title: Induction of transient and adaptive vs maladaptive unfolded protein responses of the endoplasmic reticulum in pancreatic beta cells

city: Indianapolis

last: Kalwat

address: 1210 Waterway Blvd Suite 2000

department: Lilly Diabetes Center of Excellence

institute: Indiana Biosciences Research Institute

summary: Whole RNA-sequencing of MIN6 cell samples from a drug-treatment time course to investigate how hypersecretion vs. ER calcium disruption influence stress and function adaptation.

overall_design: Three independent experiments were peformed in MIN6 beta cells. Each experiment contained 13 samples: A 0h baseline and 1,2,6,24 h for DMSO (0.1%), SW016789 (5uM), and thapsigargin (100nM) as an ER stress control treatment. RNA was collected from all samples at the end of the time course.


title: Analysis of gene regulation by the TP63 isoforms deltaNp63alpha and deltaNp63beta using ChIPseq of p63 and H3K27ac

status: Public on Dec 13 2024

city: Albany

first: Morgan

middle: A

last: Sammons

state: NY

address: 1400 Washington Ave, Life Sciences 2075

department: Biological Sciences

country: USA

institute: University at Albany - SUNY

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Our goal was to assess transcriptional changes of the colon carcinoma cell line HCT116 TP53-/- in response to expression of the TP63 isoforms deltaNp63alpha or deltaNp63beta. The goal was to understand how these C-terminal isoforms of p63 might differentially regulate gene expression. We also assessed how  differential binding of these isoforms and how they affect  local changes in H3K27ac.

overall_design: ChIP-seq of 24hr induced (+dox) conditions for either a negative control, deltaNp63alpha, or deltaNp63beta HCT116 TP53-/- cell lines using an antibody targeting the deltaN-domain of p63.  ChIP-seq of 24hr induced (+dox) conditions for either a negative control, deltaNp63alpha, or deltaNp63beta HCT116 TP53-/- cell lines using an antibody targeting H3K27ac (histone H3 lysine 27 acetylation)

first: McCann

last: Abby

first: Sammons

last: Morgan


title: CTCF-mediated 3D chromatin predetermines the gene expression program in the male germline (Hi-C)

city: Davis

first: Satoshi

middle: H

last: Namekawa

state: CA

address: One Shields Avenue

department: Department of Molecular and Cellular Biology

institute: University of California, Davis

summary: Spermatogenesis is a unidirectional differentiation process to produce haploid sperm. However, it remains largely unknown how the gene expression program is determined to direct a unidirectional differentiation. Here we unveil the high-resolution 3D chromatin architecture of male germ cells and show that CTCF-mediated 3D chromatin predetermines the gene expression program required for spermatogenesis. At the mitosis-to-meiosis transition, the germline-specific Polycomb protein SCML2 resolves chromatin loops specific to mitotic spermatogonia. On autosomes, CTCF-mediated 3D chromatin manifests the structural feature of meiosis-specific super-enhancer (meiotic SE) loci already in undifferentiated spermatogonia. In meiotic spermatocytes, the master transcription factor A-MYB is recruited to these meiotic SE loci to strengthen their 3D contacts to instruct the burst of meiotic gene expression. Further, SCML2 and A-MYB establish unique 3D chromatin of the sex chromosomes in meiotic sex chromosome inactivation. We propose that CTCF-mediated 3D chromatin underlines epigenetic priming to direct unidirectional differentiation, thereby determining the cellular identity of the male germline.

overall_design: Determination of 3D chromatin structures of THY1+ and KIT+ spermatogonia, pachytene spermatocytes and round spermatids by Hi-C analyses.

first: Yuka

last: Kitamura

first: Kazuki

last: Takahashi

first: So

last: Maezawa

first: Yasuhisa

last: Munakata

first: Akihiko

last: Sakashita

first: Noam

last: Kaplan


title: Giardiavirus rewires host translation and glycolytic metabolism to support its replication in Giardia duodenalis

status: Public on Dec 31 2024

city: changchun

first: lu

last: li

country: China

institute: Jilin university

type: Expression profiling by high throughput sequencing

summary: Giardia duodenalis is a prevalent intestinal pathogens known to cause giardiasis, a condition characterized by diarrhea and frequently linked to malnutrition and growth impairments in children. The presence of Giardiavirus (GLV) in Giardia strains has been associated with heightened immune cytokine responses in the host compared to the GLV-free strains. However, the transmission mode and biological significance of GLV remain unclear. In this study, by using G. duodenalis DH (GLV-containing) and WBC6 (GLV-free) strains, we demonstrated that the DH strain produced extracellular vesicles (EVs), which originated from unique peripheral vesicle and bead-like structures in the ventrolateral flange. Nanoparticle tracking analysis revealed that GLV infected-G. duodenalis DH strain secreted fewer EVs than the GLV-free WBC6 strain. Biochemical and electron microscopy demonstrated that GLV virions can exploit the Giardia EVs pathway to facilitate their spread among parasites. Giardia uptake of GLV-containing EVs occured through clathrin-mediated endocytosis, leading to rapid infection of trophozoites by GLV. Furthermore, GLV infection enhanced messenger RNA translation efficiency, influencing protein abundance in Giardia trophozoites. The presence of GLV also upregulated the glycolytic pathway, with Giardia enolase closely associated with GLV replication. Importantly, Giardia infected with GLV alleviated the pathogenicity of parasite compared to the GLV-freeGiardia strain. These findings highlight the pivotal role of GLV in regulating Giardia biology, suggesting its potential for informing the development of novel intervention strategies against Giardia infections.

overall_design: Comparative genes translation efficiency on the three species by Monosomes RNA-seq and polysomes RNA-seq analyses

first: Lu

last: Li


title: Transcriptional regulation of the germline piRNA pathway in the Drosophila ovary [ovo KD RNA-Seq]

status: Public on Dec 23 2024

city: Cambridge

first: Azad

last: Alizada

state: Cambridgeshire

address: University of Cambridge Li Ka Shing Centre, Robinson Way

department: Cancer Research UK Cambridge Institute

country: United Kingdom

institute: University of Cambridge

summary: The gene-regulatory mechanisms behind the female germline piRNA pathway in ovaries of metazoan species remain largely unexplored. Here, using the Drosophila model and combinations of ChIP-seq, RNA-seq, ATAC-seq, and single-cell approaches, we reveal the transcription factors that regulate the female germline piRNA pathway in animal ovaries.

overall_design: Differential RNA-seq analysis between ovo siRNA-knockdowns and renilla siRNA-knockdowns in Δl(3)mbt OSCs

first: Benjamin

last: Nicholson

first: Gregory

middle: J

last: Hannon


title: Arginine Promotes The Activation of Human Lung Fibroblasts Independent of Its Metabolism

status: Public on Dec 12 2024

city: Chicago

first: Kun Woo

last: Shin

state: IL

address: 5841 S Maryland Ave

department: Medicine

institute: The University of Chicago

summary: We investigated TGF-beta-induced gene expression in normal human lung fibroblasts (NHLFs) under different media setting, and diffrenece in TGF-beta-induced gene expression between control human male fibroblasts and human male IPF fibroblasts cultured in HPLM using RNA-seq.

overall_design: Comparative gene expression profiling analysis of RNA-seq data from Normal Human Lung Fibroblasts (HLFs) under TGF-beta treatment in different media ( HPLM and DMEM ) and different source of HLFs across different clones ( Control Male and IPF Male ) cultured in HPLM.

first: Gokhan

last: Mutlu

first: Robert

last: Hamanaka

first: KunWoo

first: Rengul

last: Cetin-Atalay


title: Diverse immune suppressing cancer communications with myeloid cells prevent T cell activation and recruitment and provide resistance to cell cycle inhibitor therapy

status: Public on Dec 11 2024

city: Houston

first: Jeffrey

last: Chang

address: 6431 Fannin St

institute: University of Texas

summary: Cancer cells evolve and acquire the ability to develop resistance to therapy and change their environment. A key mechanism involves altering communication with non-malignant cells in the tumor microenvironment (TME). By corrupting the signals for growth and survival, evolving tumors engineer a pro-tumor TME which contributes to the development of treatment resistance. However, the specific interactions between malignant and non-malignant cells that predispose drug resistance and their changes during treatment remain widely unknown. Here we examined the composition, communication, and phenotypic diversity of tumor-associated cell populations in serial biopsies from the FELINE clinical trial. Early-stage ER+ breast cancers receiving endocrine therapy (letrozole) alone or in combination with the CDK4/6 cell cycle inhibitor ribociclib were analyzed using single-cell RNA sequencing (scRNAseq). Our analyses reveal cancer cells from ribociclib resistant tumors stimulate macrophage differentiation towards an immune-suppressive phenotype through upregulation of a broad diversity of cytokines and growth factors. This shift in phenotype leads to reduced macrophage cell IL-15/-18 crosstalk with CD8+ T-cell via IL-2/15RA/18R receptors, resulting in reduced T-cell activation and recruitment. Given cell cycle inhibitors known inhibitory effect on immune cell activation, in addition to also blocking cancer cell proliferation, maintenance of immune stimulatory cellular populations and signals that promote T-cell abundance and cytotoxicity is critical for treatment response. Our results show how altered communication promotes an immune-cold TME that predisposes tumors to develop CDK4/6 inhibitor resistance. A successful treatment strategy will require both the prevention of cancer division and also activation of a cytotoxic T-cell response.

overall_design:
Single-nucleus RNA sequencing of 27 ER+ breast cancer patients undergoing neoadjuvant therapy profiled at 0, 14, and 180 days.
Raw data will be submitted to the dbGaP database, due to patient privacy concerns.

first: Jason

last: Griffiths

first: Aritro

last: Nath

first: Patrick

last: Cosgrove

first: Eric

last: Medina

first: Qamar

last: Khan

first: Fred

last: Adler

middle: T

first: Andrea

last: Bild


title: Dynamics of Th1/Th17 Responses and Antimicrobial Pathways in Leprosy Skin Lesions

status: Public on Dec 21 2024

city: Los Angeles

first: Priscila

middle: Ribeiro

last: Andrade

state: CALIFORNIA

address: 615 Charles E Young Drive South, room 246

institute: UCLA

summary: Reversal reactions (RR) in leprosy provide a unique opportunity to study the dynamics of the immune response against intracellular bacteria in humans. These episodes are often severe and difficult to treat, frequently progressing to permanent disabilities. We performed RNA sequencing on paired skin biopsy specimens from nine leprosy patients before and during RR, identifying a 64-gene antimicrobial response signature that correlated with the concomitant decrease in Mycobacterium leprae bacilli in RR patients. The upstream regulators of this signature included both innate (IL-1β, TNF) and adaptive (IFN-γ, IL-17) cytokines, indicating induction of both Th1 and Th17 responses. Using a machine learning classifier to identify proteins with predicted membrane-permeating activity, we identified 28 genes in RR with previously unknown direct antimicrobial activity, including S100A8. We validated the antimicrobial activity of four proteins (S100A7, S100A8, CCL17, CCL19) against M. leprae in infected macrophages and axenic culture. Scanning electron microscopy revealed distinct membrane damage in bacteria exposed to these antimicrobial proteins. Our findings reveal dynamic antimicrobial gene regulation during RR and identify new host defense effectors, supporting therapeutic strategies that boost Th1/Th17 function to improve outcomes in mycobacterial infections.

overall_design: We performed a longitudinal bulk-RNA-sequencing study in paired skin biopsy specimens from nine L-lep patients at the time of clinical diagnosis (pre-RR) and at the onset of RR (reversal reaction). We further confirmed our findings in skin lesions of groups of leprosy patients without multidrug therapy (MDT) that included 10 borderline-tuberculoid (T-lep), 12 RR (RR pre-MDT) and 5 new lepromatous-lepromatous (LL).

middle: R

first: Feiyang

last: Ma

first: Jing

last: Lu

first: Matteo

last: Pellegrini

middle: L

last: Modlin


title: Effect of nonsense-mediated decay inhibition  on expression of 3' UTR spliced transcripts in colorectal carcinoma cell line HCT116

status: Public on Dec 20 2024

city: Sheffield

first: Jack

last: Riley

address: Firth Court, Western Bank

department: School of Biosciences

institute: The University of Sheffield

summary: The presence of an exon junction more than 55nt downstream of a stop codon is generally considered to be a trigger for nonsense-mediated decay (NMD). Thus, transcripts containing 3’UTR introns (3UIs) are often considered as transcriptional noise. Interestingly, upon knockdown of NMD factor UPF1 we see examples of both increased and decreased expression of the 3UI-spliced isoforms. In the oncogene HRAS, upon UPF1 knockdown, we see the 3UI-retaining isoform increases in expression where the spliced isoform decreases, suggesting that 3UI splicing protects the transcript against UPF1-mediated degradation.

overall_design: To investigate the role of nonsense-mediated decay on expression of 3'UTR spliced transcripts, HCT116 cells were treated with either siUPF1 or siDsRed (control) in the presence or absence of an NMD small molecule inhibitor. Following RNAseq, we conducted differential transcript usage and differential splicing analyses.

first: Cristina

middle: N

last: Alexandru

first: Ian

middle: M

last: Sudbery


title: PANDORA-seq uncovers sensitive mitochondrial small RNA alterations associated with increased lysosome activity in an Alzheimer’s Disease Mouse Model

status: Public on Dec 17 2024

city: Salt Lake City

first: Qi

last: Chen

state: Utah

department: Department of Human Genetics

institute: university of utah

type: Non-coding RNA profiling by high throughput sequencing

summary: Emerging small noncoding RNAs (sncRNAs), including tRNA-derived small RNAs (tsRNAs) and rRNA-derived small RNAs (rsRNAs), are critical in diverse biological processes, such as neurological diseases. Traditional sncRNA-seq protocols often miss these sncRNAs due to their modifications. We have recently developed PANDORA-seq, a method enabling more comprehensive detection of modified sncRNAs by overcoming the RNA modifications. Using PANDORA-seq, we have revealed an updated sncRNA profile enriched by tsRNAs/rsRNAs in the mouse cortex and found a particularly significant downregulation of mitochondrial tsRNAs and rsRNAs in an Alzheimer's disease (AD) mouse model, compared to genomic tsRNAs and rsRNAs. Moreover, our integrated analysis of cortex gene expression and sncRNA profiles reveals that those downregulated mitochondrial sncRNAs are negatively correlated with enhanced lysosomal activity, suggesting a crucial interplay between mitochondrial RNA dynamics and lysosomal function in AD. Given the versatile tsRNA/tsRNA molecular actions in cellular regulation, our data provides insights for future mechanistic study of AD with potential therapeutic strategies.

overall_design: Brain cortex tissue was collected from wild-type (WT) and Alzheimer's disease (AD) model mice, both with a C57BL/6J genetic background at 6 months of age. For each group, 8 biological replicates were analyzed using PANDORA-seq. In addition, traditional small non-coding RNA sequencing (sncRNA-seq) was performed with 7 replicates for the AD the WT group.

first: Xudong

last: Zhang

first: Tong

last: Zhou

first: Yumei

middle: E

last: Feng


title: Human RNaseH2 subunit upregulation counteracts oncogene- and chemotherapy-induced replication stress

status: Public on Dec 16 2024

city: Liverpool

first: Claire

last: Wilson

address: Ashton Street

institute: University of Liverpool

summary: RNaseH2 is a heterotrimeric endoribonuclease that resolves RNA:DNA hybrids and mis-incorporated ribonucleotides, which are implicated in DNA replication stress and cancer development and potential therapeutic targets. Individual RNaseH2 subunit mRNA or protein levels are found elevated in some cancers, but little is known about the mechanisms behind and impact of RNaseH2 subunit upregulation. We report that RNaseH2 subunits are upregulated at the protein level in response to replication stress induced by oncogenes and chemotherapy drugs hydroxyurea, gemcitabine and camptothecin in human cancer and non-cancer cell lines. We show that inducible overexpression of the RNaseH2B subunit increases the levels of the whole active RNaseH2 complex and can counteract drug-induced RNA:DNA hybrid formation. RNaseH2B overexpression reduces replication fork stalling induced by camptothecin and hydroxyurea but can itself increase RNA:DNA hybrid levels and cause replication stress. RNaseH2 subunit levels do not strongly impact survival of chemotherapy drug treatments but may have more subtle effects on genomic instability and cytoplasmic DNA signalling. In contrast, increased RNaseH2 subunit levels in presence of oncogenic HRAS are required to limit not only RAS-induced replication fork stalling but also cell death. Our findings shed new light on the functions of RNaseH2 and suggest that upregulation of RNaseH2 subunits may be an important aspect of replication stress responses in cancer.

overall_design: To test whether RNaseH2B overexpression alters gene expression, we performed RNA sequencing on HCT116 cells overexpressing RNaseH2B. Sequencing was peformed on HCT116 cells before and 48 hours after doxycycline-induced RNaseH2B induction. Parental HCT116 cells before and after doxycycline treatment were used as a control. We then performed gene expression profiling analysis using data obtained from RNA-Seq.

first: Rosanna

last: Wilkins

first: Richard

middle: D

last: Kelly

first: Aditi

last: Kanhere

first: Eva

last: Petermann


title: Differential Roles of Astrocytic CSF1 in Alzheimer's Disease and Cerebral Amyloid Angiopathy: Insights from Transcriptomic Analysis

status: Public on Dec 15 2024

city: Beijing

first: Wei

last: Ge

state: State...

address: 5 Dong Dan San Tiao, Dongcheng District

department: Department of Immunology

institute: Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences

summary: Alzheimer's disease (AD) and cerebral amyloid angiopathy (CAA) are neurodegenerative disorders characterized by the pathological deposition of amyloid-beta (Aβ) in the brain. Although both conditions share common pathogenic pathways, they exhibit distinct cellular manifestations and disease progression. Our study focused on the differential expression and role of astrocytic colony-stimulating factor 1 (CSF1) in these diseases. Using transcriptomic analysis of 248 brain tissue samples from the hippocampal-entorhinal system of 50 individuals, we identified a significant increase in CSF1 expression in the CA4 subfield of AD patients compared to a marked decrease in CAA. Functional investigations revealed that astrocytes with elevated CSF1 levels displayed neurotoxicity associated with AD-like pathology, whereas reduced CSF1 expression in astrocytes was linked to vascular damage typical of CAA. These findings suggest that CSF1 plays divergent roles in AD and CAA, contributing to their distinct pathological profiles. Our study highlights the potential of targeting astrocytic CSF1 expression as a differential diagnostic marker and therapeutic approach in managing these overlapping yet distinct neurological conditions.

overall_design: RNA sequencing (RNA-Seq) was used to examine gene expression in postmortem frozen human brain tissues from five subfields of the hippocampal-entorhinal system [cornu ammonis (CA) 1, CA2, CA3, CA4 and the entorhinal cortex (EC)].


title: BMP-2 generates a functional bone marrow niche by inducing the differentiation of local mesenchymal cells into CAR cells

city: Okayama

first: Ikue

last: Tosa

state: Okayama

address: Shikata-cho2-5-1, Kita-ku

country: Japan

institute: Okayama University

summary: In this study, we demonstrated that BMP-2 induced ectopic bone marrow presents microstructure, cellularity and functional hematopoiesis, and is a fully optimized microenvironment capable of sustaining the hematopoietic stem cells and the bone marrow niche-associated CXC-chemokine-ligand-12 (CXCL12)-abundant reticular (CAR) cells, comparably to native bone marrow.

overall_design: BMP-2 (8 µg)/β-TCP (40 mg) was implanted subcutaneouslly into C57BL6 or Cxcl12-GFP mice. BMP-2 induced ectopic bone marrow was harvested 4 weeks after implantation and compared with femur bone marrow. Hematopoietic cells were collected from C57BL6J mice and CAR cells were collected from Cxcl12-GFP mice by collagenase digestion.

first: Mitsuaki

last: Ono

first: Anh

last: Dang

first: Ziyi

last: Wang

first: Maiko

last: Kimura

first: Wakana

last: Kitagawa

first: Kei

last: Ishibashi

first: Hang

last: Do

first: Kun

last: Zhao

first: Emilio

middle: S

last: Hara

first: Noboru

last: Asada

first: Takashi

last: Nagasawa

first: Toshitaka

last: Oohashi

first: Takuo

last: Kuboki


title: Gene Expression Profiling of Triple Negative Breast Cancer Cells Responding to Adipocyte Secreted Factors

status: Public on Dec 02 2024

city: Atlanta

first: Karmella

last: Haynes

state: GA

address: 1760 Haygood Dr NE

department: Wallace H. Coulter Department of Biomedical Engineering

institute: Emory University

summary: Adipocyte-secreted factors (ASFs) induce lipid accumulation in breast cancer cells. These lipid droplets are thought to serve a cell-protective, pro-oncogenic role. ASFs including insulin, CCL5, and apelin interact with receptors that stimulate mTOR signaling in neighboring cells. One of the major transcription factors that acts downstream of this signaling pathway is SREBP1, which translocates into the nucleus and activates the expression of metabolic enzymes and other proteins involved in fatty acid synthesis, transport, and storage. Their associated enzymatic activities increase or decrease pools of metabolites that serve as substrates and cofactors for chromatin remodeling, such as acetyl-CoA, alpha-ketoglutarate (α-KG), flavin adenine dinucleotide (FAD+), and nicotinamide adenine dinucleotide (NAD+), but the impact on gene regulation in this context is not completely understood. To model biochemical signaling from adipocytes to breast cancer epithelial cells, we cultured differentiated mouse OP-9 cells, collected the adipocyte conditioned medium (ACM) after three days, and treated human breast epithelial BT-549 cells with ACM. Three days after treatment, we observed accumulation of lipid droplets in the cytoplasm of most BT-549 cells, whereas none were visible in cells grown in unconditioned medium (UCM). Transcription profiling verified the activation of lipid droplet biosynthesis genes, and revealed repression of loci at every chromosome, consistent with the idea that lipogenesis affects the availability of substrates and cofactors for global chromatin remodeling.

overall_design: We performed gene expression profiling analysis of RNA-seq data obtained of BT-549 cells treated with adipocyte-conditioned media (ACM) or unconditioned media (UCM) for 24 hours.

first: Yifei

last: Wu

first: Ashley

last: Townsel

first: Maya

last: Jaffe

first: Cara

last: Shields


title: Gene expression analysis of different motor neuron genotypes and their response to biological exposure and AAV treatment

status: Public on Dec 01 2024

city: Amsterdam

first: Andreia

last: Duarte

state: Select a State

address: Science Park 408

country: Netherlands

institute: VectorY

summary: Wild-type motor neurons and motor neurons carrying ALS-relevant mutations (TDP-43 - M337V; SOD1 - G93A) were cultured and exposed different biological treatments in the absence and presence of AAV(s) (1-5) transduction and consequent protein expression. Their response to these variables under different combinations was assessed with regards to gene expression using nCounter Gene Expression Panels (nanoString), Neuropathology and Neuroinflammation Panels.

overall_design: In total, 27 samples were analyzed using nanoString Technology across both panels. Several controls were included for: (a) gene mutation (wild-type vs mutated), (b) biological exposure (PSPC vs PONPC) and (c) AAV transduction (AAVs 1-5). 2-3 technical replicates were included per condition.

first: Sofia

last: Pascoal


title: Gene expression analysis of different motor neuron genotypes and their response to biological exposure and AAV treatment [neuropathology]


title: Analysis of gene regulation by the TP63 isoforms deltaNp63alpha and deltaNp63beta

first: Morgan

last: Sammons

summary: We analyzed the transcriptome changes of the colon carcinoma cell line HCT116 TP53-/- in response to expression of the TP63 isoforms deltaNp63alpha or deltaNp63beta. The goal was to understand how these C-terminal isoforms of p63 might differentially regulate gene expression.

overall_design: PolyA+ RNAseq (in biological triplicate) of HCT116 TP53-/- cell lines expressing either human deltaNp63alpha or deltaNp63beta in uninduced (- doxycyline) or induced (+ doxycycline) conditions for 24hrs.


title: CTCF-mediated 3D chromatin predetermines the gene expression program in the male germline (RNA-Seq)

first: Satoshi

last: Namekawa

summary: Spermatogenesis is a unidirectional differentiation process that generates haploid sperm, but how the gene expression program that directs this process is established is largely unknown. Here we determine the high-resolution 3D chromatin architecture of male germ cells during spermatogenesis and show that CTCF-mediated 3D chromatin predetermines the gene expression program required for spermatogenesis. In undifferentiated spermatogonia, CTCF-mediated chromatin interactions between meiosis-specific super-enhancers (SE) loci and target genes precede activation of meiosis-specific SEs on autosomes. These meiotic SE recruit the master transcription factor A-MYB in meiotic spermatocytes, which strengthens their 3D contacts and instructs a burst of meiotic gene expression. We also find that at the mitosis-to-meiosis transition, the germline-specific Polycomb protein SCML2 resolves chromatin loops that are specific to mitotic spermatogonia. Moreover, SCML2 and A-MYB establish the unique 3D chromatin organization of sex chromosomes during meiotic sex chromosome inactivation. We propose that CTCF-mediated 3D chromatin organization enforces epigenetic priming that directs unidirectional differentiation, thereby determining the cellular identity of the male germline.

overall_design: Testis were collected from P17 or 18 A-myb+/- and A-myb-/- mice and pachytene spermatocytes were isolated using fluorescence-activated cell sorting (FACS) with Vybrant DyeCycle Violet Stain (DCV)

first: Yuka

last: Kitamura

first: Kazuki

last: Takahashi

first: So

last: Maezawa

first: Yasuhisa

last: Munakata

first: Akihiko

last: Sakashita

first: Noam

last: Kaplan


title: Single-cell transcriptome analysis reveals abnormal angiogenesis and placentation by loss of imprinted glutaminyl-peptide cyclotransferase

city: Shanghai

first: jihong

last: zheng

state: Shanghai

address: Siping Road, Yangpu District

institute: Tongji University

summary: We provided a comprehensive transcriptome profile of Qpct-/- and Qpct+/+ mouse placentae and fills the gap of function of maternally expressed gene Qpct in mouse placental development

overall_design: We performed the single-cell and single-nucleus RNA library in Qpct+/+ and Qpct-/- mouse placentae (5 placenta as a pool n=5)

first: Jing

last: Guo

first: Jihong

last: Zheng

first: Ruixia

last: Li

first: Jindong

last: Yao

first: Xu

last: Wang

first: Chao

last: Zhang


title: Multinucleated giant cells are hallmarks of ovarian aging with unique immune and degradation-associated molecular signatures

status: Public on Dec 03 2024

first: Aubrey

last: Converse

address: 303 E superior

institute: Northwestern University Feinberg School of Medicine

summary: The ovary is one of the first organs to exhibit signs of aging, characterized by reduced tissue function, chronic inflammation, and fibrosis. Multinucleated giant cells (MNGCs), formed by macrophage fusion, typically occur in chronic immune pathologies, including infectious and non-infectious granulomas and the foreign body response, but are also observed in the aging ovary. The function and consequence of ovarian MNGCs remain unknown as their biological activity is highly context-dependent, and their large size has limited their isolation and analysis through technologies such as single-cell RNA sequencing. In this study, we define ovarian MNGCs through a deep analysis of their presence across age and species using advanced imaging technologies as well as their unique transcriptome using laser capture microdissection. MNGCs form complex interconnected networks that increase with age in both mouse and nonhuman primate ovaries. MNGCs are characterized by high Gpnmb expression, a putative marker of ovarian and non-ovarian MNGCs. Pathway analysis highlighted functions in apoptotic cell clearance, lipid metabolism, proteolysis, immune processes, and increased oxidative phosphorylation and antioxidant activity. Thus, MNGCs have signatures related to degradative processes, immune function, and high metabolic activity. These processes were enriched in MNGCs compared to primary ovarian macrophages, suggesting discrete functionality. MNGCs express CD4 and colocalize with T-cells, which were enriched in regions of MNGCs, indicative of a close interaction between these immune cell types. These findings implicate MNGCs in modulation of the ovarian immune landscape during aging given their high penetrance and unique molecular signature that supports degradative and immune functions.

overall_design: Transcriptomic profiling of ovarian multinucleated giant cells, stroma, and primary macrophages were compared

first: Francesca

last: Duncan

first: Michele

last: Pritchard


title: CTCF-mediated 3D chromatin predetermines the gene expression program in the male germline

first: Satoshi

last: Namekawa

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

overall_design: Refer to individual Series


title: CLIP-seq analysis of WT and Gag chimera HIV-1 virions 2

status: Public on Nov 30 2024

city: Saint Louis

first: Sebla

middle: Bulent

last: Kutluay

state: MO

address: 660 S. Euclid Avenue, Campus Box 8230

institute: Washington University School of Medicine

summary: To investigate how the major HIV-1 structural protein Gag packages viral genomes efficiently, we replaced the NC domain of Gag with RNA binding domains from heterologous cellular RNA binding proteins to generate Gag chimeras.  We hypothesize that the Gag chimeras that preferentially bind to purine-rich RNA sequences will package viral genomes efficiently. We performed CLIP-seq to identify the RNA sequences bound by WT Gag and Gag chimeras in cells, at the plasma membrane, and in virions.

overall_design: 293T cells in 10 cm dishes were transfected with PR- NL4-3-derived proviral plasmids expressing the indicated Gag. One day following transfection and 14-16 hours before cells and virions are harvested, 100 μM 4-thiouridine (4SU) or 6-thioguanosine (6SG) was added to the cell culture medium. For cell Gag CLIP and virion Gag CLIP, whole cells and virions were UV-crosslinked and lysed, respectively. For plasma membrane (PM) Gag CLIP, cells were UV irradiated and then fractionated via membrane flotation. Gag-RNA complexes were immunoprecipitated from whole cell lysates, virion lysates, and PM fraction. Gag-bound RNAs were subsequently extracted for CLIP-seq library construction. Sequencing was done on Nextseq500 platform.


title: Atlas of multilineage stem cell differentiation reveals TMEM88 as a developmental regulator of blood pressure

status: Public on Dec 05 2024

city: St Lucia

first: Sophie

last: Shen

state: Queensland

address: 306 Carmody Road

department: Institute for Molecular Bioscience

country: Australia

institute: The University of Queensland

summary: Pluripotent stem cells provide a scalable approach to analyse molecular regulation of cell differentiation across multiple any developmental lineage trajectories. In this study, we engineered barcoded iPSCs to generate an atlas of multilineage differentiation from pluripotency, encompassing a time-course of WNT-induced differentiation perturbed using modulators of WNT, BMP, and VEGF signalling. Computational mapping of in vitro cell types to in vivo developmental lineages revealed a diversity of iPSC-derived cell types comprising mesendoderm lineage cell types including lateral plate and paraxial mesoderm, neural crest, and primitive gut. Coupling this atlas of in vitro differentiation with Summary data-based Mendelian Randomisation analysis of human complex traits, we identify the WNT-inhibitor protein TMEM88 as a putative regulator of mesendodermal cell types governing development of diverse cardiovascular and anthropometric traits. Using genetic loss of function models, we show that TMEM88 is required for differentiation of diverse endoderm and mesoderm cell lineages in vitro and that TMEM88 knockout in vivo results in a significant dysregulation of arterial blood pressure. This study provides an atlas of multilineage iPSC differentiation coupled with new molecular, computational, and statistical genetic tools to dissect genetic determinants of mammalian developmental physiology.

overall_design: We first used scRNA-seq to test our genomic barcode (BC)-based multiplexing method in undifferentiated human induced pluripotent stem cells (hiPSCs), pooling 18 different samples into one scRNA-seq experiment. Next, we used these 18 barcoded hiPSC lines across three scRNA-seq libraries to assess effects of nine different small molecule signalling modulation treatments (including a control) on iPSC differentiation towards diverse mesendoderm lineages across day 2, day 5, and day 9 of differentiation, with two replicates per permutation. We separately use Cell Hashing HTOs to multiplex eight time points, each 24 hours apart, from day 2 to day 9 of this mesendoderm differentiation under control conditions for a scRNA-seq time course reference. We generated two additional time course scRNA-seq datasets with Cell Hashing matching the time points from the reference. The first dataset employs a CRISPRi-mediated TMEM88 knockdown in hiPSCs to assess the effect of TMEM88 loss-of-function on mesendoderm differentiation (gain of Wnt signalling), while the hiPSCs from the second dataset are treated with Wnt inhibitor XAV-939 to observe effects of an opposite direction of Wnt signalling change.

first: Tessa

last: Werner

first: Samuel

middle: W

last: Lukowski

first: Stacey

last: Andersen

first: Yuliangzi

last: Sun

first: Woo Jun

last: Shim

first: Dalia

last: Mizikovsky

first: Sakurako

last: Kobayashi

first: Jennifer

last: Outhwaite

first: Han Sheng

last: Chiu

first: Xiaoli

last: Chen

first: Gavin

last: Chapman

first: Ella

last: Martin

first: Di

last: Xia

first: Duy

last: Pham

first: Zezhuo

last: Su

first: Daniel

last: Kim

first: Pengyi

last: Yang

first: Men Chee

last: Tan

first: Enakshi

last: Sinniah

first: Qiongyi

last: Zhao

first: Sumedha

last: Negi

first: Meredith

last: Redd

first: Joseph

last: Powell

first: Sally

last: Dunwoodie

first: Patrick

last: Tam

first: Mikael

last: Bodén

first: Joshua

last: Ho

first: Quan

last: Nguyen

first: Nathan

last: Palpant


title: CLIP-seq analysis of WT and Gag chimera HIV-1 virions 3

first: Sebla

last: Kutluay


title: RNA-Seq results of cervical spinal cords from NEAT1-KO; hSOD1G93A mice, compared to NEAT1-WT; hSOD1G93A

city: Ngoya

first: Yu

last: Kawakami

address: Showaku, Tsurumai, 65

institute: Nagoya University

summary: Transcriptional changes of NEAT1-KO; hSOD1G93A mice and NEAT1-WT; hSOD1G93A mice were compared. RNA-seq gene expression analysis revealed that differentially expressed genes were associated with protein folding as biological process.

overall_design: We crossed NEAT1-KO mice with hSOD1G93A mice to generate NEAT1-KO; hSOD1G93A and NEAT1-WT; hSOD1G93A mice. We performed gene expression profiling analysis using data obtained from RNA-seq of 3-4 samples from each genotype mice at 8-week-old.

first: Yohei

last: Iguchi

first: Masahisa

last: Katsuno


title: Progenitor Cell Dynamics in AGA: Insights from Spatially Resolved Transcriptomics

status: Public on Nov 27 2024

city: Rangsit

first: Sasin

last: Charoensuksira

state: Pathum Thani

address: 99 Moo 18 Klong Luang

country: Thailand

institute: Thammasat University

summary: Research has shown that, apart from androgen-sensitive dermal papilla cells, hair follicle progenitor cell dynamics play a critical role in androgenetic alopecia (AGA). This study focuses on these progenitor cells, applying the spatial transcriptome technique to reveal molecular disturbances in the progenitor cells (CD34+ area) between AGA patients (PG-A) and normal controls (PG-C).

overall_design: The spatial transcriptome technique was utilized to specifically analyze the hair follicle progenitor cell areas. This approach enabled the identification of key molecular differences between the progenitor cells from AGA patients and those from normal donors, providing new insights into the cellular mechanisms influencing AGA.

first: Saranyoo

last: Ponnikorn

first: Piyaporn

last: Surinlert

first: Wilai

last: Thanasarnaksorn

first: Aungkana

last: Krajarng

first: Suradej

last: Hongeng

first: Juthapa

last: Pongklaokam


title: PIP4K2C inhibition reverses autophagic flux impairment induced by SARS-CoV-2

city: Stanford

first: Halise Busra

last: Cagirici

state: California

address: 300 Pasteur Drive

institute: Stanford University

summary: In search for broad-spectrum antivirals, we discovered a small molecule inhibitor, RMC-113, that potently suppresses the replication of multiple RNA viruses including SARS-CoV-2 in human lung organoids. We demonstrated selective dual inhibition of the lipid kinases PIP4K2C and PIKfyve by RMC-113 and target engagement by its clickable analog. Advanced lipidomics revealed alteration of SARS-CoV-2-induced phosphoinositide signature by RMC-113 and linked its antiviral effect with functional PIP4K2C and PIKfyve inhibition. We discovered PIP4K2C’s roles in SARS-CoV-2 entry, RNA replication, and assembly/egress, validating it as a druggable antiviral target. Integrating proteomics, single-cell transcriptomics, and functional assays revealed that PIP4K2C regulates virus-induced impairment of autophagic flux. Reversing this autophagic flux impairment via promoting degradation of viral protein is a mechanism of antiviral action of RMC-113. These findings reveal virus-induced autophagy regulation via PIP4K2C, an understudied kinase, and propose dual inhibition of PIP4K2C and PIKfyve as a candidate strategy to combat emerging viruses.

overall_design: Adult lung organoid (ALOs) were infected with SARS-CoV-2 virus (MOI=1) fand treated with RMC-113 compound or DMSO for 4 h or 24 h. Uninfected ALOs were treated with RMC-113 and DMSO for 4h and 24 h.

first: Marwah

last: Karim

first: Manjari

last: Mishra

first: Chieh-Wen

last: Lo

first: Sirle

last: Saul

first: Halise

last: Busra Cagirici

first: Do

last: Hoang Nhu Tran

first: Aditi

last: Agrawal

first: Luca

last: Ghita

first: Amrita

last: Ojha

first: Michael

middle: P

last: East

first: Karen

last: Anbro Gammeltoft

first: Malaya

last: Kumar Sahoo

first: Gary

last: Johnson

first: Soumita

last: Das

first: Pieter

last: Leyssen

first: Johan

last: Neyts

first: Winston

last: Chiu

first: Courtney

last: Cohen

first: Jens

last: Bukh

first: Judith

last: Gottwein

first: John

last: Dye

first: Norma

last: Neff

first: Benjamin

last: Pinsky

first: Tuomo

last: Laitinen

first: Tatu

last: Pantsar

first: Antti

last: Poso

first: Fabio

last: Zanini

first: Steven

last: De Jonghe

first: Christopher

last: Asquith

first: Shirit

last: Einav


title: CLIP-seq analysis of WT and Gag chimera HIV-1 virions 1

first: Sebla

last: Kutluay


title: Human eIF2A has a minimal role in translation initiation and in uORFmediated translational control

status: Public on Nov 29 2024

city: Heidelberg

first: Mykola

last: Roiuk

address: Im Neuenheimer Feld 580

country: Germany

institute: DKFZ

summary: We apply ribosome profiling here to assess the role of eIF2A in translation initiation. For this we test the change in translation efficiency between HeLa control and eIF2A-KO cells, however we do not find any transcript to depend on eIF2A. Since eIF2A is thought to take over the function of eIF2 when eIF2 is inhibited, we also test conditions where the integrated stress response is activated, thereby leading to eIF2 inactivation. In none of our assays, however, could we detect a role of eIF2A in translation initiation.

overall_design: To find, if there are any transcripts dependent on eIF2A, we generated HeLa eIF2A knock-out cell line and performed ribosome profiling on it and an isogenic control HeLa cell line. This experiment was performed in triplicate. To test if inhibition of eIF2 redirects translation initiation to eIF2A we again performed ribosome profiling on the above-mentioned cell lines in combination with tunicamycin (16 hours) treatment. This experiment was performed in duplicates.

first: Aurelio

last: Teleman


title: A histone hyperacetylation pulse induces cellular memory of gene expression and 3-dimensional genome folding [ATAC-seq]

status: Public on Nov 22 2024

city: Montpellier

first: Marco

last: Di Stefano

address: 141 rue de la Cardonille

country: France

institute: Institut de Génétique Humaine - CNRS

summary: Epigenetic memory enables the stable propagation of gene expression patterns in response to transient developmental and environmental stimuli. Although 3-dimensional (3D) organisation is emerging as a key regulator of genome function due its role in gene expression control, it is unknown whether it contributes to cellular memory. Here, we establish that acute perturbation of the epigenome can induce cellular memory of gene expression in mouse embryonic stem cells (mESCs). Specifically, we uncover how a pulse of histone deacetylase inhibition translates to changes in the histone acetylation and methylation landscape, as well as global and local genome folding. While most epigenomic and transcriptional changes are initially reversed once the perturbation is removed, architectural features are not fully reestablished. Upon a second transient pulse of hyperacetylation, hundreds of genes maintain their dysregulated state. Using ultra-deep Micro-C, we associate memory of gene expression with enhancer-promoter contacts and repressive chromatin topology mediated by Polycomb. These results uncover the complex interplay between different epigenetic regulatory layers and establish a novel link between genome folding and cellular memory.

overall_design: Micro-C experiments in mouse ESCs in control (DMSO), TSA-treated or 24-hour recovery (REC) conditions.

first: Flora

last: Paldi

first: Michael Florian

last: Szalay

first: Giacomo

last: Cavalli


title: Ggnbp2 and Ifih1 knockout in HoxA9 immortalised R26-CreERT2 Adar1fl/E861A myeliod cell line.

status: Public on Nov 21 2024

city: Meblourne

first: Alistair

middle: Morgan

last: Chalk

state: VIC

address: 1G Royal Parade

department: Bioinformatics

institute: WEHI

first: J

last: Heraud-Farlow

first: A

first: S

last: Taylor

last: Gupte

first: C

last: Walkley


title: A histone hyperacetylation pulse induces cellular memory of gene expression and 3-dimensional genome folding [RNA-seq]

summary: Epigenetic memory enables the stable propagation of gene expression patterns in response to transient developmental and environmental stimuli. Although 3-dimensional (3D) organisation is emerging as a key regulator of genome function due its role in gene expression control, it is unknown whether it contributes to cellular memory. Here, we establish that acute perturbation of the epigenome can induce cellular memory of gene expression in mouse embryonic stem cells (mESCs). Specifically, we uncover how a pulse of histone deacetylase inhibition translates to changes in the histone acetylation and methylation landscape, as well as global and local genome folding. While most epigenomic and transcriptional changes are readily reversible once the perturbation is removed, architectural features are not fully reestablished. Upon a second transient pulse of hyperacetylation, hundreds of genes maintain their dysregulated state. Using ultra-deep Micro-C, we associate memory of gene expression with enhancer-promoter contacts and repressive chromatin topology mediated by Polycomb. These results uncover the complex interplay between different epigenetic regulatory layers and establish a novel link between genome folding and cellular memory.

overall_design: RNA-seq experiments in wild type mouse ESCs, NPCs, PCGF4-/- PCGF2fl/fl ESCs and mouse gastruloids.

first: Solene

last: Dufau


title: Transient histone deacetylase inhibition induces cellular memory of gene expression and three-dimensional genome folding


title: Fractionation of sgControl, sgIfih1 and sgGgnbp2 in HoxA9 immortalized Rosa26-CreERT2 Adar1fl/E861A immortalised myeloid cells.

overall_design: Fractionation was performed on three independent validated clones of sgControl, sgIfih1 and sgGgnbp2 Rosa26-CreERT2 Adar1fl/E861A immortalised myeloid cells. 12x106 myeloid cells (either 1 day or 3 days in 200nM tamoxifen as described above) were treated with AEBSF (0.5mM) for 30 minutes in culture media to inhibit proteases prior to lysis. 2x106 cells were taken for total RNA extraction as previously described using Bioline RNA isolation kit.


title: On the way to elucidate the effects of cannabidiol (CBD) on living organisms - assessment of changes in the global transcription profile of kidney in a mouse model

status: Public on Nov 26 2024

city: Kraków

first: Artur

last: Gurgul

address: Rędzina 1C

department: Center for Experimental and Innovative Medicine

country: Poland

institute: University of Agriculture in Krakow

summary:
Cannabidiol (CBD) is a phytocannabinoid known for its anti-inflammatory, immunomodulatory, antioxidant, anxiolytic, antipsychotic, and anticonvulsant effects. The systemic administration of CBD also has the potential to impact kidney function, either beneficially or adversely. This study aims to examine the effects of CBD on behavior, immune system activity, and stress levels, and to determine its impact on transcriptomic alterations in selected organs of mice.
(Study funded by the National Science Centre, Poland;  project no. 2020/39/O/NZ9/00821)

overall_design:
Male mice of the standard C57BL/6J strain were used. The animals were added to the long-term intraperitoneal CBD administration (28 days). There were two subgroups receiving different doses of CBD - medium (10 mg/kg bw) and high, used in some clinical trials (20 mg/kg bw). Additionally, a control group (placebo/vehicle) was maintained. Chemically pure Cannabidiol (CBD) was administered intraperitoneally (in the research group) to ensure that the substances reached the blood vessels of the digestive system. CBD was administered in a single daily dose. The mice were housed under standard conditions and with standard feeding and watering ad libitum. They were also examined every third day to determine well-being and health. After the period of CBD administration, the mice were immediately euthanized and organs such as the brain, spleen and liver, kidneys, and others were collected. In this part of the experiment, kidney samples were used to isolate the total RNA. The obtained RNA was quality assessed and used to prepare directional libraries for high throughput transcriptome sequencing using CORALL mRNA-Seq V2 kit (Lexogen).  The libraries were sequenced commercially in a 2x150bp paired-end run. The obtained sequence data were used to analyze the differential expression between the different CBD dosages and the control group. Differentially expressed genes were classified into particular processes or functions and were subjected to statistical testing for overrepresentation. The synthesis of the obtained data will allow to propose of potential mechanisms of CBD action and create an atlas of changes taking place in the transcriptome of the main target organs.
(Study funded by the National Science Centre, Poland;  project no. 2020/39/O/NZ9/00821)

first: Mikołaj

last: Rokicki

first: Jakub

last: Żurowski

first: Ewa

last: Ocłoń

first: Tomasz

last: Szmatoła

first: Igor

last: Jasielczuk

first: Karolina

last: Mizera-Szpilka


title: The genetic architecture of cell-type-specific cis-regulation [RNA-seq]

status: Public on Nov 19 2024

city: Athens

first: Robert

last: Schmitz

address: B416 Davison Life Sciences

department: Genetics

institute: University of Georgia

summary: An understanding of how genetic variants affect chromatin accessibility in unique cellular context remains poorly understood. Here, we generated single-cell chromatin accessibility profiles in nearly 200 diverse maize inbred lines, revealing for the first time variants that affect chromatin accessibility in distinct cellular contexts.

overall_design: snRNA-seq on maize 7 DAS seedlings (B73 and Mo17)

first: Alexandre

last: Marand


title: The genetic architecture of single-cell chromatin accessibility in maize


title: Spatial deconvolution of tertiary lymphoid structure in human IPF lungs revealed novel CCL19-expressing mural cells

city: New Brunzwick

first: Zhi

last: Yang

state: New Jersey

address: 89 French Street

department: Child Health Institute of New Jersey

institute: Rutgers, the state university of New Jersey

summary: Tertiary lymphoid structures (TLS) are commonly observed in human idiopathic pulmonary fibrosis (IPF) lungs. The specific immune and non-immune cells in the TLS of IPF patients, and the factors that drive TLS formation, remain largely unknown. Here we spatially deconvoluted immune and non-immune cells in the TLS of human IPF lungs, and examined the signals underlying TSL development in IPF patients. We identified a novel subset of CCL19hi-IPF-associated-mural cells (CCL19hiIAMC) that enveloped the vessels inside the TLS.  CCL19hiIAMCs were the major source of CCL19 inside the TLS of IPF lungs, attracting T/B lymphocytes and unique CCR7+DCs to drive TLS formation. CCL19hiIAMCs also provided other pro-TLS and proinflammatory molecules to promote lymphocyte maturation and tissue inflammation. Our data also reveal that various IPF-associated fibroblasts surrounded the TLS, promoting TLS development and plasma cells dissemination via CXCL12/CXCR4 signaling. CCR7 and CXCR4 signaling attracted lymphocytes and DCs that upregulated pro-TLS genes in the IPF microenvironment to enhance TLS formation. Together, these findings have deconvoluted the cell subsets and their transcriptomes in the TLS of IPF lungs and suggest that novel CCL19hiIAMCs may drive TLS formation in collaboration with various immune cells and fibroblasts.

overall_design: Nine human IPF tissues from different patients were used for spatial gene expression transcriptomics, including one fresh frozen sample and eight PPFE samples.

first: Qi


title: GGNBP2 regulates MDA5 sensing triggered by self double stranded RNA following loss of ADAR1 editing

first: Alistair

summary: Adenosine-to-Inosine (A-to-I) editing of double stranded RNA (dsRNA) is an essential modifier of dsRNA immunogenicity. Loss of Adenosine deaminase acting on RNA 1 (ADAR1), or its A-to-I editing activity, triggers a lethal MDA5-dependent autoinflammatory response to endogenous unedited dsRNA. We performed a genome-wide suppressor screen to map the genetic landscape regulating the response to a loss of ADAR1 mediated A-to-I editing. This identified that the interacting proteins GGNBP2, CNOT10 and CNOT11 regulate immunogenic sensing following loss of A-to-I editing. GGNBP2 acts between transcription and cytoplasmic MDA5 sensing. Loss of GGNBP2 modified the distribution of dsRNA within cells, reducing the cytoplasmic dsRNA load and preventing induction of type I IFN and lethal autoinflammation. GGNBP2, CNOT10 and CNOT11 are novel regulators of the cellular response to unedited dsRNA.


title: The genetic architecture of cell-type-specific cis-regulation [ATAC-seq]

summary: Gene expression and complex phenotypes are determined by the activity of cis-regulatory elements. However, an understanding of how extant genetic variants affect cis-regulatory activity remains limited. Here, we investigated the consequences of cis-regulatory diversity using single-cell genomics of >0.7 million nuclei across 172 maize inbreds. Our analyses pinpointed cis-regulatory elements distinct to domesticated maize and how transposons rewired the regulatory landscape. We found widespread chromatin accessibility variation associated with >4.6 million genetic variants with largely cell-type-specific effects. Variants in TEOSINTE BRANCHED1/CYCLOIDEA/PROLIFERATING CELL FACTOR binding sites were the most prevalent determinants of chromatin accessibility. Finally, integration of genetic variants associated with chromatin accessibility, organismal trait variation, and population differentiation revealed how local adaptation has rewired regulatory networks in unique cellular context to alter maize flowering phenotypes.

overall_design: Single-cell genomics for seven day old seedlings across 172 maize inbred lines.


title: Tamoxifen timecourse in HoxA9 immortalized R26-CreERT2 Adar1fl/+ and R26-CreERT2 Adar1fl/E861A myeliod cell lines.

overall_design: Tamoxifen time course in two independent R26-CreERT2 Adar1fl/+ and two independent R26-CreERT2 Adar1fl/E861A immortalised myeloid cell lines. Days 0, 2, 4 and 7.

first: J


title: CTCF-mediated 3D chromatin predetermines the gene expression program in the male germline (ChIP-seq)

status: Public on Dec 13 2024

city: Davis

first: Satoshi

last: Namekawa

address: One Shields Avenue

institute: University of California, Davis

summary: Spermatogenesis is a unidirectional differentiation process to produce haploid sperm. However, it remains largely unknown how the gene expression program is determined to direct a unidirectional differentiation. Here we unveil the high-resolution 3D chromatin architecture of male germ cells and show that CTCF-mediated 3D chromatin predetermines the gene expression program required for spermatogenesis. At the mitosis-to-meiosis transition, the germline-specific Polycomb protein SCML2 resolves chromatin loops specific to mitotic spermatogonia. On autosomes, CTCF-mediated 3D chromatin manifests the structural feature of meiosis-specific super-enhancer (meiotic SE) loci already in undifferentiated spermatogonia. In meiotic spermatocytes, the master transcription factor A-MYB is recruited to these meiotic SE loci to strengthen their 3D contacts to instruct the burst of meiotic gene expression. Further, SCML2 and A-MYB establish unique 3D chromatin of the sex chromosomes in meiotic sex chromosome inactivation. We propose that CTCF-mediated 3D chromatin underlines epigenetic priming to direct unidirectional differentiation, thereby determining the cellular identity of the male germline.

overall_design: Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for CTCF in THY1+ spermatogonia and pachytene spermatocytes.

first: Yuka

last: Kitamura

first: Kazuki

last: Takahashi

first: So

last: Maezawa

first: Yasuhisa

last: Munakata

first: Akihiko

last: Sakashita

first: Noam

last: Kaplan


title: Genome-wide CRISPR screen of HoxA9 immortalised R26-CreERT2 Adar1fl/+ and R26-CreERT2 Adar1fl/E861A cells

last: Chalk

summary: ADAR1 E861A CRISPR knockout pooled library screen

overall_design: Genome-wide CRISPR knockout screen, Cas9 HoxA9 immortalised R26-CreERT2 Adar1fl/+ and R26-CreERT2 Adar1fl/E861A cells with a genome-wide Brie sgRNA library. 2 independent replicates of each genotype, sampled at 0, 7 and 14 days post tamoxifen .

last: Heraud-Farlow

first: A

first: S

last: Taylor

last: Gupte

first: I

last: Nikolic

first: K

last: Simpson

first: C

last: Walkley


title: Krill oil supplementation in vivo promotes increased fuel metabolism and protein synthesis in cultured human skeletal muscle cells

status: Public on Nov 20 2024

city: Oslo

first: Parmeshwar

middle: Bajirao

last: Katare

state: Oslo

address: Gydas vei 8

department: Section for Pharmacology and Pharmaceutical Biosciences, Department of Pharmacy,

country: Norway

institute: University of Oslo

summary: Krill oil is a dietary supplement derived from Antarctic krill; a small crustacean found in the ocean. Krill oil is a rich source of omega-3 fatty acids, specifically eicosapentaenoic acid and docosahexaenoic acid, as well as the antioxidant astaxanthin. The aim of this study was to investigate the effects of krill oil supplementation, compared to placebo oil (high oleic sunflower oil added astaxanthin), in vivo on energy metabolism and substrate turnover in skeletal muscle cells. Skeletal muscle cells (myotubes) were obtained before and after a 7-week krill oil or placebo oil intervention, and glucose and oleic acid metabolism and leucine accumulation, as well as effects of different stimuli in vitro, were studied in the myotubes. In vivo intervention with krill oil increased oleic acid oxidation and leucine accumulation in skeletal muscle cells, however no effects were observed on glucose metabolism. The krill oil-intervention-induced increase in oleic acid oxidation correlated negatively with changes in serum low-density lipoprotein (LDL) concentration. In addition, myotubes were also exposed to krill oil in vitro. The in vitro study revealed that 24 h of krill oil treatment increased both glucose and oleic acid metabolism, enhancing energy substrate utilization. Transcriptomic analysis comparing myotubes obtained before and after krill oil-supplementation identified differentially expressed genes associated with e.g. glycolysis/gluconeogenesis, metabolic pathways and calcium signaling pathway, while proteomic analysis demonstrated upregulation of e.g. LDL-receptor in myotubes obtained after krill oil intervention. These findings suggest that krill oil intervention promotes increased fuel metabolism and protein synthesis in human skeletal muscle cells, with potential implications for metabolic health.

overall_design: To investigate the effects of krill oil supplementation, in vivo on energy metabolism and substrate turnover in skeletal muscle cells. Skeletal muscle cells (myotubes) were obtained before (AA) and after (BB) a 7-week krill oil intervention, and glucose and oleic acid metabolism and leucine accumulation, as well as effects of different stimuli in vitro, were studied in the myotubes.

middle: B

first: Andrea

last: Fernandez

first: Abel

last: Mengeste

first: Farnaz

last: Navabakbar

first: Håvard

last: Hamarsland

first: Stian

last: Ellefsen

first: Rolf

middle: K

last: Berge

first: Hege

middle: G

last: Bakke

first: Tuula

last: Nyman

first: Eili

last: Kase

first: Arild

middle: C

last: Rustan

last: Thoresen


title: A histone hyperacetylation pulse induces cellular memory of gene expression and 3-dimensional genome folding [microC_3]

first: Marco

last: Di Stefano

first: Flora

last: Paldi

first: Solene

last: Dufau

first: Michael Florian

last: Szalay

first: Giacomo

last: Cavalli


title: Ggnbp2 knockout in HoxA9 immortalised R26-CreERT2 Adar1fl/+ myeliod cell line.

first: Alistair

last: Chalk

overall_design: Ggnbp2 and EV knockout in HoxA9 immortalised R26-CreERT2 Adar1fl/+ myeliod cell line, in triplicate.

first: J

last: Heraud-Farlow

first: A

first: S

last: Taylor

last: Gupte

first: C

last: Walkley


title: Effect of PHRF1 depletion on gene expression in HeLa cells

status: Public on Nov 18 2024

city: Chapel Hill

first: Kanishk

last: Jain

state: NC

address: 120 Mason Farm Rd. CB #7260, Genetic Medicine Building, Ste 3010

institute: UNC Chapel Hill

summary: The goal of this study was to understand how PHRF1 regulates gene expression in HeLa cells. By conducting RNA sequencing of 3 WT and 3 KO (PHRF1 knockout) samples, we found that DNA damage response pathways were upregulated and splicing-related pathways were downregulated in KO cells versus WT cells.

overall_design: PHRF1 deletion in HeLa cells was generated through CRISPR-Cas9 mediated gene editing. Total RNA was extracted from 3 WT and 3 KO Hela cell populations and prepared for RNA sequencing. Differential gene expression analysis based on the sequencing data was conducted, comparing gene expression in  PHRF1 KO cells to WT cells.


title: Single-cell RNA sequencing of human endometrium across the luteal phase of the menstrual cycle

status: Public on Nov 06 2024

city: Coventry

first: Jan

last: Brosens

address: University Hospital Coventry and Warwickshire

department: Warwick Medical School

institute: University of Warwick

summary: The fate of the human endometrium is determined during the mid-luteal window of implantation, a crucial period when endometrial stromal cells (EnSCs) differentiate into specialized decidual cells. Upon embryo implantation, these differentiating EnSCs transform the endometrium into the decidua of pregnancy. Conversely, in the absence of pregnancy, decreasing progesterone levels trigger tissue breakdown, leading to menstruation. Despite our understanding of these processes, the precise mechanisms governing this tissue transformation remain elusive. To bridge this knowledge gap, we conducted single-cell RNA sequencing, mapping the transcriptomic profiles of timed endometrial biopsies throughout the luteal phase of the menstrual cycle.

overall_design: Fresh endometrial biopsies across the secretory phase (LH+5-11) were recovered and subjected to 10 x single-cell RNA sequencing experiment

first: Emma

last: Lucas

first: Pavle

last: Vrljicak

first: Thomas

last: Rawlings

first: Chow Seng

last: Kong


title: RNA-Seq results of a neuronal cell model of SBMA, AR-97Q cells, compared to AR-24Q cells

status: Public on Nov 05 2024

city: Nagoya

first: Madoka

last: Iida

state: Aichi

address: 65 Tsurumai-cho Showa-ku

department: Neurology department

summary: Spinal and bulbar muscular atrophy (SBMA) is a neuromuscular disorder caused by a polyglutamine expansion in the androgen receptor (AR). Transcriptional changes were compared between a muscle cell model of SBMA (AR-97Q cells) and a control model (AR-24Q cells). RNA-seq gene expression analysis revealed that differentially expressed genes were associated with upregulation of transforming growth factor β (TGFβ) signaling, mitogen-activated protein kinase (MAPK), tumor necrosis factor (TNF) signaling, calcium signaling, and NFκB signaling.

overall_design: C2C12 AR-24Q cells and AR-97Q cells were genetically modified to stably express full-length human AR with 24 or 97 poly-glutamine repeats.

first: Kenji

last: Sakakibara

first: Ryuji

last: Kato

first: Masahisa

last: Katsuno


title: Crypt density and recruited enhancers underlie intestinal tumour initiation [Bisulfite-seq]

status: Public on Nov 15 2024

city: Boston

first: Ramesh

middle: A.

last: Shivdasani

state: MA

address: 450 Brookline Ave. SM1060

department: Medical Oncology

institute: Dana-Farber Cancer Institute

type: Methylation profiling by high throughput sequencing

summary: Oncogenic mutations that drive colorectal cancer can be present in healthy intestines for long periods without overt consequence. Mutation of Adenomatous polyposis coli (Apc), the most common initiating event in conventional adenomas, activates Wnt signaling, hence conferring fitness on mutant intestinal stem cells (ISCs). Apc mutations may occur in ISCs that arose by routine self-renewal or by dedifferentiation of their progeny. Although ISCs of these different origins are fundamentally similar, it is unclear if both generate tumours equally well in uninjured intestines. Also unknown is whether cis-regulatory elements are substantively modulated upon Wnt hyperactivation or as a feature of subsequent tumours. Here, we show in two mouse models that adenomas are not an obligatory outcome of Apc deletion in either ISC source but require proximity of mutant intestinal crypts. Reduced crypt density abrogates, and aggregation of mutant colonic crypts augments, adenoma formation. Moreover, adenoma-resident ISCs open chromatin at thousands of enhancers that are inaccessible in Apc-null ISCs not associated with adenomas. These cis-elements explain adenoma-selective gene activity and persist, with little further expansion of the repertoire, as other oncogenic mutations accumulate. Thus, cooperativity between neighbouring mutant crypts and new accessibility at specific enhancers are key steps early in intestinal tumourigenesis.

overall_design: To validate the effect of Apc loss (constitutive Wnt activation) on the intestinal stem cell (ISC) epigenome, we performed whole-genome bisulfite sequencing (WGBS) on primary intestinal stem cells (ISCs) isolated from the duodenum 14 days after Apc gene deletion.

first: Liam

last: Gaynor

first: Harshabad

last: Singh

first: Guodong

last: Tie

first: Shariq

last: Madha

first: Unmesh

last: Jadhav

first: Andrew

last: Mancini

first: Mikio

last: Hoshino

first: Frederic

last: de Sauvage

first: Kazutaka

last: Murata

middle: A


title: Effect of PHRF1 depletion on gene expression in HCT116 cells

first: Kanishk

last: Jain

summary: The goal of this study was to understand how PHRF1 regulates gene expression in HCT116 cells. By conducting RNA sequencing of 3 WT and 3 KO (PHRF1 knockout) samples, we found that DNA damage response pathways were upregulated and splicing-related pathways were downregulated in KO cells versus WT cells.

overall_design: PHRF1 deletion in HCT116 cells was generated through CRISPR-Cas9 mediated gene editing. Total RNA was extracted from 3 WT and 3 KO Hela cell populations and prepared for RNA sequencing. Differential gene expression analysis based on the sequencing data was conducted, comparing gene expression in  PHRF1 KO cells to WT cells.


title: Crypt density and recruited enhancers underlie intestinal tumour initiation

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

overall_design: Refer to individual Series


title: Transcriptomic profiling across 20 CC-RIX genetic backgrounds with and without high-fat high-sugar (HFHS) diet, and with and without metformin [liver]

city: Bar Harbor

first: Gary

last: Churchill

state: Maine

address: 600 Main St

institute: The Jackson Laboratory

summary:
Adipose, liver, and skeletal muscle tissue from 6-month and 12-month CC-RIX cohorts underwent transcriptomic profiling. CC-RIX cohorts were exposed to sensitizer (HFHS diet) and a widely used pharmaceutical intervention (metformin). Chow (control) or high fat high sugar (HFHS) diet started at 6 weeks of age.. At six months of age, a subset of mice from each chow group and of each sex were euthanized for tissue collection.
At 7 months of age, half of the remaining mice in each diet treatment group will receive metformin treatment.  In a subset of the 12 month cohort animals, metformin (5 mg/mL) was delivered ad libitum in the drinking water (Chang, Moro et al. 2018). Terminal phenotyping and tissue collection was done at 12 months of age.

overall_design: Type 2 Diabetes relevant phenotypes across 20 CC-RIX genetic backgrounds, two sexes, and metformin intervention.

first: Candice

last: Baker


title: Effects of CDK7 inhibition on fibrotic gene expression

city: South San Francisco

first: Jason

middle: Anthony

last: Vander Heiden

state: CA

address: 1 DNA Way

department: Biological Research AI Development

institute: Genentech

summary: Targeted CDK7, a cyclin-dependent kinase with dual functionality in both cell cycle and transcription, by small molecule inhibitors may attenuate fibrotic response both in vivo and in vitro. However, the exact role of CDK7 in this pathogenic process remains quite elusive. Herein, we want to address this MOA question using RNA-seq. In this RNA-seq experiment we set out to determine (1) the role of CDK7 in fibroblast gene expression and (2) the role of CDK7 in TGF beta-driven gene expression in normal human lung fibroblasts.

overall_design: Triplicate cultures of normal human lung fibroblasts were treated with DMSO only (0.10%), the CDK7 inhibitor THZ1 (50nM), TGFb (10ng/mL), or combined TGFb and THZ1 (50nM).

first: Ning

last: Ding

first: Hsiao-Yen

last: Ma

first: Soren

last: Muller


title: Virulence regulates and boosts CRISPR-Cas9 immunity in Group B Streptococcus [RNA-seq]

status: Public on Nov 04 2024

city: Paris

first: Rachel

last: Legendre

address: 28, rue du docteur Roux

department: Research and Resource Center for Scientific Informatics

institute: Institut Pasteur

summary: Bacterial CRISPR-Cas9 immune systems protect against foreign DNA. However, immune efficiency is constrained by Cas9 off-target effects and toxicity. Here, we demonstrate that CRISPR-Cas9 immunity is regulated by CovR, the major regulator of virulence in Group B Streptococcus, a pathobiont responsible for neonatal invasive infections. We show that CovR binds to and represses a distal promoter of the cas operon, embedding immunity in the virulence regulatory network. Releasing CovR repression enhances immune efficiency against suboptimal spacers, originating from old immune memory or mutations, thereby transiently expanding the sequence space recognized and cleaved by Cas9. Furthermore, CovR inactivation promotes the acquisition of new spacers, enhancing immune memory. CovR-mediated immune regulation is conserved at the species level, with lineage-specific variability in the constitutive cas promoter and Cas9 variants, suggesting different evolutionary trajectories. Overall, we describe a coordinated regulatory mechanism between immunity and virulence that enhances the immune repertoire during acute infection phases.

overall_design:
To investigate the regulation of CRISPR-cas9 transcrition, we generated covR deletion mutant (∆covR) in three WT strains: NEM316, 2603V/R, and A909RF belonging to different clonal complexes. We then performed differential gene expression analysis by RNA-seq between the wild-type parental strain and the corresponding ∆covR mutant.
RNA are purified from exponentialy growing cultures (OD600 = 0.5) in rich media (THY) incubated at 37°C in static condition. Biological triplicate (Replicate 1, 2, and 3) are done on different days.

first: Maria-Vittoria

last: MAZZUOLI

first: Elise

last: JACQUEMET

last: LEGENDRE

first: Odile

last: SISMEIRO

first: Arnaud

last: FIRON


title: Virulence regulates and boosts CRISPR-Cas9 immunity in Group B Streptococcus [CrispRseq]

sample_organism:
Escherichia coli
Streptococcus agalactiae

overall_design:
The experiment investigates how single mismatch in the protospacer affect immunity in the BM110 wild-type strain and in the isogenic ∆covR mutant. We first constructed in vitro a vector collection (Input pool) containing the protospacer  4 or 8 with single mutation at each base position with all four nucleotide options (A, T, G, C). We then transformed the pool of vector into GBS and selected for kanamycin resistant  clones. Transformants were pooled and plasmid purified (Output pools).
Purified pools of vectors were used as matrix to amplify by PCR the protospacers. The primers included a common annealing sequence  and an unique identifiers (index) for multiplex sequencing. Amplicon (197 bp) were pooled and Illumina adapters added (PCR-free). After amplicon sequencing, Illumina adapters were removed and amplicon demultiplexed.
PCR amplification and sequencing were done starting from 3 independent plasmid purification of the input pool (before and after long term -80°C storage). This should be considered as technical replicate.
As a control, the input pool was transformed into a double mutant ∆covR ∆P2cas in which the immune system is inactive. A single experiment was done.
Input pool were transformed indepently two times in the WT and in the ∆covR mutant to give two biological duplicate of the corresponding output pools.


title: Virulence regulates and boosts CRISPR-Cas9 immunity in Group B Streptococcus.

platform_organism:
Escherichia coli
Streptococcus agalactiae


title: LSK immunophenotype hematopoietic stem cells after irradiation at day points in 3 weeks

city: 重庆Chongqing

first: Zeyu

last: Fu

state: 重庆Chongqing

address: 高滩岩正街

department: College of Preventive Medicine

institute: AMU

summary: In this study, we aimed to investigate the differentiation dynamics of LSK (Lin-Sca1+c-Kit+) cells following radiation injury. LSK cells, which are a critical component of the hematopoietic stem and progenitor cell population, play a vital role in the regeneration and repair of the hematopoietic system after damage. By utilizing these cells, we sought to understand the mechanisms underlying their response to radiation, focusing on their capacity for self-renewal and differentiation into various blood cell lineages.

overall_design: To perform 10X Genomics single-cell sequencing on mouse bone marrow LSK cells, begin by euthanizing the mouse following ethical guidelines and extracting the femurs and tibias. Flush the bone marrow using a syringe with a suitable buffer, such as PBS with 2% FBS, to obtain a single-cell suspension. Filter this suspension through a 40 µm cell strainer to remove debris and clumps. Next, enrich the LSK (Lineage^-, Sca-1^+, c-Kit^+) cells using magnetic-activated cell sorting (MACS) or fluorescence-activated cell sorting (FACS). Count the cells and assess viability using trypan blue staining or a similar method. Adjust the concentration of LSK cells to the required density, typically 700-1200 cells/µL, for optimal performance in the 10X Genomics Chromium Controller. Load the single-cell suspension, reagents, and gel beads into the appropriate wells of a 10X Genomics Single Cell 3' Chip and run the chip in the Chromium Controller to encapsulate single cells into nanoliter-scale Gel Bead-In-EMulsions (GEMs). Perform reverse transcription within the GEMs to generate barcoded cDNA from the mRNA of individual cells, then break the emulsion and clean up the barcoded cDNA. Amplify the cDNA via PCR and assess the quality and quantity using an Agilent Bioanalyzer or similar instrument. Construct sequencing libraries from the amplified cDNA, following the 10X Genomics protocol. Sequence the libraries on an Illumina platform, such as NovaSeq or NextSeq, according to the recommended read length and depth for single-cell RNA-seq. Finally, use the 10X Genomics Cell Ranger pipeline to process the raw sequencing data, perform alignment, and generate a gene expression matrix. Analyze the data using bioinformatics tools to identify cell populations, gene expression patterns, and potential changes in cell fate or differentiation pathways.

first: Song

last: Wang

first: Shilei

last: Chen

first: Xinhai

last: Yang

first: Junping


title: Transient gene melting governs the timing of oligodendrocyte maturation [CUT&RUN]

status: Public on Nov 02 2024

city: Cleveland

first: Paul

last: Tesar

state: Ohio

address: 10900 Euclid Ave.

department: Genetics & Genome Sciences

institute: Case Western Reserve University

summary: Cellular maturation is a crucial step for tissue formation and function, distinct from the initial steps of differentiation and cell fate specification. In the central nervous system, failure of oligodendrocyte maturation is linked to diseases such as multiple sclerosis. Here, we report a transcriptional mechanism that governs the timing of oligodendrocyte maturation. After progenitor cells differentiate into immature oligodendrocytes, the transcription factor SOX6 redistributes from super enhancers to cluster across specific gene bodies. These sites exhibit extensive chromatin decondensation and transcription, which abruptly turn off upon maturation. Suppression of SOX6 deactivates these immaturity loci, accelerating the transition to mature, myelinating oligodendrocytes. Notably, cells harboring this immature SOX6 gene signature are enriched in multiple sclerosis patient brains. Antisense oligonucleotide-mediated Sox6 knockdown drives precocious oligodendrocyte maturation in mice. Our findings establish SOX6 as a key regulator of oligodendrocyte maturation and highlight its potential as a therapeutic target to promote myelination in disease.

overall_design: CUT & RUN of pluripotent stem cell derived oligodendrocyte progenitor cells (OPCs) and oligodendrocytes (OLs)

first: Jesse

middle: J

last: Zhan

first: Erin

middle: F

last: Cohn

first: Kevin

last: Allan


title: Divergence from the human astrocyte developmental trajectory in glioblastoma

status: Public on Oct 31 2024

city: Atlanta

first: Steven

last: Sloan

state: Georgia

address: 615 Michael Street, Room 375

institute: Emory University

summary: Glioblastoma (GBM) is defined by heterogenous and resilient cell populations that closely reflect neurodevelopmental cell types. While it is clear that GBM echoes early and immature cell states, identifying the specific developmental programs disrupted in these tumors has been hindered by a lack of high-resolution trajectories of glial and neuronal lineages. Here, we delineate the course of human astrocyte maturation to uncover discrete developmental stages and attributes mirrored by GBM. We generated a transcriptomic and epigenomic map of human astrocyte maturation using cortical organoids maintained in culture for nearly two years. Through this approach, we chronicled a multi-phase developmental process orchestrated by a series of transcription factor and gene regulatory networks. Our time course of human astrocyte maturation includes a novel and molecularly distinct intermediate period that serves as a lineage commitment checkpoint upstream of mature quiescence. This intermediate stage acts as a site of developmental deviation separating IDH-wildtype neoplastic astrocyte lineage cells from quiescent astrocyte populations. Interestingly, IDH1-mutant tumor astrocyte lineage cells are the exception to this developmental perturbation, where immature properties are suppressed as a result of D2HG oncometabolite exposure. We propose that this defiance is likely a consequence of IDH1mt-associated epigenetic dysregulation and we identified biased DNA hydroxymethylation (5hmC) in maturation genes as a possible mechanism. Together, this study illustrates a novel cellular state aberration in GBM astrocyte lineage cells and presents new developmental targets for experimental and therapeutic exploration.

overall_design: This study consists of multiple sequencing datasets. (1) We purified astrocytes from fresh human cortical organoids and GBM tumor and margin tissue samples for bulk RNA-seq and ATAC-seq. (2) Nuclei were isolated from frozen GBM tumor and margin tissue samples for single-nucleus multiome (ATAC- and RNA-seq) sequencing. (3) We performed MeDIP and hMe-Seal with flash-frozen tissue from IDH1mt and IDHwt tumors (with corresponding margin samples) for 5mC and 5hmC analyses. (4) Bulk RNA-seq data for ASCL1 overexpression experiments and D2HG functional experiments with human fetal astrocytes.

first: Sojka

last: Caitlin

first: Wang

last: Hsiao-Lin V.

first: Bhatia

last: Tarun

first: Li

last: Yangping

first: Chopra

last: Pankaj

first: Sing

last: Anson

first: Voss

last: Anna

first: King

last: Alexia

last: Feng

first: Joseph

last: Kevin

first: Ravi

last: Vidhya

first: Olson

last: Jeffrey

first: Hoang

last: Kimberly

first: Nduom

last: Edjah

first: Corces

last: Victor

first: Yao

last: Bing

first: Sloan

last: Steven


title: Transcriptomic profiling across 20 CC-RIX genetic backgrounds with and without high-fat high-sugar (HFHS) diet, and with and without metformin [adipose]

first: Gary

last: Churchill

first: Candice

middle: N

last: Baker


title: A histone hyperacetylation pulse induces cellular memory of gene expression and 3-dimensional genome folding

city: Montpellier

first: Marco

last: Di Stefano

address: 141 rue de la Cardonille

summary: Epigenetic memory enables the stable propagation of gene expression patterns in response to transient developmental and environmental stimuli. Although 3-dimensional (3D) organisation is emerging as a key regulator of genome function due its role in gene expression control, it is unknown whether it contributes to cellular memory. Here, we establish that acute perturbation of the epigenome can induce cellular memory of gene expression in mouse embryonic stem cells (mESCs). Specifically, we uncover how a pulse of histone deacetylase inhibition translates to changes in the histone acetylation and methylation landscape, as well as global and local genome folding. While most epigenomic and transcriptional changes are initially reversed once the perturbation is removed, architectural features are not fully reestablished. Upon a second transient pulse of hyperacetylation, hundreds of genes maintain their dysregulated state. Using ultra-deep Micro-C, we associate memory of gene expression with enhancer-promoter contacts and repressive chromatin topology mediated by Polycomb. These results uncover the complex interplay between different epigenetic regulatory layers and establish a novel link between genome folding and cellular memory.

overall_design: ChIP-seq experiments in mouse ESCs in control (DMSO), TSA-treated or 24-hour recovery (REC), sequential TSA (reTSA) and recovery (reTSA) conditions. ChIP-seq experiments in PCGF4-/- PCGF2fl/fl cell line in UT-DMSO, OHT-DMSO, OHT-TSA and OHT-REC conditions.

first: Flora

last: Paldi

first: Michael Florian

last: Szalay

first: Solene

last: Dufau

first: Hadrien

last: Reboul

first: Giacomo

last: Cavalli


title: Transcriptomic profiling across 20 CC-RIX genetic backgrounds with and without high-fat high-sugar (HFHS) diet, and with and without metformin


title: Divergence from the human astrocyte developmental trajectory in glioblastoma [MeDIP-seq]

first: Steven

last: Sloan

first: Caitlin

last: Sojka

first: Hsiao-Lin

middle: V

last: Wang

first: Tarun

last: Bhatia

first: Yangping

last: Li

first: Pankaj

last: Chopra

first: Anson

last: Sing

first: Anna

last: Voss

first: Alexia

last: King

first: Feng

first: Kevin

last: Joseph

first: Vidhya

last: Ravi

first: Jeffrey

last: Olson

first: Kimberly

last: Hoang

first: Edjah

last: Nduom

first: Victor

last: Corces

first: Bing

last: Yao


title: Transcriptomic profiling across 20 CC-RIX genetic backgrounds with and without high-fat high-sugar (HFHS) diet, and with and without metformin [skeletal muscle]

first: Gary

last: Churchill

first: Candice

last: Baker


title: VIRMA-mediated m6A modification regulates forebrain formation through facilitating ribosome biogenesis

status: Public on Oct 29 2024

city: Shenzhen

first: Tao

last: Zhou

address: Shenzhen Institutes of Advance Technology, 1068 Xueyuan Avenue, Shenzhen University Town, Nanshan District, Shenzhen

institute: The Brain Cognition and Brain Disease Institute of Shenzhen Institute of Advanced Technology, Chinese Academy of Sciences

summary: N6-methyladenosine (m6A) modification plays crucial roles in tissue development and homeostasis. However, the mechanisms underlying cellular adaptation of m6A modification and their impact on protein synthesis machinery remain unclear. VIRMA, the largest and evolutionarily conserved core of the m6A methyltransferase complex, is highly expressed in the embryonic brain and various cancers. Here, we demonstrate that VIRMA-mediated m6A modification is essential for active ribosome biogenesis. VIRMA depletion destabilizes the entire writer complex and reduces m6A levels, leading to decreased proliferation and increased apoptosis of neural progenitor/stem cells (NPCs), ultimately causing severe forebrain developmental defects. Mechanistically, VIRMA depletion impairs ribosome biogenesis by inhibiting mRNA decay, triggering a p53-dependent stress response and compromising global protein synthesis. Importantly, these findings extend to human cancer cells, suggesting a potential conservation of this mechanism. Overall, our study reveals the critical role of m6A in adapting protein synthesis machinery during brain development and potentially in cancer.

overall_design: We perform high-throughput RNA sequencing of cultured neural progenitor/stem cells (NPCs) with or without deletion of VIRMA to profile RNA half-lives. Wiltype (WT) and conditional knockout (cKO) NPCs were treated with actinomycin-D (ActD) to halt new RNA synthesis. At 0, 3, and 6 hours post-treatment, cells were collected for total RNA extraction followed by RNA-seq. Four replicates were conducted for each genotype at each timepoint.

first: Min

last: Wu

first: Bin

last: Shen

first: Haifeng

last: Sun

first: Xiaoli


city: Nanjing City

state: JiangSu province

address: Tianyuan East Road 818

institute: Nanjing Medical University

overall_design: We performed high-throughput sequencing of mouse developing forebrain (E13.5) with or without deletion of VIRMA to profile m6A methylated RNA and RNA expression. Both wildtype (WT) and conditional homozygous knockout (cKO) conditions were assayed in four biological replicates each.


title: High-resolution Hi-C reveals increased chromatin looping with senescence associated with hypomethylation and retrotransposon derepression [Long Read DNA-seq]

status: Public on Oct 30 2024

city: Providence

first: Nicola

last: Neretti

state: RI

address: 225 Dyer St.

institute: Brown University

summary: This study presents the highest-resolution chromatin map of cellular senescence to date, shedding light on how genomic architecture is altered with this damaging phenotype. Senescence, a driver of aging, is a pro-inflammatory state of proliferative arrest caused by DNA damage; it is associated with epigenetic changes, including those to chromatin organization. We created ~3kb Hi-C contact maps of proliferating, quiescent, and replicative senescent lung fibroblasts, and also compared these to oncogene-induced senescence. Our findings confirm a loss of heterochromatin, with a shift towards the A compartment and A subcompartments. We establish a novel loop analysis framework, revealing the ~six times more unique loops with senescence, which lose methylation at their anchors. Additionally, we present a custom long-read reference genome highlighting structural changes supporting retrotransposon derepression, particularly at a defined ‘hotspot’. These architectural changes contribute to senescence, as they promote cell cycle arrest and inflammation, as well as epigenetic drift.

overall_design: To investigate the role of genomic structure in cellular senescence we generated Hi-C data, total RNA-seq, and long-read DNA sequencing for proliferating and late replicative senescent human diploid lung fibroblasts (LF1). We also generated Hi-C and total RNA-seq for quiescent cells. We compared the proliferating, quiescent, and senescent cell data to determine characteristic architectural features of senescence.

first: Audrey

last: Dalgarno

first: Shane

last: Evans

first: Maxfield

last: Kelsey

first: Thomas

last: Nunez

first: Azucena

last: Rocha

first: Kelly

last: Clark

first: John

middle: M

last: Sedivy


title: Divergence from the human astrocyte developmental trajectory in glioblastoma [bulk RNA]

first: Steven

last: Sloan

overall_design:
This study consists of multiple sequencing datasets. (1) We purified astrocytes from fresh human cortical organoids and GBM tumor and margin tissue samples for bulk RNA-seq and ATAC-seq. (2) Nuclei were isolated from frozen GBM tumor and margin tissue samples for single-nucleus multiome (ATAC- and RNA-seq) sequencing. (3) We performed MeDIP and hMe-Seal with flash-frozen tissue from IDH1mt and IDHwt tumors (with corresponding margin samples) for 5mC and 5hmC analyses. (4) Bulk RNA-seq data for ASCL1 overexpression experiments and D2HG functional experiments with human fetal astrocytes.

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GSM8492612-GSM8492624 missing raw data files
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first: Caitlin

last: Sojka

first: Hsiao-Lin

last: Wang

first: Tarun

last: Bhatia

first: Yangping

last: Li

first: Pankaj

last: Chopra

first: Anson

last: Sing

first: Anna

last: Voss

first: Alexia

last: King

first: Feng

first: Kevin

last: Joseph

first: Vidhya

last: Ravi

first: Jeffrey

last: Olson

first: Kimberly

last: Hoang

first: Edjah

last: Nduom

first: Victor


title: Decreased mRNA expression of malonyl-CoA decarboxylase emerges from an inborn error-like defect in the Mlycd gene of DBA/2J mice

status: Public on Oct 26 2024

city: New York

first: Sander

last: Houten

state: NY

address: 1425 Madison Avenue, Box 1498

department: Genetics and Genomic Sciences

institute: Icahn School of Medicine at Mount Sinai

summary: Mlycd encodes malonyl-CoA decarboxylase (MCD), which is an enzyme that localizes in the cytosolic, mitochondrial, and peroxisomal compartments and catalyzes the conversion of malonyl-CoA into acetyl-CoA. Malonyl-CoA can be converted into malonylcarnitine (C3DC). Patients with an autosomal recessive defect of MCD and MCD KO mice have pronounced elevations of C3DC. Analysis of plasma C3DC levels in the BxD genetic reference population revealed increased levels in BxD strains that harbor the DBA/2J haplotype at the site of the Mlycd gene. RNA sequencing was performed on two samples of DBA/2J mouse livers and two C57BL/6J mouse livers. Decreased expression of Mlycd gene as well as intronic reads in intron 2 were observed in DBA/2J livers. Long-read sequecing of DBA/2J livers in the Mlycd region confirmed an intracisternal A-particle (IAP) retrotransposon in intron 2 of the DBA/2J Mlycd sequence. To confirm the causal nature of the variant, DBA/2J mice with and without the C57BL/6J variant of Mlycd spliced in were tested for products of MCD enzymatic activity, and the C57BL/6J variant was able to rescue the phenotype seen in the DBA/2J mice.

overall_design: We analyzed mRNA expression in liver samples from two DBA/2J mice and two C57BL/6J mice using RNA sequencing

first: Carmen

last: Argmann

first: Aaron

last: Bender


title: Spatial transcriptomics in human mid luteal endometrial biopsies

status: Public on Oct 27 2024

first: Jan

last: Brosens

summary: To test if cell states within the endometrium are spatially organized, we performed spatial transcriptomics on 8 mid-luteal phase superficial endometrial biopsies.

overall_design: Endometrial mRNA profiles of 8 endometrial pipelle biopsies were generated by spatial transcriptomics.

first: Joanne

last: Muter

first: Pavle

last: Vrljicak

first: Sascha

last: Ott


title: Divergence from the human astrocyte developmental trajectory in glioblastoma [hMe-Seal-seq]

first: Steven

last: Sloan

first: Caitlin

last: Sojka

first: Hsiao-Lin

last: Wang

first: Tarun

last: Bhatia

first: Yangping

last: Li

first: Pankaj

first: Anson

last: Corces

first: Bing

last: Yao


title: Spatially defined multicellular functional units in colorectal cancer revealed from single cell and spatial transcriptomics - Slide-Seq

status: Public on Oct 18 2024

city: Cambridge

first: Simon

last: Mages

address: 415 Main St.

institute: Broad Institute

summary: While advances in single cell genomics have helped to chart the cellular components of tumor ecosystems, it has been more challenging to characterize their specific spatial organization and functional interactions. Here, we combine single cell RNA-seq, spatial transcriptomics by Slide-seq, and in situ multiplex RNA analysis, to create a detailed spatial map of healthy and dysplastic colon cellular ecosystems and their association with disease progression. We profiled inducible genetic CRC mouse models that recapitulate key features of human CRC, assigned cell types and epithelial expression programs to spatial tissue locations in tumors, and computationally used them to identify the regional features spanning different cells in the same spatial niche. We find that tumors were organized in cellular neighborhoods, each with a distinct composition of cell subtypes, expression programs, and local cellular interactions. Comparing to scRNA-seq and Slide-seq data from human CRC, we find that both cell composition and layout features were conserved between the species, with mouse neighborhoods correlating with malignancy and clinical outcome in human patient tumors, highlighting the relevance of our findings to human disease. Our work offers a comprehensive framework that is applicable across various tissues, tumors, and disease conditions, with tools for the extrapolation of findings from experimental mouse models to human diseases.

overall_design: Mouse colons were flashed with cold PBS and a segment that includes the lesion and surrounding tissue (or a respective healthy segment from normal mice) was dissected. Samples were then mounted in cold OCT, flash frozen on dry ice covered with ETOH until and long-term stored in -80℃. 10microm sections were cut and the Slide-seq V2 protocol was used as previously described. Four and six arrays were collected from normal colons and premalignant lesions respectively. The muscularis was fit onto the array of both healthy and dysplastic lesions to allow appropriate orientation.

first: Inbal

last: Avraham-Davidi

first: Leah

last: Caplan

first: Evan

last: Murray


title: Spatially defined multicellular functional units in colorectal cancer revealed from single cell and spatial transcriptomics - scRNAseq v2 with cell multiplexing

overall_design: Single-cell suspensions from healthy colon or dysplastic lesions were processed to generate single cell suspensions. All cell suspensions were counted to achieve a uniform concentration of 7,000 cells per microliter  before pooling for capture  by 10x Chromium controller following the manufacturer protocol for the v2 3’ kit (10X Genomics, Pleasanton, CA). antibody_index_XXX.csv gives the mapping from HTOs to samples

first: Danielle

last: Dionne

first: Toni

last: Delorey


title: Spatially defined multicellular functional units in colorectal cancer revealed from single cell and spatial transcriptomics - scRNAseq v3 with cell multiplexing

overall_design: Single-cell suspensions from healthy colon or dysplastic lesions were processed to generate single cell suspensions. All cell suspensions were counted to achieve a uniform concentration of 7,000 cells per microliter  before pooling for capture  by 10x Chromium controller following the manufacturer protocol for the v3 3’ kit (10X Genomics, Pleasanton, CA). antibody_index_XXX.csv gives the mapping from HTOs to samples


title: Divergence from the human astrocyte developmental trajectory in glioblastoma [bulk ATAC-seq]

last: Chopra

last: Sing

first: Anna

last: Voss

first: Alexia

last: King

first: Feng

first: Kevin

last: Joseph

first: Vidhya

last: Ravi

first: Jeffrey

last: Olson

first: Kimberly

last: Hoang

first: Edjah

last: Nduom

first: Victor


title: edu-Ψ-seq Reveals snoRNA-guided mRNA Pseudouridylation that Regulates Protein Synthesis [CLIP-seq]

city: Guangzhou

first: Jian-Hua

last: Yang

address: No. 135, Xingang Xi Road

department: School of Life Sciences

institute: Sun Yat-sen University

summary:
Purpose:Pseudouridine (Ψ) is the most abundant RNA epigenetic modification and plays vital roles in various biological processes. However, its biogenesis and functions in mRNAs remain elusive.
Method:Here, we developed an approach for enriching Ψ sites with deep sequencing (edu-Ψ-seq), which utilizes both exoribonuclease and restriction endonuclease activity to eliminate other nucleosides or reads without Ψ modifications.
Results:Using edu-Ψ-seq approach in the dyskerin pseudouridine synthase 1 (DKC1) knockdown cells, we identified that more than 100 pseudouridines could be catalyzed by DKC1 in human mRNAs and non-coding RNAs (ncRNAs). Surprisingly, we discovered, for the first time, 10 pseudouridines in mRNAs might be guided by 11 small nucleolar ncRNAs (snoRNAs). Interestingly, we found that SNORA10 could guide DKC1 protein to induce the pseudouridylation of RPL15 mRNA. Importantly, loss of SNORA10 impaired the protein synthesis of RPL15 and inhibited cell proliferation.
Conclusion:Overall, our study not only developed a powerful method for detecting Ψ sites but also uncovered another layer of gene expression regulation that involves crosstalk among snoRNAs, mRNA pseudouridylation and protein synthesis.

overall_design: DKC1 CLIP-seq

first: Jianhua

first: Wujian

last: Zheng


title: Single Cell RNAseq REX-1 RFP mESC 2019

status: Public on Oct 16 2024

city: Detroit

first: Daniel

last: Rappolee

state: MI

address: 275 E Hancock St

department: OBGYN

institute: Wayne State University

summary:
This experiment examines how increasing stress (sorbitol) can override normal stemness (NS) conditions, which LIF maintains, causing lineage imbalance and otherwise affecting cells in the early embryo.
It also compares NS to normal differentiation (ND) (without LIF).
D3 are from 8 blastocysts of 129/Sv+/+ strain mice germ line competent 1. No sex is given as these may actually be a product of different sexes of the 8 blastocysts. The D3 cells were obtained for ATCC (American Type Culture Collection).
1. Doetschman TC, Eistetter H, Katz M, Schmidt W, Kemler R. The in vitro development of blastocyst-derived embryonic stem cell lines: formation of visceral yolk sac, blood islands and myocardium. J Embryol Exp Morphol 1985;87:27-45.
Rex1 promoter- Red fluorescence protein (RFP) transgenic viable reporter ESC were created and validated previously2.
2. Li Q, Gomez-Lopez N, Drewlo S, et al. Development and Validation of a Rex1-RFP Potency Activity Reporter Assay That Quantifies Stress-Forced Potency Loss in Mouse Embryonic Stem Cells. Stem Cells Dev 2016;25:320-8.

overall_design: Single Cell RNAseq of Rex1 promoter- Red Fluorescent Protein (RFP) mESC cultured for 72hr with different stimuli. Single replicates.

first: Ximena

middle: L

last: Ruden

first: Mohammed

last: Abdulhasan

first: Katherine

last: Gurdziel

first: Hao

last: Feng

first: Wen

last: Tang