

summary: To investigate the interactions between FoxP3 and chromatinized templates, particularly assessing whether FoxP3 can still bind significantly to TnG-repeat DNA, we conducted FoxP3 pulldown experiments using nucleosomal DNA. Consitent with the result using naked genomic DNA, the results highlighted TnG repeats as one of the most significant motifs in these interactions.

overall_design: We conducted FoxP3 pulldown-seq using nucleosomal DNA isolated from EL4 cells. Subsequently, we performed motif analysis using the enriched peaks to assess the DNA binding pattern of FoxP3 on nucleosomal DNA.

first: Sun

last: Hur

first: Xi

last: Wang


title: Ladder-like multimerization of FoxP3 enables microsatellite recognition and DNA bridging


title: FoxP3 mRNA-seq in mouse CD4+T cells

summary: To investigate the impact of FoxP3 multimerization mutants on FoxP3's genome-wide gene expression profiles, we conducted FoxP3 RNA-seq experiments using mouse CD4+ T cells. The results indicated that all of these multimerization mutants had an effect on FoxP3 gene expression profiles, including some well-established Treg signatures.

overall_design: mRNA-seq on Wild-type FoxP3 overexpressed in mouse CD4+ T cells, alongside an empty vector control and four distinct multimerization mutants. Each sample was quadruplicated for robustness.


title: Differentially expressed genes in SCC13 NRF3-KO cells and SCC13 cells that re-express endogenous concentrations of recombinant NRF3

city: Zürich

first: Selina

last: Gurri

address: Otto-Stern-Weg 7

department: Institute of Molecular Health Sciences

country: Switzerland

institute: ETH Zürich

summary: We identified a tumor-suppressing function of NRF3 in skin cancer. To determine the underlying mechanism of action we wanted to compare the transcriptome of SCC13 NRF3-KO cells and SCC13 cells that re-express endogenous concentrations of recombinant NRF3.

overall_design: First Crispr/Cas9 KO of NRF3 was performed in SCC13 cells whereafter the cells were transduced with a pInducer20 contrusct containing the NRF3 cDNA, which allows doxycycline-inducible re-expression of NRF3. The cells were treated with DMSO (vehicle) or 20 ng/ml Dox to allow for endogenous expression of NRF3 and translocation to the nucleus. Differentially expressed genes were identified using RNA-seq.

first: Sabine

last: Werner


title: AhR promotes dedifferentiation in MYCN amplified neuroblastoma [RNA-seq]

first: Mark

last: Long

overall_design:
Neuroblastoma cell lines (Kelly, BE2C) were subject to transcriptome profiling by RNA-seq
RNAseq analysis of Neuroblastoma cell lines +/- shAhR knockdown

first: Anna

last: Bianchi-Smiraglia


title: RNA sequence (RNA-seq) profiling of SynVIII strain

city: New York

first: Jitong

last: Cai

address: 435 E 30th street

institute: NYU Langone Health

summary: Building synthetic yeast chromosomes with thousands of designer modifications empowers probing of genome flexibility on an unprecedented scale. Here, we describe the assembly and characterization of synthetic chromosome VIII (synVIII), which served as a platform for further genome manipulation of an essential functional element, the centromere. Recent findings suggest that centromeres may have distinct characteristics and thus relocating the centromere to various positions along a single chromosome (VIII) enables us to directly test the suitability of several ectopic destinations for centromere function and stability. We used a novel CRISPR/Cas9-mediated approach, Single-Step Intrachromosomal Centromere Transplantation (SSICT), to relocate the 118 bp point centromere sequence of CEN8. Successful transplantation of CEN8 invariably resulted in chromosome VIII aneuploidy regardless of ectopic centromere position in wild-type or synVIII strains and euploid structural variants hinted that including left-flanking pericentromeric sequences of CEN8 allowed ectopic function. Plasmid loss rate assays revealed that specific left-flanking sequences increased stability of a CEN8-containing circular minichromosome and reduced incidence of aneuploidy during SSICT. Together, these results show that the minimal point centromere of budding yeast is insufficient to confer chromosome stability when transplanted ectopically, and is rescued by left-flanking pericentromeric sequence. This is an important consideration for future genome engineering projects involving transplantation of functional elements.

overall_design: We perform RNA-sequencing to assess the gene expression changes in the synVIII strain ySLL185 against wild-type strain BY4741.

first: Stephanie

last: Lauer

first: Jef

last: Boeke


title: The three-dimensional structure of the EBV genome plays a crucial role in regulating viral gene expression in EBVaGC [RNA-seq]

first: Italo

last: Tempera

overall_design: RNA sequencing of Human and EBV transcriptome in YCCEL1 and SNU719 cell lines

first: Davide

last: Maestri

first: Giorgia

last: Napoletani

first: Andrew

last: Kossenkov

first: Sarah

last: Preston-Alp

first: Lisa

last: Caruso


title: ChIP-Seq of SA1, SA2, and Rad21 in RPE-1 and CHM13 cells

city: Kansas City

first: Chris

last: Seidel

state: MO

department: Genomics

institute: Stowers Institute

summary: The biorientation of sister chromatids on the mitotic spindle, essential for accurate sister chromatid segregation, relies on critical centromere components including cohesin, the centromere-specific H3 variant CENP-A, and centromeric DNA. Centromeric DNA is highly variable between chromosomes yet must accomplish a similar function. Moreover, how the 50 nm cohesin ring, proposed to encircle sister chromatids, accommodates inter-sister centromeric distances of hundreds of nanometers on the metaphase spindle is a conundrum. Insight into the 3D organization of centromere components would help resolve how centromeres function on the mitotic spindle. We used ChIP-seq and super-resolution microscopy to examine the geometry of essential centromeric components on human chromosomes. ChIP-seq of SA1, SA2, and Rad21 in human cells demonstrates that cohesin subunits are depleted in -satellite arrays where CENP-A nucleosomes and kinetochores assemble. Cohesin is instead enriched at pericentromeric DNA. Structured illumination microscopy of sister centromeres is consistent, revealing a non-overlapping pattern of CENP-A and cohesin.

overall_design: ChIP-Seq of Cohesin subunits SA1, SA2, and Rad21 in human cell lines RPE-1 in duplicate, and CHM13 in triplicate.

first: Ayantika

last: Sen Gupta

first: Jennifer

last: Gerton


title: Dual roles of the Arabidopsis PEAT complex in histone H2A deubiquitination and H4K5 acetylation

status: Public on Oct 08 2023

city: Beijing

first: xinjian

last: He

address: No. 7, zhongguancun life science park road, changping district

institute: National Institute of Biological Sciences


title: Profiling stress-triggered RNA condensation with photocatalytic proximity labeling

first: Ziqi

last: Ren

address: Chengfu Road No.292

department: College of Chemistry and Molecular Engineering

institute: Peking University

summary: Stress granules (SGs) are highly dynamic cytoplasmic membrane-less organelles that assemble when cells are challenged by stress. At different stages of assembly, various RNA molecules are sorted into SGs where they play important roles in maintaining the structural stability of SGs and regulating gene expression. Despite recent efforts of fluorescence microscopy and biochemical fractionation analysis, there still lacks a complete description of the dynamic changes in the molecular inventory of SG components during different stages of assembly and disassembly. In this study, we applied a proximity-dependent RNA labeling method, CAP-seq, to comprehensively investigate the content of local transcriptome in SGs, in the context of live mammalian cells. CAP-seq captures 457 and 822 RNAs in arsenite- and sorbitol-induced SGs in HEK293T cells, respectively, revealing that SG enrichment is positively correlated with RNA length and AU content, but negatively correlated with translation efficiency. The high spatial specificity of CAP-seq dataset is validated by single-molecule FISH imaging. We further applied CAP-seq profile RNA components in microscopically invisible SG cores, both before stress and after recovery from stress, thus mapping the dynamic changes in SG-proximal transcriptome along the time course of granule assembly/disassembly processes. Our data portray a model of AU-rich and translationally repressed SG nanostructure that are memorized long after the removal of stress.

overall_design: Expression profiling by next-generation sequencing form SGs CAP-seq libraries from different treatment including arseite stress, sorbitol stress, basal condition, and 1 hour or 3 hours recovery post arsenite stress.

first: Wei

last: Tang

first: Luxin

last: Peng

first: Peng

last: Zou


title: RNA-sequencing of differentiated mouse skeletal muscle cells following the depletion of EYA3

status: Public on Oct 06 2023

city: Chapel Hill

first: Jimena

last: Giudice

state: NC

address: 6340B MBRB, Campus Box 7545

department: Cell Biology and Physiology

institute: UNC - Chapel Hill

summary: Mouse skeletal muscle cells were subjected to treatment with siRNAs targeting EYA3 or a non-targeting siRNA (si-Ctrl). Differential gene expression analysis was performed on the resulting data.

overall_design: 9 samples total, 3 replicates each with three conditions (si-Ctrl, si-Eya3#1, and si-Eya3#2)

first: Hannah

last: Wiedner


title: AhR promotes dedifferentiation in MYCN amplified neuroblastoma [ATAC-seq]

first: Mark

last: Long

summary:
Neuroblastoma cell lines (Kelly, BE2C) were subject to chromatin profiling by ATAC-seq
ATAC-seq analysis of Neuroblastoma cell lines +/- shAhR knockdown

overall_design: Anna, Bianchi-Smiraglia


title: Identifying the potential  targets of METTL3 in macrophages by high-throughput m6A-seq

city: Shanghai

first: Xiao

last: Han

state: Select state...

address: 399 Wanyuan Rd, Minhang, Shanghai, China

institute: Children’s Hospital of Fudan University

summary: N6-methyladenosine (m6A), the most prevalent internal messenger RNA modification, plays critical roles in diverse biological processes. To characterize the potential involvement of METTL3 in macrophage function,  we mapped m6A methylomes of METTL3-deficent and WT macrophages by m6A sequencing (m6A-seq) and RNA-seq, with independent biological replicates. Notably, among the  m6A peaks with remarkable changes upon METTL3-depletion, the vast majority exhibited a significant (p < 0.05) decrease in m6A abundance.The most common m6A motif GGAC is significantly enriched in the m6A peaks in both WT and METTL3-deficent cells. A similar pattern of total and common m6A distribution in control and METTL3-deficient cells was observed, when the m6A peaks were especially abundant in the vicinity of CDS and 3’UTR regions.

overall_design: The m6A profiles of  WT and METTL3-depleted macrophages under IL-4 stimulation. Two or three  independent experiments were performed at  control siRNA or METTL3 siRNA treatment, respectively.

first: Wenfeng

last: Xiao


title: FoxP3 ChIP-seq in mouse CD4+T cells

first: Wenxiang

last: Zhang

state: Massachusetts

summary: To investigate the impact of FoxP3 multimerization mutants on FoxP3's genome-wide DNA binding, with a particular focus on TnG-repeats binding, we conducted FoxP3 ChIP-seq experiments using mouse CD4+ T cells. The results revealed that all of these multimerization mutants have the potential to influence FoxP3 DNA binding, including its interaction with TnG-repeats.

overall_design: We performed ChIP-seq experiments on Wild-type HA-tagged FoxP3 overexpressed in mouse CD4+ T cells, along with an empty vector control and four distinct multimerization mutants.

first: Sun

last: Hur


title: MBP-FoxP3 pulldown-seq on naked DNA

summary: To re-evaluate FoxP3 sequence specificity, we performed an unbiased pull-down of genomic DNA with recombinant FoxP3 protein. Use of genomic DNA, as opposed to synthetic DNA oligos, enables testing sequence specificity in the context of naturally existing repertoire of sequences. De novo motif analysis showed a strong enrichment of TnG repeats (n=2-5) by FoxP3 pull-down, using either MBP alone pull-down or input as a control.

overall_design: We isolated genomic DNA from mouse EL4 cells, fragmented it to approximately 100-200 bp in size, and then incubated it with purified MBP-tagged mouse FoxP3. Subsequently, we conducted an MBP pull-down, followed by next-generation sequencing (NGS) of the co-purified DNA, a process referred to as FoxP3 PD-seq. We utilized the enriched peaks from this dataset to perform motif analysis, shedding light on the DNA binding patterns of FoxP3 on naked genomic DNA.


title: Prevalence of chromosomal alterations in first-trimester spontaneous pregnancy loss

status: Public on Oct 05 2023

city: Maastricht

first: Rick

last: Essers

state: Limburg

address: P. Debyelaan 25

department: Clinical Genetics

institute: Maastricht University Medical Centre (MUMC+)

summary:
Extensive genome analysis of pregnancy loss products by genome haplarithmisis.
Pregnancy loss (PL) is the primary pregnancy complication, mostly caused by chromosomal abnormalities of the conceptus. However, the nature and prevalence of these abnormalities and the allocation of (ab)normal cells in embryonic and placental compartments during intrauterine development remains elusive. We analyzed 1,745 spontaneous PLs and found that ~50% were karyotypically normal. We applied genome haplarithmisis to 91 PL families with normal karyotypes, following whole-genome genotypes of the parents as well as of the extraembryonic mesoderm (EM) and chorionic villi (CV), representing embryonic and placental cells, of the product of conception (POC), which allowed characterizing genomic landscape of both lineages. 36.4% of these PLs have chromosomal aberrations not previously detected by karyotyping. In contrast to viable pregnancies where mosaic chromosomal abnormalities are often restricted to the CV, we find that in spontaneous abortions, the situation is reversed with a higher degree of mosaic chromosomal imbalances in EM rather than CV.

overall_design: From 1987 to 2021, A total of 1745 spontaneous PLs were analyzed using karyotyping. 866 (49.6%) were classified as normal and 879 (50.4%) as abnormal. The karyotypes of fetal tissue was determined by conventional metaphase analysis (n=1745) or additional testing by comparative genomic hybridization (CGH) and fluorescence in situ hybridization (FISH), as described previously37,38. All the samples with abnormal karyotype were excluded for downstream haplarithmisis. Of the karyotypically normal cases, 111 families were randomly selected for SNP haplotyping, given that fetal (EM and CV) and parental blood samples were available without previously identified genetic predisposition was in any of the couples. Twenty families have been excluded due to the DNA of one or more family members being of insufficient quality causing low SNP call rates or due to one or both parents not being the biological parent, making proper haplotyping analysis impossible . Ninety-one families have been successfully analyzed by haplarithmisis. Of those, forty-two were categorized as SPL (loss of one pregnancy) and forty-nine as RPL (loss of two or more consecutive pregnancies).

first: Igor

last: Lebedev

first: Ants

last: Kurg

first: Elizaveta

middle: A

last: Fonova

first: Servi

last: Stevens

first: Rebekka

last: Koeck

first: Ulrike

last: von Rango

first: Lloyd

last: Brandts

first: Spyridon

last: Deligiannis

first: Tatyana

middle: V

last: Nikitina

first: Elena

last: Sazhenova

first: Ekatarina

last: Tolmacheva

first: Anna

last: Kashevarova

first: Dmitry

last: Fedotov

first: Viktoria

last: Demeneva

first: Gleb

last: Drozdov

first: Salwan

last: Al-Nasiry

first: Merryn

last: Macville

first: Arthur

last: van den Wijngaard

first: Jos

last: Dreesen

first: Aimee

last: Paulussen

first: Alexander

last: Hoschen

first: Han

last: Brunner

first: Andres

last: Salumets

first: Masoud

last: Esteki


title: Combination with ERR gamma agonist and mechanical stretching enhances maturation of 3D cardiac tissue and manifests the hypertrophic cardiomyopathy phenotype

city: Fujisawa City

first: Yuya

last: Fujiwara

state: Kanagawa Prefecture

address: 2-26-1 Muraokahigashi

institute: Kyoto University

summary: Here, we established a method for maturating engineered cardiac tissue and creating manifested its HCM model using  human induced prulipotent stem cells dericed cardiomyocytes and its isogenic cells.

overall_design: RNA-seq data for ECTs treated with static condition under DMSO treatment,  static condition under T112 treatment, mechanical stimulation under DMSO tratment and mechanical stimulation under T112 to assess maturation. And RNA-seq data for ECTs treated mechanical stimulation under T112 harbouring WT, MYH7 R719Q or MYBPC3 G115*.

first: Masako

last: Sasaki

first: Megumi

last: Kumazaki

first: Yoshinori

last: Yoshida


title: Differentially expressed genes in wt and Nrf3-ko mouse keratinocytes before and after 24 h 100 mJ/cm2  UVB irradiation

first: Selina

last: Gurri

summary: We identified a pro-apoptotic function of Nrf3 in keratinocytes after UVB irradiation. To determine the underlying mechanism of action we wanted to compare the transcriptome of wt and Nrf3-ko mouse keratinocytes before and after 24 h 100 mJ/cm2  UVB irradiation

overall_design: Wt and Nrf3-ko mice were mock-treated or UVB-irradiated and sacrificed 24 h later. Differentially expressed genes in the epidermis were identified using RNA-seq.

first: Beat

last: Siegenthaler

first: Sabine

last: Werner


title: Transcriptome of murine aorta

first: Yongting

last: luo

state: Beijing

address: No.10 Tianxiu Road

institute: Beijing Advanced Innovation Center for Food Nutrition and Human Health

summary: This experiment aims to explore the transcriptome of aorta from mice treated without or with coQ10, TPA, or T-5224.

overall_design: aortic tissue transcirptome prolifes from 9 mouse aorta

last: Luo

first: Junjie

first: Peng

last: An

first: Fazheng

last: Ren