title: The lipopeptide Pam3CSK4 inhibits Rift Valley fever virus infection and protects from encephalitis

city: Philadelphia

first: Trevor

last: Griesman

state: PA

address: 459 Stemmler hall, 3450 Hamilton Walk

department: Pathology and laboratory medicine

institute: University of Pennsylvania

summary: Rift Valley fever virus (RVFV) is an encephalitic bunyavirus that can infect neurons in the brain. There are no approved therapeutics that can protect from RVFV encephalitis. Innate immunity, the first line of defense against infection, canonically antagonizes viruses through interferon signaling. We found that interferons did not efficiently protect primary cortical neurons from RVFV, unlike other cell types. To identify alternative neuronal antiviral pathways, we screened innate immune ligands and discovered that the TLR2 ligand Pam3CSK4 inhibited RVFV infection, and other bunyaviruses. Mechanistically, we found that Pam3CSK4 blocks viral fusion, independent of TLR2. In a mouse model of RVFV encephalitis, Pam3CSK4 treatment protected animals from infection and mortality. Overall, Pam3CSK4 is a bunyavirus fusion inhibitor active in primary neurons and the brain, representing a new approach toward the development of treatments for encephalitic bunyavirus infections.

overall_design:
To define the neuronal response to TLR2 stimulation, we performed RNAseq on rat cortical neurons (7 days ex vivo) stimulated with vehicle, Pam3CSK4 (10ug/mL), or Pam2CSK4 (10ug/mL) for 6 hours. Three replicates per condition.
We then used differential gene analysis to compare genes induced by Pam3CSK4 or Pam2CSK4 relative to vehicle

first: Cynthia

last: McMillen

first: Kanupriya

last: Whig

first: Seble Getenet

last: Negatu

first: Lenka

last: Dohnalová

first: Mark

last: Dittmar

first: Jesse

middle: J

last: Hulahan

first: Christoph

last: Thaiss

first: Kellie

last: Jurado

first: David

last: Schultz

first: Amy

last: Hartman

first: Sara

last: Cherry


title: Role of Yap/Taz in mouse embryonic stem cells

status: Public on Jun 18 2024

city: Muenster

first: Hyun-Woo

last: Jeong

state: NRW

department: Dept. Tissue Morphogenesis

country: Germany

institute: Max-Planck-Institute for Molecular Biomedicine

type: Expression profiling by high throughput sequencing

summary: Here, we show that the loss of Yap/Taz function does not affect the undifferentiated status of the ESCs but these factors play a pivotal role in the maintenance of the pluripotent cells by keeping the process of apoptosis at bay.

overall_design: RNA-seq analysis of wildtype, Taz knockout, and Yap/Taz double knockout ESCs

first: Rui

last: Chen

last: Fan

first: Fei

first: Niraimathi

last: Govindasamy

first: Martin

last: Stehling

first: Ralf

middle: H

last: Adams

first: Ivan

last: Bedzhov


title: STAG2 Mutations Reshape the Cohesin-Structured Spatial Chromatin Architecture to Drive Gene Regulation in Acute myeloid Leukemia [ChIP-Seq]

city: Regensburg

first: Alexander

last: Fischer

address: Franz-Josef-Strauß-Allee 11

institute: Leibniz Institute for Immunotherapy

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Cohesin shapes the chromatin architecture, including enhancer-promoter interactions. Its components, especially STAG2 but not its paralogue STAG1, are frequently mutated in myeloid malignancies. To elucidate the underlying mechanisms of leukemogenesis, we comprehensively characterized genetic, epigenetic, transcriptional, and chromatin conformational changes in acute myeloid leukemia (AML) patient samples. Specific loci displayed altered cohesin occupancy, gene expression and local chromatin activation which were not compensated by STAG1. These changes could be linked to disrupted spatial chromatin looping in cohesin-mutated AMLs. We performed complementary depletion of STAG2 or STAG1 in primary human hematopoietic progenitors (HSPCs). We detected effects overlapping STAG2-mutant AML-specific changes following STAG2 knockdown, not invoked by depletion of STAG1. STAG2-deficient HSPCs displayed impaired differentiation capacity and maintained HSPC-like gene expression. This work establishes STAG2 as a key regulator of chromatin contacts, gene expression and differentiation in the hematopoietic system and identifies candidate target genes that may be implicated in human leukemogenesis.

overall_design:
To determine the impact of cohesin loss on genome-wide cohesin subunit (RAD21/STAG2/STAG1) occupancy and distribution as well as to define active enhancer/promotor (H3K27ac) and CTCF sites, we performed ChIP-seq on STAG2-mutant, RAD21-mutant and cohesin wildtype AML patients samples. In addition we aslo used primary CD34+ cord-blood derived HSPC cultures of independent donors electroporated with siRNAs for knockdown targeting either STAG2, STAG1, RAD21. As controls a non-targeting (luciferase-specific) control siRNA (siCtrl) transfected samples were used. HSPCs were additonally used to define Mediator binding sites (MED12) and to assay chromatin-binding of the transcription factor PU.1.

***************************************************************
Due to patient privacy concerns, raw sequencing data of adult AML patients is deposited in the European Genome Archive (EGA) under study number EGAS00001007405. Due to special legal and ethical privacy concerns regarding newborns, raw sequencing data of cord blood HSPC samples cannot be deposited in any online repository.
***************************************************************

first: Margit

last: Nuetzel

first: Hanna

last: Stanewski

first: Ute

last: Ackermann

first: Michael

last: Rehli

first: Ruud

last: Delwel

first: Claudia

last: Gebhard


title: STAG2 Mutations Reshape the Cohesin-Structured Spatial Chromatin Architecture to Drive Gene Regulation in Acute myeloid Leukemia [Hi-C]

overall_design:
We performed HiC-seq on STAG2-mutant, RAD21-mutant and cohesin wildtype AML patients samples to determine the impact of Cohesin mutations on spatial chromatin interactions.For experimental validation, we performed HiC-seq on primary CD34+ cord-blood derived HSPC cultures of independent donors electroporated with siRNAs for knockdown targeting either STAG2, STAG1 or RAD21. As controls non-targeting (luciferase-specific) control siRNA (siCtrl) transfected samples were used.

***************************************************************
Due to patient privacy concerns, raw sequencing data of adult AML patients is deposited in the European Genome Archive (EGA) under study number EGAS00001007405. Due to special legal and ethical privacy concerns regarding newborns, raw sequencing data of cord blood HSPC samples cannot be deposited in any online repository.
***************************************************************

first: Benjamín

last: Hernández-Rodríguez

first: Amadou

last: Niang

first: Noelia

last: Diaz

first: Juan

middle: M

last: Vaquerizas

first: Roger

last: Mulet-Lazaro


title: Targeting parkin-regulated metabolomic change in cartilage in the treatment of osteoarthritis II

city: Shanghai

first: Junjie

last: Gao

address: 600 Yishan Road

country: China

institute: Shanghai Sixth People’s Hospital Affiliated to Shanghai Jiao Tong University School of Medicine

summary: Articular cartilage degeneration, which may lead to osteoarthritis (OA), is a normal component of the aging process, but its underlying mechanism remains unknown. We found that chondrocytes exhibited an energy metabolism shift from glycolysis to oxidative phosphorylation (OXPHOS) during aging. Reprogrammed cartilage metabolism by parkin ablation decreased OXPHOS and increased glycolysis, with ameliorated aging-related OA. Metabolomics analysis indicated that lauroyl-L-carnitine (LLC) was decreased in aged cartilage, but increased in parkin-deficient cartilage. In vitro, LLC improved the cartilage matrix synthesis of OA chondrocytes. In vivo, intraarticular injection of LLC in mice with anterior cruciate ligament transaction (ACLT) ameliorated OA. These results suggest that metabolic changes are regulated by parkin-impaired cartilage during aging, and targeting the metabolomic changes by supplementation with LLC is a promising treatment strategy for ameliorating OA.

overall_design: To investigate the expression change regulated by parkin in articular cartilage

first: Yiyang

last: Ma

first: Changqing

last: Zhang


title: Targeting parkin-regulated metabolomic change in cartilage in the treatment of osteoarthritis I

overall_design: To investigate the expression change of articular cartilage during aging


title: The motility defect caused by absence of the transcriptional regulator LdtR in Sinorhizobium meliloti is restored by mutations in the motility genes motA and motS

city: Blacksburg

first: Birgit

middle: E

last: Scharf

state: VA

address: 970 Washington St SW

department: Biological Sciences

country: USA

institute: Virginia Tech

summary: The flagellar motor is a powerful macromolecular machine used to propel bacteria through various environments. Flagellar motility of the alpha-proteobacterium Sinorhizobium meliloti is nearly abolished in the absence of the transcriptional regulator LdtR, which is involved in peptidoglycan remodeling. We report that LdtR does not regulate motility gene transcription. Remarkably, the motility defects of the DldtR mutant can be restored by secondary mutations in the motility gene motA or a previously uncharacterized gene in the flagellar regulon, which we named motS. MotS is not essential for S. meliloti motility and may serve an accessory role in flagellar motor function. Structural modeling predicts that MotS is comprised of an N-terminal transmembrane segment, a long-disordered region, and a conserved β-sandwich domain. The C-terminus of MotS is localized in the periplasm. Genetics-based substitution of MotA with a MotAG12S variant protein also restored the ΔldtR motility defect. The MotAG12S variant causes a local polarity shift at the periphery of the MotAB stator units. We propose that MotS may be required for optimal alignment of stators in wild-type flagellar motors but becomes detrimental in cells with altered peptidoglycan. Similarly, the polarity shift in the stator units composed of MotB/MotAG12S might stabilize its interaction with altered peptidoglycan.

overall_design:
To investigate the differential gene expression profiles of Sinorhizobium meliloti ΔldtR cells compared to wild-type under motility-promoting conditions, we grew three replicates of each strain (six samples total) in Bromfield overlay plates to early log phase and harvested RNA from equivalent amounts of bacteria.
RNA-seq analysis was performed for both strains (three replicates) and the data used to perform comparative gene expression analysis between the two strains.

first: Richard

middle: C

last: Sobe


title: Epigenetic and transcriptional consequences of chemically induced transposon mobilization in the endosperm (CUT&Tag)

city: Uppsala

last: Santos-González

state: Uppsala

address: Almas Allé 5

department: Department of Plant Biology and Forest Genetics

country: Sweden

institute: SLU

summary: Genomic imprinting, an epigenetic phenomenon leading to parent-of-origin-specific gene expression, has independently evolved in the endosperm of flowering plants and the placenta of mammals—tissues crucial for nurturing embryos. While transposable elements (TEs) frequently colocalize with imprinted genes and are implicated in imprinting establishment, direct investigations of the impact of de novo TE transposition on genomic imprinting remain scarce. In this study, we explored the effects of chemically induced transposition of the Copia element ONSEN on genomic imprinting in Arabidopsis thaliana. Through the combination of chemical TE mobilization and doubled haploid induction, we generated a line with 40 new ONSEN copies. Our findings reveal a preferential targeting of maternally expressed genes (MEGs) for transposition, aligning with the colocalization of H2A.Z and H3K27me3 in MEGs—both previously identified as promoters of ONSEN insertions. Additionally, we demonstrate that chemically-induced DNA hypomethylation induces global transcriptional deregulation in the endosperm, leading to the breakdown of MEG imprinting. This study provides insights into the consequences of chemically induced TE remobilization in the endosperm, underscoring the need for cautious interpretation of the connection between TEs and genomic imprinting.

overall_design: RNAseq, WGBS and CUT&Tag datasets were obtained from INTACT purified endosperm nuclei from Col × Ler (L) reciprocal crosses at 4 days after pollination (DAP).  WGBS and CUT&Tag were generated in duplicates using the INTACT line (referred as C) or TEmob (T3, referred as Cm) genetic background in recirpocal crosses with Ler (L). RNAseq libraries were generated in triplicates from TEmob (Cm) x Ler (L) reciprocal crosses.

first: Gerardo

last: del Toro de Leon

first: Joram

last: van Boven

first: Jiao

last: Wen-Biao

first: Korbinian

last: Schneeberger

last: Köhler


title: MCM2-7 loading-dependent ORC release ensures genome-wide origin licensing

city: London

first: LMS

last: Bioinformatics Core

address: Hammersmith Hospital Campus, Du Cane Road

department: Bioinformatics Core

country: United Kingdom

institute: MRC London Institute of Medical Sciences

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

overall_design: Refer to individual Series


title: STAG2 Mutations Reshape the Cohesin-Structured Spatial Chromatin Architecture to Drive Gene Regulation in Acute myeloid Leukemia [bulk RNA-Seq]

first: Alexander

last: Fischer

overall_design:
We performed total RNAseq on STAG2-mutant, RAD21-mutant and cohesin wildtype AML patients samples to determine the impact of Cohesin mutations on the transcriptome. In addtion total RNAseq was performed on healthy adult CD34+ HSCs to serve as healthy controls for comparison to leukemia samples. To determine the impact of stable genomic STAG2 deletion on the transcriptome of healthy cells, we performed RNA-seq on primary CD34+ cord-blood derived HSPC cultures of independent donors electroporated with sgRNA Cas9 ribonucleprotien complexes  targeting the STAG2 gene (SA2KO). As controls, cells were transfected with a non-targeting guide RNA (NC) Cas9 complex.

***************************************************************
Due to patient privacy concerns, raw sequencing data of adult AML patients is deposited in the European Genome Archive (EGA) under study number EGAS00001007405. Due to special legal and ethical privacy concerns regarding newborns, raw sequencing data of cord blood HSPC samples cannot be deposited in any online repository.
***************************************************************

first: Margit

last: Nuetzel

first: Stanley

last: van Herk

first: Hanna

last: Stanewski

first: Peter

last: Valk

first: Michael

last: Rehli

first: Ruud

last: Delwel

last: Gebhard


title: Opto-seq reveals input-specific immediate early gene induction in ventral tegmental area cell types

city: Seattle

first: Marta

last: Soden

state: WA

address: 1959 NE Pacific St

institute: University of Washington

summary: The ventral tegmental area (VTA) is a critical node in circuits governing motivated behavior and is home to diverse populations of neurons that release dopamine, GABA, glutamate, or combinations of these neurotransmitters. The VTA receives inputs from many brain regions, but a comprehensive understanding of input-specific activation of VTA neuronal subpopulations is lacking. To address this, we combined optogenetic stimulation of select VTA inputs with single-nucleus RNA sequencing (snRNAseq) and highly multiplexed in situ hybridization to identify distinct neuronal clusters and characterize their spatial distribution and activation patterns. Quantification of immediate early gene (IEG) expression revealed that different inputs activated select VTA subpopulations, which demonstrated cell-type specific IEG programs. Within dopaminergic subpopulations IEG induction levels correlated with differential expression of ion channel genes. This new transcriptomics-guided circuit analysis reveals the diversity of VTA activation driven by distinct inputs and provides a resource for future analysis of VTA cell types.

overall_design: To perform transcriptomic analysis of the mouse VTA following optogenetic stimulation of specific inputs, we injected Slc32a1-Cre (Vgat-Cre) mice bilaterally in the LH or NAc, and Slc17a7-Cre (Vglut1-Cre) mice bilaterally in the PFC with a Cre-dependent virus encoding channelrhodopsin (AAV1-FLEX-ChR2-YFP). Control mice were injected with AAV1-FLEX-YFP, and all mice were implanted with bilateral fiber optic cannulas above the VTA. Following recovery from surgery and extensive acclimation to the patchcord, mice received 15 min of 20 Hz blue light stimulation. Immediately after stimulation the VTA was microdissected and snap frozen. VTA tissue from 3-4 mice per group was pooled and nuclei were isolated and sequenced using the 10x Genomics Next GEM platform.

first: Rhiana

last: Simon

first: Mary

last: Loveless

first: Joshua

middle: X

last: Yee

first: Koichi

last: Hashikawa

first: Garret

middle: D

last: Stuber

first: Larry

middle: S

last: Zweifel


title: A HDAC4-HDAC2 complex composes an epigenetic sensor that links the DNA damage response to the senescent program by erasing H2BK120 acetylation [II]

city: Udine

first: Emiliano

last: Dalla

state: UD

address: Piazzale Kolbe 4

department: Department of Medicine

country: Italy

institute: Università degli Studi di Udine

summary:
Purpose: Identification of SEs in time course after HDAC4 depletion and correlation to the accumulation of DNA damage.
Outcome:  We discovered a novel epigenetic complex formed by HDAC4 and HDAC2 that is involved in the erasure of H2BK120 acetylation. The HDAC4/HDAC2 complex, through the dynamic deacetylation of H2BK120, modulates the efficiency of DNA repair by homologous recombination (HR) by controlling the recruitment of BRCA1 and CtIP to the site of lesions. Degradation of HDAC4 during senescence has a dual effect. First, it contributes to super-enhancers activation by limiting HDAC3 activity, and second, it causes the accumulation of damaged DNA.

overall_design: Identification of SEs (H3K27ac and H3K4me1 co-presence) in SK-LMS-1 Leiomyosarcomas cells deleted for HDAC4 using CRISPR/Cas9 and re-expressing a tamoxifen-inducible PAM mutant version of HDAC4 (HDAC4-/-/HDAC4-/-PAM-ER) at 0, 12 and 36h from HDAC4 depletion.

first: Claudio

last: Brancolini

first: Eros

last: Di Giorgio


title: A HDAC4-HDAC2 complex composes an epigenetic sensor that links the DNA damage response to the senescent program by erasing H2BK120 acetylation [I]

summary:
Purpose: Recognize the specific H2BK120ac profile around HDAC4 binding sites in proliferating, pre-senescent, senescent and senescence escaped cells during the accumulation of dsDNA damage, compared to undamaged regions.
Outcome:  We discovered a novel epigenetic complex formed by HDAC4 and HDAC2 that is involved in monitoring H2BK120 acetylation. The HDAC4/HDAC2 complex, through the dynamic deacetylation of H2BK120, modulates the efficiency of DNA repair by homologous recombination (HR) by controlling the recruitment of BRCA1 and CtIP to the site of lesions. Degradation of HDAC4 during senescence has a dual effect. First, it contributes to super-enhancers activation by limiting HDAC3 activity, and second, it causes the accumulation of DNA damage.

overall_design: Evaluation of H2BK120 and H4K16 acetylation, H2AX phosphorylation (gammaH2AX) and of HDAC4 binding in BJ-hTERT or hTERT/ HRAS-ER or hTERT/ HRAS-ER /HDAC4 cells at 2 or 8 days from 4-OHT treatment (to induce HRAS G12V).

first: C

first: E


title: A HDAC4-HDAC2 complex composes an epigenetic sensor that links the DNA damage response to the senescent program by erasing H2BK120 acetylation


title: Anti-cancer effects of 7-oxabicyclo[2.2.1]heptene sulfonamide (OBHSA) derivative 17e on gene expression of breast cancer T47D cells

city: Wuhan

first: Jian

last: Huang

address: Bayi Road 299

institute: Wuhan University

summary:
To investigate the anti-cancer effects of PROTAC-like OBHSA derivative  17e, we exposed estrogen receptor alpha positive (ERα+) breast cancer cell line T47D to compound 17e at 10 μM for 12 hours .
We then performed gene expression profiling analysis using data obtained from RNA-seq of T47D cells treated with DMSO or 17e (10 μM) in three replicates.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for T47D cells treated with DMSO (control) and T47D treated with 10 μM 17e (perturbation).


title: Species-specific progenitor domain extends neurogenesis and increases motor neuron production

status: Public on Jun 17 2024

city: New York

first: Sumin

last: Jang

state: NY

institute: Columbia University

summary: Human and mouse cells undergoing in vitro motor neurogenesis were sampled at high temporal resolution to identify species' differences in the progenitor pools that give rise to disproportionate timescale changes of motor neurogenesis between human and mouse.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Human iPS cells and mouse ES cells were subjected to in vitro motor neuron differentiation, which recapitulates species-specific timescales of motor neurogenesis. Cells were sampled every 24 (mouse) and 48 (human) hours during neurogenesis and profiled using single-cell RNA-seq (3').

first: Hynek

last: Wichterle


title: Reorganization of H3K9me heterochromatin leads the neuronal impairment via the cascading destruction of KDM3B-centered epigenomic network

city: Seoul

first: Jung-Woong

last: Kim

address: 84 Heuksuk-ro Dongjak-ku

department: Life Science

country: South Korea

institute: Chung-Ang University

summary: Histone H3K9 methylated heterochromatin silences repetitive non-coding sequences and lineage-specific genes during development, but how tissue-specific genes escape from heterochromatin in differentiated cells is unclear. Here, we examine age-dependent transcriptomic profiling of terminally differentiated mouse retina to identify epigenetic regulators involved in heterochromatin reorganization. The single-cell RNA sequencing analysis reveal a gradual downregulation of Kdm3b in cone photoreceptors during aging. Disruption of Kdm3b (Kdm3b+/-) of 12-month-old mouse retina leads to the decreasing number of cones via apoptosis, and it changes the morphology of cone ribbon synapses. Integration of transcriptome with epigenomic analysis in Kdm3b+/- retinas demonstrate gains of heterochromatin feature in synapse assembly and vesicle transport genes that are downregulated via the accumulation of H3K9me1/2. Contrary, losses of heterochromatin in apoptotic genes exacerbated retinal neurodegeneration. We propose that KDM3B-centered epigenomic network is crucial for balancing of cone photoreceptor homeostasis via the modulation of gene-set specific heterochromatin features during aging.

overall_design: We compared gene expression profiles between mouse retina of wild type and Kdm3b heterozygous. ChIP-seq for H3K9me1, H3K9me2 and H3K9me3 were performed in mouse retina between wild type and heterozygous.


title: An olfactory bulb GnRH neuronal population translates social relevant odors into reproductive behavior in male mice

city: Cologne

first: Sophie

last: Steculorum

address: Gleueler Straße 50

department: Metabolism Research

institute: Max Planck Institute

summary: Hypothalamic gonadotropin-releasing hormone (GnRH) neurons are central regulators of fertility and integrate endogenous hormonal status with environmental cues to ensure reproductive success. Here, we found that extra-hypothalamic GnRH neurons in the olfactory bulb of adult mice and humans (GnRHOB) can mediate social recognition. We show that GnRHOB neurons extend neurites into the vomeronasal organ and olfactory epithelium and project to the hypothalamic median eminence. We demonstrate that male GnRHOB neurons express vomeronasal and olfactory receptors, are activated by female odors in vivo, and mediate gonadotropin release in response to female urine. We find that male preference for female odors is enhanced upon chemogenetic activation of GnRHOB neurons and is impaired after genetic inhibition or ablation of these cells and relies on GnRH signaling in the posterodorsal medial amygdala. Taken together, these results establish GnRHOB neurons as a central regulatory hub regulating fertility, sex recognition, and mating in males.

overall_design: The olfactory bulbs of 18 weeks old male C57Bl/6N mice were microdissected and immediately frozen in liquid nitrogen. Two biological replicates were analyzed, and each replicate comprised three olfactory bulbs. Singlet olfactory bulb nuclei were isolated by fluorescent-activated cells sorting through the presence of DAPI and subjected to 10x snRNA-Sequencing.

first: Decoster

last: Laurine

first: Trova

last: Sara

first: Zucca

last: Stefano

first: Bulk

last: Janice

first: Gouveia

last: Ayden

first: Ternier

last: Gaetan

first: Lhomme

last: Tori

first: Legrand

last: Amandine

first: Gallet

last: Sarah

first: Boehm

last: Ulrich

first: Wyatt

last: Amanda

first: Wahl

last: Vanessa

first: Wartenberg

last: Phillip

first: Hrabovszky

last: Erik

first: Rácz

last: Gergely

first: Luzzati

last: Federico

first: Nato

last: Guilia

first: Fogli

last: Marco

first: Peretto

last: Paolo

first: Schriever

last: Sonja C.

first: Bernecker

last: Miriam

first: Pfluger

last: Paul T

first: Steculorum

last: Sophie M

first: Bovetti

last: Serena

first: Rasika

last: Sowmyalakshmi

first: Prevot

last: Vincent

first: Silva

last: Mauro S B.

first: Giacobini


title: Targeting z-Crystallin by aspirin restores the sensitivity to cisplatin in resistant A2780 ovarian cancer cells

city: Florence

first: Matteo

last: Lulli

address: viale Morgagni 50

institute: University of Florence

summary: Ovarian cancer is the deadliest gynaecologic malignancies worldwide. Platinum based chemotherapy is the mainstay treatment for ovarian cancer; however, frequent recurrence and chemoresistance onset in patients with advanced diseases remain a therapeutic challenge. Although mechanisms underlying the development of chemoresistance are still ambiguous, the B-cell lymphoma-2 (Bcl-2) family is closely associated with chemoresistance in ovarian cancer. We previously disclosed that Zeta-Crystallin (CryZ) is a post-transcriptional regulator of Bcl-2 gene expression, by binding to Bcl-2 mRNA and increasing its half-life. Here, we investigated the role of CryZ as a novel therapeutic target in A2780 ovarian carcinoma cells by modulating the protein activity with acetylsalicylic acid (ASA) to restore chemosensitivity. Molecular docking and fragment-mapping based approach revealed potential interaction of ASA within CryZ protein. Inhibition of CryZ binding activity to Bcl-2 and Bcl-xl mRNA targets by ASA was demonstrated in A375 cells. Cytotoxicity assays were conducted in A2780S and A2780R ovarian cancer cells to evaluate if CryZ binding activity inhibition and CryZ silencing were able to reverse cisplatin resistance. ASA-treatment determined a downregulation of Bcl-2 and Bcl-xl mRNA levels in A2780S and A2780R cells. ASA-treatment or CryZ silencing were able to increase and restore the chemosensitivity in both sensitive and resistant A2780 ovarian cancer cells, respectively. In this research article we demonstrated that the pharmacological or genetic inhibition of CryZ restores the sensitivity to cisplatin in a model of sensitive or resistant ovarian cancer cells. These findings suggest a new gene-targeted chemotherapeutic approach to restore the cytotoxicity in drug-resistant ovarian cancers and increase the sensitivity in non-resistant cells.

overall_design: We investigated the role of CryZ as a novel therapeutic target in A2780 ovarian carcinoma cells by modulating the protein activity with acetylsalicylic acid (ASA) to restore chemosensitivity. Molecular docking and fragment-mapping based approach revealed potential interaction of ASA within CryZ protein

first: Andrea

last: Lapucci


title: Primary nasal viral infection rewires the tissue-scale memory response

city: Boston

first: Jose

last: Ordovas-Montanes

state: MA

address: 300 Longwood Ave

department: Gastroenterology, Hepatology and Nutrition Research

institute: Boston Children's Hospital

summary: The nasal mucosa is frequently the initial site of respiratory viral infection, replication, and transmission, but the cellular composition at homeostasis, during infection, and during a memory response are poorly understood. Here, we generated a single-cell RNA-sequencing (scRNA-seq) atlas of the murine nasal mucosa sampling three distinct regions before and during primary and secondary influenza A virus (IAV) infection. Primary infection was largely restricted to respiratory mucosa and induced stepwise changes in immune and epithelial cell type, subset, and state composition over time. Following viral clearance (14 dpi), rare, previously undescribed Krt13+ nasal immune-interacting floor epithelial (KNIIFE) cells expressing multiple genes with immune communication potential increased concurrently with tissue-resident memory T (TRM)-like cells and early IgG+/IgA+ plasmablasts. Proportionality analysis coupled with cell-cell communication inference, alongside validation by in situ microscopy, underscored the CXCL16–CXCR6 signaling axis between MDMs and effector CD8 T cells 8dpi and KNIIFE cells and TRM cells 14 dpi. Secondary influenza challenge with a homologous or heterologous strain administered 60 dpi induced an accelerated and coordinated myeloid and lymphoid response without epithelial proliferation, illustrating how tissue-scale memory to natural infection engages both myeloid and lymphoid cells to reduce epithelial regenerative burden. Raw and processed single-cell counts matrices can be accessed and downloaded from the Broad Institute Single-Cell Portal from studies SCP2216 and SCP 2221.

overall_design:
6-8 week old C57BL/6 mice were innoculated with 10^4 plaque forming units of influenza A virus PR8 (H1N1) in the upper respiratory tract through intranasal instilation (5 µl/nostril). Three mice per time point were sacrificed and respiratory mucosa (RM), olfactory mucosa (OM), and lateral nasal gland (LNG) were harvested. Single-cell RNA-sequencing was performed using 10x Genomics 3' v3.1 transcriptomics kit with mouse replicates (per tissue region) combined together and labeled with antibody-barcoded "hashes" to differentiate between samples (using Biolegend TotalSeqB). For rechallenge and memory experiments, PR8 infected mice were rechallenged with the same dose of PR8 or 10^5 plaque forming units of X31 (H3N2) and only RM tissue was collected and processed.
------------------------------
ADT/HTO      barcode sequence      hashtagged samples
TotalSeq-B0301 anti-mouse Hashtag 1      ACCCACCAGTAAGAC      mouse 1, all samples
TotalSeq-B0302 anti-mouse Hashtag 2      GGTCGAGAGCATTCA      mouse 2, all samples
TotalSeq-B0303 anti-mouse Hashtag 3      CTTGCCGCATGTCAT      mouse 3, all samples

first: Samuel

middle: W

last: Kazer

first: Colette

last: Match

first: Ulrich

last: von Andrian


title: Epigenetic and transcriptional consequences in the endosperm of chemically induced transposon mobilization in Arabidopsis

first: Juan

last: Santos-González

type: Methylation profiling by high throughput sequencing

summary: Genomic imprinting, an epigenetic phenomenon leading to parent-of-origin-specific gene expression, has independently evolved in the endosperm of flowering plants and the placenta of mammals-tissues crucial for nurturing embryos. While transposable elements (TEs) frequently colocalize with imprinted genes and are implicated in imprinting establishment, direct investigations of the impact of de novo TE transposition on genomic imprinting remain scarce. In this study, we explored the effects of chemically induced transposition of the Copia element ONSEN on genomic imprinting in Arabidopsis thaliana. Through the combination of chemical TE mobilization and doubled haploid induction, we generated a line with 40 new ONSEN copies. Our findings reveal a preferential targeting of maternally expressed genes (MEGs) for transposition, aligning with the colocalization of H2A.Z and H3K27me3 in MEGs-both previously identified as promoters of ONSEN insertions. Additionally, we demonstrate that chemically-induced DNA hypomethylation induces global transcriptional deregulation in the endosperm, leading to the breakdown of MEG imprinting. This study provides insights into the consequences of chemically induced TE remobilization in the endosperm, revealing that chemically-induced epigenome changes can have long-term consequences on imprinted gene expression.

overall_design:
RNAseq, WGBS and CUT&Tag datasets were obtained from INTACT purified endosperm nuclei from Col × Ler (L) reciprocal crosses at 4 days after pollination (DAP).
WGBS and CUT&Tag were generated in duplicates using the INTACT line (referred as C) or TEmob (T3, referred as Cm) genetic background in recirpocal crosses with Ler (L).
RNAseq libraries were generated in triplicates from TEmob (Cm) x Ler (L) reciprocal crosses.


title: ChIP–seq on periosteal cells derived from the pre-invasive group using antibodies to acetylated histone H3 Lys27 (H3K27ac), H3K4me3 and HIF-1α

city: Bunkyo-ku

first: Masayuki

last: Tsukasaki

state: Tokyo

department: Department of Immunology Graduate School of Medicine and Faculty of Medicine

country: Japan

institute: The University of Tokyo

summary: To test whether HIF-1a can directly induce Timp1 expression in periosteal cells at the pre-invasive stage of head and neck squamous cell carcinoma (HNSCC), we performed Chromatin immunoprecipitation followed by sequencing (ChIP–seq) on periosteal cells derived from the pre-invasive stage of a mouse HNSCC bone invasion model by using antibodies to acetylated histone H3 Lys27 (H3K27ac), H3K4me3 and HIF-1α.

overall_design: Primary periosteal cells (Ter119–CD45–CD31–PDPN+PDGFRα+ cells) were isolated from the tumour-excised calvarial tissues from the pre-invasive stage of a mouse HNSCC bone invasion model. Chromatin immunoprecipitation on 100,000 periosteal cells was performed with CUT&RUN Assay Kit (CST, #86652) using antibodies to acetylated histone H3 Lys27 (H3K27ac) (CST,#8173s), H3K4me3 (CST,#9751) and HIF-1α(CST,#36169). Illumina sequencing libraries were prepared using the NEBNext Ultra II DNA Library Prep Kit for Illumina (Cat No. 7645) according to the manufacturer’s instructions. Barcoded libraries were run on the NovaSeq 6000 in a PE150 run, and an average of 40 million paired reads were generated per sample.

first: Kazutaka

last: Nakamura


title: Whole-exome sequencing analysis of HNM007, AKR, and NOOC1 cells.

summary: To characterize the mutational status of mouse head and neck squamous cell carcinoma (HNSCC) cell lines, we performed whole-genome sequencing analysis on HNM007, AKR, and NOOC1 cells.

overall_design: Genomic DNA was extracted from HNM007, AKR, and NOOC1 cells by phenol–chloroform extraction and then purified with TIANamp Genomic DNA Kit (Spin Column). Whole-exome libraries were constructed with the Agilent SureSelectXT Mouse All Exon Kit. Libraries were constructed using 600 ng of DNA and were sequenced on the Illumina NovaSeq 6000 platform using 150 bp paired-end reads at the Rhelixa Inc, Tokyo, Japan.


title: Single-cell landscape of the mouse HNSCC-related bone invasive lesion

summary: To understand the invasive microenvironment of head and neck squamous cell carcinomas (HNSCCs) at a single-cell resolution, we performed single-cell RNA-sequencing (scRNA-seq) using cells derived from calvaria of tumor-free control mice and tumor-excised calvarial tissues from pre-invasive stage（scratch-negative）and invasive stage（scratch-positive）groups 1 week after the implantation of  C57BL/6-derived HNSCC cell line HNM007 cells.

overall_design: Single-cell RNA-sequencing was performed on live cells, mixed CD45+ and CD45– cells in a 1:1 ratio, derived from calvarial tissues of tumour-free control mice and tumour-excised pre-invasive stage（scratch-negative）and invasive stage（scratch-positive）mice 1 week after the implantation of HNM007 cells. The HNM007 cells were implanted onto the calvaria with or without prior scratching of the bone surface using a 21 G needle. This procedure mechanically removes the periosteum.


title: Differential regulation by CD47 and thrombospondin-1 of extramedullary erythropoiesis in mouse spleen

city: Bethesda

first: Thomas

middle: Joshua

last: Meyer

state: Maryland

address: 41 Center Drive, Building 41, Room B620

department: National Cancer Institute

institute: National Institutes of Health


title: An intestinal T effector cell subset leading to cancer initiation

city: Bron

first: Hector

last: Hernandez-Vargas

state: Rhone

address: 36 bis Rue de la Batterie

country: France

institute: Genomics Consulting

summary: Here, we revealed that an intestinal T cell subset, derived from already differentiated TH17 cells, induces the spontaneous transformation of the healthy epithelium. This subset produces a large panel of inflammatory cytokines and its tumorigenic potential is dependent on the transcriptional factors KLF6 and T-BET.

overall_design: Single cell RNA sequencing was performed in FACS purified Th17 cells.

first: Valentin

last: Thevin

first: Julien

last: Marie


title: Mitochondrial complex I promotes kidney cancer metastasis

city: Dallas

first: Divya

last: Bezwada

state: Texas

address: 5323 Harry Hines Blvd

institute: University of Texas Southwestern Medical Center

summary: Most kidney cancers display metabolic dysfunction but how this relates to cancer progression in humans is unknown. We infused 13C-labeled nutrients during surgical tumour resection in over 80 patients with kidney cancer. Labeling from [U-13C]glucose varies across subtypes, indicating that the kidney environment alone cannot account for all metabolic reprogramming in these tumours. Compared to the adjacent kidney, clear cell renal cell carcinomas (ccRCC) display suppressed labelling of tricarboxylic acid (TCA) cycle intermediates in vivo and in organotypic cultures ex vivo, indicating that suppressed labeling is tissue intrinsic. Infusions of [1,2-13C]acetate and [U-13C]glutamine in patients, coupled with measurements of respiration in mitochondria isolated from kidneys and tumours, reveal electron transport chain (ETC) defects in ccRCC. However, ccRCC metastases unexpectedly have enhanced TCA cycle labeling compared to primary ccRCCs, indicating a divergent metabolic program during metastasis in patients. In mice, stimulating respiration or NADH recycling in kidney cancer cells is sufficient to promote metastasis, while inhibiting ETC complex I decreases metastasis. These findings indicate that metabolic properties and liabilities evolve during kidney cancer progression in humans, and that mitochondrial function is limiting for metastasis but not for growth at the original site.

overall_design: RNA Sequencing of kidney cancer patient tissue samples. Patient samples are de-identified and are either from the adjacent kidney, a kidney cancer primary tumor, or clear cell renal cell carcinoma (ccRCC) metastasis.

first: Ralph

last: DeBerardinis


title: Differential regulation by CD47 and thrombospondin-1 of extramedullary erythropoiesis in mouse spleen (Bulk RNA-Seq)

summary: Extramedullary erythropoiesis is not expected in healthy adult mice, but erythropoietic gene expression was elevated in lineage-depleted spleen cells from cd47−/− mice. Elevated expression of several genes associated with early stages of erythropoiesis was observed in mice lacking CD47 or its signaling ligand thrombospondin-1, consistent with previous evidence that this signaling pathway inhibits expression of multipotent stem cell transcription factors in spleen. In contrast, cells expressing markers of committed erythroid progenitors including erythropoietin receptor, aquaporin-1, glycophorin A, and erythrocyte membrane-associated protein were more abundant in cd47−/− spleens but significantly depleted in thbs1−/− spleens. Single cell transcriptome analysis indicated that loss of CD47 is associated with accumulation and increased proliferation of CD34− committed erythroid progenitors in spleen, consistent with the known function of CD47 to limit turnover of aging erythrocytes. Conversely, loss of thrombospondin-1 delays turnover, which suppresses erythropoiesis in thbs1−/− spleens relative to the basal level in wild type mice.

overall_design: CD8+ T Cells were Isolated CD8a+ T Cell Isolation Kit mouseCD8a+ T Cell Isolation Kit  (Order no. 130-104-075) from Miltenyi biotec according to manufactures instructions. The freshly isolated CD8+ T cells were washed with filtered buffer solution containing phosphate-buffered saline (PBS), pH 7.2, 0.5% bovine serum albumin (BSA), and 2 Mm EDTA. The clumps of isolated cells were removed using Pre-Separation Filters (30 µm) and subjected to negative selection of CD8+ T cells. The cells were immediately placed at -80 °C until the RNA was extracted.

first: Sukhbir

last: Kaur

first: Maggie

last: Cam

first: David

last: Roberts

first: Rajdeep

last: Banerjee


title: PHF2 maintains neural progenitor genome stability by preserving pericentric heterochromatin integrity

city: Barcelona

last: Aguirre Infantes

state: Barcelona

address: Baldiri Reixac 15-21

department: Department of Structural and Molecular Biology

country: Spain

institute: IBMB-CSIC

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

overall_design: Refer to individual Series


title: Genomic heterogeneity drives mechanical adaptation in human tumor cells [RNA-seq]

city: College Station

first: Ting-Ching

last: Wang

address: 101 Bizzell St

institute: Texas A&M University

summary: The progression of many solid tumors is accompanied by temporal and spatial changes in the stiffness of the extracellular matrix (ECM). Cancer cells adapt to soft and stiff ECM through mechanisms that are not fully understood. In particular, it is well known that there is significant genetic heterogeneity from cell to cell in tumors, but how ECM stiffness as a parameter might interact with that genetic variation is not known. Here, we used the method of experimental evolution to study response of genetically variable and clonal tumor cell populations to ECM stiffness. Cell fitness increased on soft ECM over a period of several weeks in genetically variable but not clonal populations. DNA barcode-enabled clonal tracking revealed that sustained culture on soft ECM selects for a few genetic variants. These data provide the first evidence that ECM stiffness exerts natural selection on genetically variable tumor populations. Genome-wide analysis including RNA-seq, ATAC-seq and DNA methylation profiling reveal substantial differences in gene expression between selected populations and ancestral cells which are partially explained by epigenetic modifications. Soft-selected cells are highly migratory with enriched oncogenic signatures and exhibit highly unusual behaviors like spreading and traction force generation on ECMs as soft as 1 kPa. Cell spreading is the directly selected trait but not levels of integrins or adhesion proteins like talin. Overall, these data show that ECM stiffness in solid tumors may drive malignant behaviors through evolution by natural selection.

overall_design: To explore the mechanism underlying cell evolution on soft ECM, we performed whole genome transcriptome sequencing (RNA-seq).

first: Suchitaa

last: Sawhney

first: Daylin

last: Morgan

first: Richard

middle: L

last: Bennett

first: Richa

last: Rashmi

first: Marcos

middle: R

last: Estecio

first: Amy

last: Brock

first: Irtisha

last: Singh

first: Charles

middle: F

last: Baer

first: Jonathan

last: Licht

first: Tanmay

middle: P

last: Lele


title: A centrosomal defect of intestinal stem cells predisposes to Crohn’s Disease [RNA-Seq]

city: St. Louis

first: Vincent

last: Peng

state: MO

address: 660 S. Euclid Ave.

institute: Washington University School of Medicine in St. Louis

summary: The intestinal epithelium is a single-cell layer comprising a constellation of specialized cell types that functionally interconnect host physiological systems with extrinsic microbial communities, metabolites, and dietary factors1. Active renewal of the intestinal epithelium relies on stem cells that reside in invaginations of the epithelium known as crypts. Inflammatory Bowel Disease (IBD) disrupts normal functions and/or proportions of cells in the intestinal epithelium, including stem cell programming during epithelial restitution. Genome-wide association studies have provided a powerful means to identify loci and genes contributing to IBD risk and functional annotation of candidate genes have unveiled cellular programs and molecular mechanisms involved in IBD etiology. Here we find that risk for a major form of IBD, Crohn’s disease (CD), is linked to a variant of the Fibroblast Growth Factor Receptor 1 Oncogene Partner (FGFR1OP) locus, which encodes a protein of the centrosome, the microtubule organizing center that generates microtubule structures involved in cell division, polarity, and motility. Deletion of Fgfr1op in mouse intestinal epithelium impaired stem cell division and intestinal crypt architecture causing crypt loss, inflammation and death. Transcriptional, biochemical and morphological analyses demonstrated that Fgfr1op coordinates the centrosome with the actin cytoskeleton and with specialized cell junctions known as desmosomes. We conclude that a genetically inherited centrosomal defect predisposes to CD by affecting renewal of the intestinal epithelium.

overall_design: Small intestinal spheroids from WT and Fgfr1op KO mice at day 3 post-hydroxytamoxifen in vitro (4 mice each group). Small intestinal crypt-enriched cells from WT and Fgfr1op KO mice at days 6 and 9 post-tamoxifen (3 mice each group). Scramble or shFgfr1op Caco-2 cells at day 3 post-doxycycline.

first: Tihana

last: Trsan

first: Ramnik

last: Xavier

first: Marco

last: Colonna


title: PHF2 maintains neural progenitor genome stability by preserving pericentric heterochromatin integrity (ChIP-Seq)

first: Samuel

summary: Heterochromatin stability is crucial for progenitor proliferation during development. It relays on the maintenance of local hubs of H3K9me. However, the mechanisms underlying the establishment of competent local levels of H3K9me remain poorly understood. To address this intriguing question, we used a neural stem cell (NSC) model to analyze the significance of the H3K9me2 demethylase PHF2, which is crucial for progenitor proliferation. Through mass spectroscopy and genome-wide assays, we uncovered that PHF2 interacts with heterochromatin components and it is enriched at pericentromeric heterochromatin (PcH) boundaries. This binding is essential for maintaining silenced the satellite repeats and to prevent DNA damage and genome instability. To do that, PHF2 balances H3K9me3 levels at these boundaries to ensure high H3K9me3 levels at satellite repeats. Mechanistically, we discover that while its catalytic and PHD domains are indispensable, the intrinsically disordered region within PHF2 is dispensable for stabilizing PcH. Altogether, our study sheds light on the intricate relationship between heterochromatin stability and progenitor proliferation during mammalian neurogenesis.

overall_design: Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for Histone 3 lysine 9 trimetilated (H3K9me3) and Histone 3 lysine 4 trimetilated (H3K4me3) were performed on Neural Stem Cells (NSCs) under shControl and shPHF2 conditions by duplicate. In addition, chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for PHF2 was performed by duplicate. Input of each immunoprecipitation is included.

last: Aguirre

first: Stella

last: Pappa

first: Núria

last: Serna-Pujol

first: Natalia

last: Padilla

first: Silvina

middle: A

last: Nacht

first: Guillermo

last: Vicent

first: Albert

last: Jordan

first: Xavier

last: de la Cruz

first: Marian

last: Martínez-Balbás


title: A centrosomal defect of intestinal stem cells predisposes to Crohn’s Disease [scRNA-Seq]

overall_design: 3 WT and 3 FGFR1OP KO samples


title: MHC II Heterozygosity Limits T Cell Receptor Variability in CD4 T Cells

status: Public on Jun 16 2024

city: Denver

first: Philippa

last: Marrack

state: CO

address: 1400 Jackson St

department: Immunology & Genomic Medicine

institute: National Jewish Health

summary: αβ T cell receptor (TCR) V(D)J genes code for billions of TCR combinations.  However only some appear on peripheral T cells in any individual because, to mature, thymocytes must react with low affinity but not high affinity with thymus expressed MHC/peptides. MHC proteins are very polymorphic. Different alleles bind different peptides. Therefore any individual might express many different MHC alleles to ensure that some peptides from an invader are bound to MHC and activate T cells.  However, most individuals express limited numbers of MHC alleles. To explore this we compared the TCR repertoires of naïve, CD4 T cells in mice expressing one or two MHC II alleles.  Surprisingly, the TCRs in heterozygotes were less diverse that those in the sum of their MHC II homozygous relatives. Our results suggest that thymus negative selection cancels out the advantages of increased thymic positive selection in the MHC heterozygotes.

overall_design: The aim of this study was to find out how MHC heterogeneity affects the T cell receptor (TCR) repertoire of naive CD4 T cells in mice. This work aimed to provide a mechanistic link between genetic background and phenotypic TCR repertoire. The focus of this work is to address a basic unsolved immunological question. However, understanding how different MHC alleles dictate the specific selection of diverse TCR repertoires could have enormous practical use in guiding the development of personalized vaccines, in monitoring autoimmune development, and predicting the responses of patients to infectious disease or immunotherapy checkpoint blockade. Because it is, at the moment, impossible to identify all the TCRs expressed in an individual wild type mouse we bred animals on the C57BL/6 background that were heterozygous for TCRalpha expression and also expressed a transgenic TCRbeta chain and no other TCRbetas. Consequently, in these animals, the transgenic TCRbeta is the only one expressed and each T cell expresses a single, varying TCRalpha chain. Thus the TCR repertoire in the animal can be understood by sequencing the expressed TCRalpha genes. Deposited samples are RNA sequencing reads of these recombined TCRalpha genes. Animals of this genotype were bred to express on the C57BL/6 background, homozygously, H-2b, H-2f, H-2g7 or H-2s. MHC heterozygotes were produced by intercrossing these animals.

first: Alexander

last: Brown

first: Janice

last: White

first: Laura

last: Shaw

first: Jimmy

last: Gross

first: Andrei

last: Slabodkin

first: Ella

last: Kushner

first: Victor

last: Greiff

first: Jennifer

last: Matsuda

first: Laurent

last: Gapin

first: James

last: Scott-Browne

first: John

last: Kappler


title: A centrosomal defect of intestinal stem cells predisposes to Crohn’s Disease

first: Vincent

last: Peng


title: Effect of mccc-1(ww4) mutation on gene expression in Caenorhabditis elegans at young adult stage fed on Comamonas aquatica DA1877

city: Worcester

first: Yonguk

last: Lee

state: Massachusetts

address: 55 N Lake Avenue

department: Department of Systems Biology

institute: University of Massachusetts Chan Medical School

summary:
mccc-1(ww4) mutant animals has no obvious developmental defect, but we observed different abundance of many metabolites by LC-MS/MS analysis.
Then, we assumed different metabolite abundance is associated with different metabolic gene expression, sucha that RNA-seq was performed to examine differently expressed gene.

overall_design: Comparative gene expression profiling analysis of RNA-seq data between control animal (Strain name: VL749) and mccc-1(ww4) mutant animals (Strain name: VL1080)

first: Yong-Uk

last: Walhout


title: A centrosomal defect of intestinal stem cells predisposes to Crohn’s Disease [array]

overall_design: Whole jejunum from WT and Fgfr1op KO mice at days 6, 9, and 12 post-tamoxifen (2 mice each).

first: Tihana

last: Trsan

first: Marco

last: Colonna

first: Ramnik

last: Xavier


title: Enterovirus 3A protein disrupts endoplasmic reticulum homeostasis through interaction with GBF1

city: Suita

first: Daisuke

last: Motooka

state: Osaka

address: 3-1, Yamadaoka

institute: NGS core facility, Research Institute for Microbial Diseases, Osaka University

summary: Enterovirus A71 (EV-A71) is a single-stranded, positive sense RNA virus causing endoplasmic reticulum stress to induce or modulate downstream signaling pathways known as the unfolded protein responses (UPR). The 3A protein of EV-A71 was identified as a major regulator of the UPR caused by infection with EV-A71. To identify the molecular pathways for the 3A protein to regulate the UPR, we performed RNA sequencing (RNA-seq) with RD cells expressing the 3A protein. Wild type or defective mutant of 3A protein that carried alanine substitutions of Ile8 or Ile10 (3A-I8A or -I10A) was expressed in the cells. The RNA-seq with cells expressing either WT or mutant 3A was performed.

overall_design: To investigate the functions of EV-A71 3A protein, we generated a defective mutant of 3A protein that carried alanine substitutions of Ile8 or Ile10 (3A-I8A or -I10A). Gene expression profiling analysis of RD cells, RD cells expressing wild type 3A protein, RD cells expressing 3A-I8A, or RD cells expressing 3A-I10A was performed.

first: Junki

last: Hirano

first: Yoshiharu

last: Matsuura


title: Differential regulation by CD47 and thrombospondin-1 of extramedullary erythropoiesis in mouse spleen (scRNA-Seq)

first: Thomas

last: Meyer

overall_design: Single cell suspensions from WT, thbs1−/−  and cd47−/−  mice spleens were depleted of all mature hematopoietic cell lineages including erythroblasts and mature RBC by passing through CD8a (Ly-2) microbeads and CD8+ T cell untouched negative selection microbeads (Miltenyi Biotec). Single cell suspensions were incubated with the supplied antibody cocktail of the CD8+ T cell isolation kit for 15 min. on ice and then passed through the column as per manufacturer’s instructions. After 3 washes, the flow through was centrifuged and incubated with the CD8a (Ly-2) microbeads and then passed through the magnetic column to obtain the lineage-depleted cell populations.

first: Sukhbir

last: Kaur

first: Maggie

last: Cam

first: David

last: Roberts

first: Rajdeep

last: Banerjee


title: Ferritinophagy is a Druggable Vulnerability of Quiescent Leukemic Stem Cells [9a]

city: paris

first: Jerome

last: Tamburini

address: 22 rue mechain

institute: Cochin Institute

summary: Acute myeloid leukemia (AML) remains a challenging hematological malignancy with poor prognosis and limited treatment options. Post-therapy relapse, particularly driven by leukemic stem cells (LSCs), contributes to therapeutic failure and adverse outcome. Here, we investigated the role of quiescence and its associated molecular mechanisms in AML pathogenesis and identified potential vulnerabilities for therapeutic intervention. We found that quiescent AML cells, enriched for LSC functions, exhibited a distinct gene set that served as a significant prognostic factor for poor outcomes in AML patients. Furthermore, quiescent cells displayed heightened autophagic activity, with a reliance on ferritinophagy, a selective form of autophagy mediated by Nuclear Receptor Coactivator 4 (NCOA4), for iron bioavailability. Inhibition of NCOA4 genetically or chemically showed potent anti-leukemic effects, particularly targeting the LSC compartment. These findings uncover that ferritinophagy inhibition may represent a promising therapeutic strategy for patients with AML.

overall_design: Eight different primary AML samples were incubated with vehicle or 1mM compound 9a for 24 h.

first: Clement

last: Larrue


title: Small molecules that induce proliferation of human iPSC-cardiomyocytes

city: Charlottesville

first: Jeffrey

middle: John

last: Saucerman

department: Biomedical Engineering

institute: University of Virginia

summary: We selected a set of five compounds with diverse known targets that induce proliferation of human iPSC-derived cardiomyocytes. We performed bulk RNA-sequencing of human iPSC-CMs in response to the five compounds to identify common mechanisms mediating compound-induced cell cycle progression. Transcriptome profiling revealed these compounds all induced a common set of pathways including cell cycle, DNA repair, and kinesins.

overall_design: Cellular Dynamics iCell Cardiomyocytes were treated with DMSO or 1 of 5 small molecules (C1, C2, C3, C4, C5) for 24 hours. Each condition had 3 replicates.

first: Laura

last: Woo


title: Ferritinophagy is a Druggable Vulnerability of Quiescent Leukemic Stem Cells [shCTL vs shNCOA4]

overall_design: Four different AML samples were transduced ex vivo with lentiviral vectors expressing mCherry-tagged NCOA4-targeting or control shRNAs. Transcriptomic analysis was performed 72 h after transduction.


title: Ferritinophagy is a Druggable Vulnerability of Quiescent Leukemic Stem Cells [scRNA-seq]

overall_design: Two PDX AML samples (TUH84 and TUH102) were analyzed seven days after xenotransplantation in vivo in immunodeficient mice.


title: Ferritinophagy is a Druggable Vulnerability of Quiescent Leukemic Stem Cells


title: Gene expression profile at single cell level of primary regulatory T cells(CD25+CD127- Treg), primary induced regulatory T cells(iTreg), and AMRT treated iPSC-derived CD4+ T cells (iCD4+Treg-like)

city: Kyoto

first: Shin

last: Kaneko

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

department: Department of Cell Growth and Differentiation

institute: Center for iPS Cell Research and Application (CiRA), Kyoto University

summary: We used single cell RNA sequencing (scRNA-seq) to analyze the difference among CD25+CD127-Treg, iTreg, and iCD4+Treg-like.  We expanded CD25+CD127-Treg by TCR stimulation in the presense of AS2863619, anti-TNFR2 antibody, and TGFbeta (AMT). We differentiated iTreg from naive CD4+T cells and iCD4+Treg-like from iPSC-derived CD4+T cells in the presence of TCR stimulation, AS2863619, anti-TNFR2 antibody, TGFbeta, and rapamycin (AMRT).

overall_design: CD25+CD127-Treg was isolated from PBMCs by FASC and expanded by TCR stimulation in the presence of AS2863619, anti-TNFR2 antibody, and TGFbeta (AMT) for 14 days. iTreg was differentiated from naïve CD4+ T cells, which were isolated from PBMCs by FACS, in the presence of TCR stimulation, AS2863619, anti-TNFR2 antibody, TGFbeta, and rapamycin (AMRT) for 14 days. iCD4+Treg-like was differentiated from iPSC-derived CD4+T cells in the presence of TCR stimulation, AS2863619, anti-TNFR2 antibody, TGFbeta, and rapamycin (AMRT) for 7 or 14 days.


title: Ferritinophagy is a Druggable Vulnerability of Quiescent Leukemic Stem Cells [cycling and quiescent fractions]

overall_design: Four different primary AML samples were sorted based on Pyronin Y / Hoechst staining between quiescent and cycling fractions.


title: Multiplexed single-cell characterization of alternative polyadenylation regulators

status: Public on Jun 15 2024

city: New York

first: Madeline

last: Kowalski

address: 101 6th Ave

institute: New York Genome Center


title: Multiplexed single-cell characterization of alternative polyadenylation regulators (scRNA-Seq)

summary: Most mammalian genes have multiple polyA sites, representing a substantial source of transcript diversity regulated by the cleavage and polyadenylation (CPA) machinery. To better understand how these proteins govern polyA site choice we introduce CPA-Perturb-seq, a multiplexed perturbation screen dataset of 42 CPA regulators with a 3’ scRNA-seq readout that enables transcriptome-wide inference of polyA site usage. We develop a framework to detect perturbation-dependent changes in polyadenylation and characterize modules of co-regulated polyA sites. We find groups of intronic polyA sites regulated by distinct components of the nuclear RNA life cycle including elongation, splicing, termination, and surveillance. We train and validate a multi-task deep neural network (APARENT-Perturb) for tandem polyA site usage, delineating a cis-regulatory code that predicts perturbation response and reveals interactions between regulatory complexes. Our work highlights the potential for multiplexed single-cell perturbation screens to further our understanding of post-transcriptional regulation.

overall_design: We conducted a pooled single-cell CRISPR screen (in both HEK293FT and K562 cell lines) targeting 42 genes previously implicated in regulating cleavage and polyadenylation.  We inferred polyA site usage at single-cell resolution from a scRNA-seq readout, and determine how each of these perturbations effects polyA site usage. We validate our polyA site quantifications by performing 3'RACE followed by Illumina sequencing at 7 loci in NT and NUDT21-perturbed cells. We also built a sequence-based deep learning model (APARENT-Perturb) to describe how sequence affects perturbation, and design an MPRA to validate these sequence-based predictions on polyA site usage.

first: Hans-Hermann

last: Wessels

first: Johannes

last: Linder

first: Carol

last: Dalgarno

first: Isabella

last: Mascio

first: Saket

last: Choudhary

first: Austin

last: Hartman

first: Yuhan

last: Hao

first: Anshul

last: Kundaje

first: Rahul

last: Satija


title: Multiplexed single-cell characterization of alternative polyadenylation regulators (MPRA)


title: Multiplexed single-cell characterization of alternative polyadenylation regulators (3PRACE)


title: Sex-biasing influence of autism-associated Ube3a gene overdosage at connectomic, behavioral and transcriptomic levels

city: Rovereto

first: Caterina

last: Montani

address: Corso Bettini 31

institute: IIT - Istituto Italiano di Tecnologia

summary: We tested the hypothesis that increase dosage of Ube3a may exert its influence on transcriptomic organization in a sex-biased manner

overall_design: We performed gene expression analysis using data obtained from RNA-seq of prefrontal cortex and hypothalamus dissected from adult male and female mice with extra copies of Ube3a (Ube3a2x) and Wild-type controls

first: Luigi

last: Balasco

first: Pierre

last: Lau

first: Giovanni

last: Provenzano

first: Yuri

last: Bozzi

first: Michael

middle: V

last: Lombardo

first: Alessandro

last: Gozzi


title: Cell state transitions are decoupled from cell division during early embryo development [II]

status: Public on Jun 14 2024

first: Kalki

last: Kukreja

address: 200 Longwood Avenue

institute: Harvard University

summary: Paper abstract: As tissues develop, cells divide and differentiate concurrently. Conflicting evidence shows that cell division is either dispensable or required for formation of cell types. To determine the role of cell division in differentiation, we arrested the cell cycle in zebrafish embryos using two independent approaches and profiled them at single-cell resolution. We show that cell division is dispensable for differentiation of all embryonic tissues during initial cell type differentiation from early gastrulation to the end of segmentation. However, in the absence of cell division, differentiation slows down in some cell types, and cells exhibit global stress responses. While differentiation is robust to blocking cell division, the proportions of cells across cell states are not but show evidence of partial compensation. This work clarifies our understanding of the role of cell division in development and showcases the utility of combining embryo-wide perturbations with single-cell RNA sequencing to uncover the role of common biological processes across multiple tissues.

overall_design: Design of experiment for this particular dataset: Forty tails from zebrafish embryos at 24, 38 and 48 hours post fertilization (hpf) were collected. Cells were dissociated from these tails and multi-seq tags were added to the cells for each time point. Cells from all time points were pooled and single-cell sequencing was performed using Indrops. Four gene expression (GEX) libraries were prepared for transcriptomic information and four multiseq tag libraries (TAG) were prepared for reading out the multiseq tags. We provide the raw data (.fastq files), processed data - both raw (counts.tsv.gz) and filtered by total UMI counts (filtered.h5ad files) and the entire filtered data after removing doublets as all_data.h5ad. We also provide counts for multi-seq tags (example, TAG_1.counts.csv). Information about pooling libraries and library indices is provided in Pooling_Samples.xlsx. Only 24 hpf data was used for Kukreja et. al. 2024 and hence the cell state information we provide in the all_data.h5ad is only for 24 hpf.

first: Nikit

last: Patel

first: Allon

last: Klein


title: A single-cell transcriptomic dataset of pluripotent stem cell-derived astrocytes via NFIB/SOX9 overexpression

city: ShenZhen

first: Ran

last: YI

state: Guangdong

address: 1068 Xueyuan Avenue

department: The Brain Cognition and Brain Disease Institute

institute: Shenzhen Institute of Advanced Technology，Chinese Academy of Sciences

summary: Astrocytes, the predominant glial cells in the central nervous system, play essential roles in maintaining brain function. Reprogramming induced pluripotent stem cells (iPSCs) to become astrocytes through overexpression of the transcription factors, NFIB and SOX9, is a rapid and efficient approach for studying human neurological diseases and identifying therapeutic targets. However, the precise differentiation path and molecular signatures of induced astrocytes remain incompletely understood. Accordingly, we performed single-cell RNA sequencing analysis on 64,736 cells to establish a comprehensive atlas of NFIB/SOX9-directed astrocyte differentiation from human iPSCs. Our dataset provides detailed information about the path of astrocyte differentiation, highlighting the stepwise molecular changes that occur throughout the differentiation process. This dataset serves as a valuable reference for dissecting uncharacterized transcriptomic features of NFIB/SOX9-induced astrocytes and investigating lineage progression during astrocyte differentiation. Moreover, these findings pave the way for future studies on neurological diseases using the NFIB/SOX9-induced astrocyte model.

overall_design: Astrocytes were generated from multiple iPSC lines (iPSC1, DYR0100 line; iPSC2, BIONi037-A line) by overexpressing Nfib and Sox9. The resulting astrocytes derived from iPSC1 and iPSC2 were profiled by scRNA-seq. Moreover, the resulting cells at multiple stages (Day 0, 1, 3, 8, 14, 21) along monoclonal iPSC1 differentiation were profiled by single-cell RNA sequencing.

last: Yi

first: Shuai

last: Chen

first: Mingfeng

last: Guan

first: Chunyan

last: Liao

first: Yao

last: Zhu

first: Jacque

last: Ip

first: Tao

last: Ye

first: Yu


title: Gut-derived memory γδ T17 cells exacerbate sepsis-induced acute lung injury [ChIP-seq]

city: Wuhan

first: Mengyuan

last: Wang

state: Hubei

address: NO.1277 JieFang Avenue

department: Department of Critical Care Medicine

institute: Union Hospital, Tongji Medical College, Huazhong University of Science and Technology

summary: Sepsis, a worldwide health crisis, is notorious for its high mortality rate, often attributed to the development of acute lung injury. The direct migration of intestinal immune cells to the lung as a mediator of acute lung injury remains unclear. Our study unveiled a process where cecal ligation and puncture-induced sepsis prompted a migration of γδT cells from the small intestine to the lung, subsequently triggering an overwhelming inflammatory response dominated by IL-17A.

overall_design: chromatin immunoprecipitation sequencing (ChIP-seq) experiments targeting Lef1 in MH-S alveolar macrophages to investigate the potential specific binding between Lef1 and CCL1


title: Gut-derived memory γδ T17 cells exacerbate sepsis-induced acute lung injury [scRNA-seq]

overall_design: CD45+ immune cells from the small intestine tissue of septic and sham-operated C57 mice were isolated Fluorescence-activated cell sorting (FACS) and performed to scRNA seq


title: Identification of Circular RNAs Regulating Cardiomyocyte Proliferation in Neonatal Pig Hearts

city: Scottsdale

first: Wuqiang

state: AZ

address: 13400 E shea blvd

department: Cardiovascular Medicine

institute: Mayo Clinic

type: Non-coding RNA profiling by high throughput sequencing

summary: Little is known about the expression patterns and functions of circular RNAs (circRNAs) in the heart of large mammals. In this study, we examined the expression profiles of circRNAs, microRNAs (miRNAs), and messenger RNAs (mRNAs) in neonatal pig hearts. Pig heart samples collected on postnatal days 1 (P1), 3 (P3), 7 (P7) and 28 (P28) were sent for total RNA sequencing. Our data revealed a total of 7000 circRNAs in the 24 pig hearts. Pathway enrichment analysis of hallmark gene sets demonstrated that differentially expressed circRNAs are engaged in different pathways. The most significant difference was observed between P1 and the other three groups (P3, P7 and P28) in pathways related to cell cycle and muscle development. Out of the ten circRNAs that were validated through real-time quantitative polymerase chain reaction (qRT-PCR) to confirm their expression, six exhibited significant effects on cell cycle activity in human induced pluripotent stem cells-derived cardiomyocytes (hiPSC-CMs) following small interfering RNA-mediated knockdown. The circRNA-miRNA-mRNA networks were constructed to understand the potential mechanisms of circRNAs in the heart. In conclusion, our study provided a dataset for exploring the roles of circRNAs in pig hearts. In addition, we identified several circRNAs that regulate cardiomyocyte cell cycle.

overall_design:
To provided insights into the complex regulatory processes involving circRNAs, miRNAs, and mRNAs during postnatal heart development in newborn pigs, we performed RNA sequencing of mRNAs and noncoding RNAs including circRNAs and miRNAs using samples from neonatal pig hearts at different developmental stages (P1, P3, P7, and P28).
The heart samples were sent to two different companies for sequencing. In the first batch, 12 samples from animals of different ages (3 animals at each age) were sent to Novogene Inc. for circRNA sequencing (annotated as “nv1”). A second batch of 12 more samples was sent to CD Genomocs Inc. (annotated as “cd”) for total RNA sequencing. In the third batch, new samples from the same 12 animals in the first batch were sent to Novogene Inc. (annotated as “nv2”) for total RNA sequencing.
We validated the sequencing data using real-time quantitative polymerase chain reaction (qRT-PCR).
Differential expression analysis and pathway enrichment analysis were done using different bioinformatics tools. Functional experiments in the cultured cardiomyocytes via small interfering RNA (siRNA) mediated knockdown of selected circRNA candidates.

first: Junwen

first: Ling

last: Tang

first: Verah

last: Nyarige

first: Pengsheng

last: Li


title: Single-cell analysis reveals lasting immunological consequences of influenza infection and respiratory immunisation in the pig lung

city: Cambridge

first: Andrew

last: Muir

address: B570, Babraham Instititute

institute: Babraham Institute

summary: The pig is a natural host for influenza viruses and integrally involved in virus evolution through interspecies transmissions between humans and swine. We employed scRNA-seq and flow cytometry to characterize the major leucocyte subsets in bronchoalveolar lavage (BAL), twenty-one days after H1N1pdm09 infection or respiratory immunization with an adenoviral vector vaccine expressing haemagglutinin and nucleoprotein with or without IL-1β. Mapping scRNA-seq clusters from BAL onto those previously described in peripheral blood facilitated annotation and highlighted differences between tissue resident and circulating immune cells. ScRNA-seq data and functional assays revealed lasting impacts of immune challenge on BAL populations. First, mucosal administration of IL-1β reduced the number of functionally active Treg. Second, influenza infection upregulated IFI6 in BAL cells, decreasing their susceptibility to virus replication in vitro. Our data provide a reference map of BAL cells following respiratory infection or immunization in a highly relevant large animal model for respiratory virus infection.

overall_design: Samples obtained from a previous experiment conducted on inbred Babraham pigs were utilized (Schmidt et al., 2023). Briefly three groups of 5 pigs each, were treated as follows: 1) infected intranasally with 8x106 PFU of A/swine/England/1353/2009 (pH1N1) using a mucosal atomization device (MAD); 2) immunized intranasally with 1x109 particles of recombinant adenoviral vectors expressing NP from H1N1/PR8/34 virus and HA from pH1N1/Texas/05/2009 (Ad-HA/NP) using a MAD and 3) immunized intranasally with a mix of 1x109 particles of Ad-HA/NP and 1x109 particles recombinant adenoviral vector encoding porcine IL-1β (Ad-HA/NP+Ad-IL-1β) using a MAD. Control pigs (n=3) received PBS intranasally. The animals were culled three weeks later, and bronchoalveolar lavage (BAL), tracheobronchial lymph nodes and blood collected. Virus shedding and immune responses in respiratory tissues and peripheral blood were described in (Schmidt et al., 2023). Cryopreserved BAL cells were thawed, washed and resuspended in PBS. Cells were incubated with antibodies against CD4 (clone 74-12-4, PerCP-Cy5.5-conjugated, BD Biosciences), CD8β (clone PPT23, FITC-conjugated, Bio-Rad), CD3 (clone PPT3, APC-conjugated, Southern Biotech), CD16 (clone G7, AF647-conjugated, Bio-Rad), CD172a (clone 74-22-15, PE-conjugated, Bio-Rad) and Fixable Viability Dye eFluor780 (ThermoFisher). After two washes in PBS, cells were fixed, washed again and sorted on a FACSAria UIII (BD Biosciences), equipped with 405, 488, 561 and 640nm lasers and BD FACSDiva 8 software. After gating on FSC-AdimSSC-Adim cells and exclusion of doublets and dead cells, CD4+, CD8β+ and CD3-CD16- (to enrich for B cells) cells were sorted (12,000 cells per population). In addition, after gating on FSC-AbrightSSC-Abright cells, doublet and dead cell exclusion and gating on CD16+CD172a+ cells, 12,000 macrophages were sorted. Cells from each fraction were pooled and subjected to partitioning and barcoding on a 10x Genomics Chromium iX Controller. For 10X, single cells were processed with the Chromium iX controller, using the Next GEM chip G, and single cell 3’ kit v3.1 library preparation kit (10X Genomics, Pleasanton, CA). An estimated 10,000 cells were targeted for recovery during partitioning. Samples were randomly grouped and run with two libraries per sequencing run on three NextSeq High Output 150 cycle reagent kits (Illumina, San Diego, CA), with 1% PhiX, targeting approximately 200 million reads per sample. Bcl files were demultiplexed using ‘cellranger mkfastq’ and aligned/counted using ‘cellranger count’ (cellranger-7.0.0) using default parameters and Sus scrofa genome (genome assembly 11.1, Ensembl release 107).

first: Basudev

last: Paudyal

first: Selma

last: Schmidt

first: Ehsan

last: Sedaghat-Rostami

first: Soumendu

last: Chakravarti

first: Sonia

last: Hernandez

first: Katy

last: Moffat

first: Noemi

last: Polo

first: Nicos

last: Angelopoulos

first: Anna

first: Matthias

last: Tenbusch

first: Graham

last: Freimanis

first: Wilhelm

last: Gerner

first: Arianne

last: Richard

first: Elma

last: Tchilian


title: Cell and tissue adaptation in the pulmonary autograft in a large animal model of the Ross procedure

city: Leuven

first: Elizabeth

last: Jones

address: Herestraat 49, bus 911

department: Centre for Molecular and Vascular Biology

country: Belgium

institute: KU Leuven

summary: In the Ross procedure, a patient's pulmonary valve is transplanted into aortic position. Reoperation is needed in many cases due to failure through dilatation of the pulmonary autograft. To further understand the failure mechanisms, gene expression profiling of the autograft tissue was performed and compared to control aortic and pulmonary sinus tissue though bulk RNA-seq. Notable results include the upregulation of inflammatory pathways and genes coding for proteins important for the structural integrity of the tissue, such as extracellular matrix proteins, collagenases and elastases.

overall_design:
In three sheep, the pulmonary root was transplanted into aortic position to form the pulmonary autograft.
Six months after the surgery, autograft sinus tissue was extracted to perform gene expression profiling though RNA-seq in these three sheep.
Pulmonary sinus and aortic sinus tissue was extracted from non-operated sheep to perform gene expression profiling through RNA-seq as control.

first: Lucas

last: Van Hoof

first: Filip

last: Rega

first: Peter

last: Verbrugghe

first: Nele

last: Famaey

first: Lauranne

last: Maes

first: Amber

last: Hendrickx


title: YY1-controlled regulatory connectivity and transcription are influenced by the cell cycle

city: Philadelphia

first: Jessica

last: Lam

state: PA

address: 3615 Civic Center Blvd

department: Perelman School of Medicine

institute: University of Pennsylvania

summary: Few transcription factors have been examined for their direct roles in physically connecting enhancers and promoters. Here acute degradation of Yin Yang 1 (YY1) in erythroid cells revealed its requirement for the maintenance of numerous enhancer-promoter loops, but not compartments or domains. Despite its reported ability to interact with cohesin, the formation of YY1-dependent enhancer-promoter loops does not involve stalling of cohesin-mediated loop extrusion. Integrating mitosis-to-G1-phase dynamics, we observed partial retention of YY1 on mitotic chromatin, predominantly at gene promoters, followed by rapid rebinding during mitotic exit, coinciding with enhancer-promoter loop establishment. YY1 degradation during the mitosis-to-G1-phase interval revealed a set of enhancer-promoter loops that require YY1 for establishment during G1-phase entry but not for maintenance in interphase, suggesting that cell cycle stage influences YY1's architectural function. Thus, as revealed here for YY1, chromatin architectural functions of transcription factors can vary in their interplay with CTCF and cohesin as well as by cell cycle stage.

overall_design: We performed ChIP-seq (YY1, CTCF, RAD21, LDB1, H3K27ac, and total Pol2) and Micro-C in asynchronous YY1-AID cells treated with auxin for 4h. RNA-seq and TT-seq were also performed in asynchronous YY1-AID cells. Additional Pol2 ChIP-seqs and Micro-C were performed on synchronized and FACS-purified YY1-AID cells (nocodazole treatment for 7h with simultaneous auxin treatment for 4h). YY1 ChIP-seq was also performed on synchronized and FACS-purified G1E-ER4 MD-mCherry cells at mitosis-to-G1 stages (nocodazole treatment for 7h).

first: Gerd

last: Blobel


title: Cranioencephalic functional lymphoid units in glioblastoma

city: Essen

first: Björn

last: Scheffler

address: Hufelandstrasse 55

department: DKTK Division Translational Neurooncology at the WTZ

institute: DKFZ (German Cancer Research Center)

summary: We used single cell RNA sequencing to profile the immune cell repertoire of tumor tissue, peripheral blood mononuclear cells (PBMC), bone marrow mononuclear cells (BMMC) from distal bone and cranial (skull) bone from human treatment-naive glioblastoma patients. For comparison, we obtained and analyzed control samples (cranial bone and PBMC) from human non-malignant intracranial disease.

overall_design: CD45+ cells were derived by magnetic cell separation from fresh clinical samples (tumor tissue, cranial bone, PBMC, BMMC) and analyzed using the 10x Genomics Chromium Next GEM Single Cell 3´ Reagent Kits v3.1 or the 10x Genomics Chromium Next GEM Single Cell 5´ Reagent Kits v2. A second set of data was derived from CD45+ single cell suspensions that underwent additional magnetic depletion of myeloid cells (CD14 negative flowthrough) and were further analyzed using the 10x Genomics Chromium Next GEM Single Cell 5´ Reagent Kits v2.

first: Celia

last: Dobersalske

first: Laurèl

last: Rauschenbach

first: Yichao

last: Hua

first: Florian

last: Rambow

first: Ulrich

last: Sure

first: Bjoern


title: scRNAseq of human blood, tonsil, lung and small intestine innate lymphoid cells

city: Saint Louis

first: Marco

last: Colonna

state: Missouri

address: 660 S Euclid Ave

department: Pathology and Immunology

institute: Washington University in St Louis School of Medicine

summary: Group 1 innate lymphoid cells (ILC1s) are cytotoxic and interferon gamma-producing lymphocytes lacking antigen-specific receptors, which include ILC1s and natural killer (NK) cells. In mice, ILC1s differ from NK cells, as they develop independently of the NK-specifying transcription factor EOMES, while requiring the repressor ZFP683 (ZNF683 in humans) for tissue residency. Here we identify highly variable ILC1 subtypes across tissues through investigation of human ILC1 diversity by single-cell RNA sequencing and flow cytometry. The intestinal epithelium contained abundant mature EOMES− ILC1s expressing PRDM1 rather than ZNF683, alongside a few immature TCF7+PRDM1− ILC1s. Other tissues harbored NK cells expressing ZNF683 and EOMES transcripts; however, EOMES protein content was variable. These ZNF683+ NK cells are tissue-imprinted NK cells phenotypically resembling ILC1s. The tissue ILC1-NK spectrum also encompassed conventional NK cells and NK cells distinguished by PTGDS expression. These findings establish a foundation for evaluating phenotypic and functional changes within the NK-ILC1 spectrum in diseases.

overall_design: Blood samples were collected from discarded leukoreduction system (LRS) chambers from apheresis procedures. Blood lymphocytes were purified via a Ficoll density gradient centrifugation. Lineage negative cells were enriched using the EasySep™ Human Pan-ILC Enrichment Kit (StemCell Technologies). Cells were then stained for sorting or flow cytometry analysis. Tonsils were obtained from elective tonsillectomies (Children’s Hospital in Saint Louis) and provided as surgical waste with no identifiers attached. Tonsils were mechanically disrupted, passed through a 0.0059-inch opening sieve, and filtered extensively through 70-μm cell strainers. CD56+ tonsil cells were enriched via positive selection with CD56 microbeads (Miltenyi), according to the manufacturer’s recommendations. Lineage negative cells were enriched using the EasySep™ Human Pan-ILC Enrichment Kit (StemCell Technologies). Cells were then stained for sorting or flow cytometry analysis. Lung tissue was procured from organ donors and explants from transplant recipients suffering from pulmonary fibrosis. Patients provided informed consent. Lungs from healthy donors and fibrotic recipients were mechanically disrupted and digested in complete RPMI medium containing 1 mg ml–1 Collagenase IV (Sigma, C-5138) at 37 °C for 45 min under agitation. Cells were filtered through 70-μm cell strainers and stained for sorting or flow cytometry analysis. All ileum samples were provided as surgical waste with no identifiers attached on written informed consent to the Digestive Disease Research Cores Center (DDRC) at Washington University. Single-cell suspension of the mucosal tissue from the terminal ileum was separated from the muscular layer and serosa and cut into small pieces. Intraepithelial lymphocyte cells were extracted by rotating the tissue at room temperature for 40 min in Hank's balanced salt solution, 10% FCS, and 5 mM ethylenediaminetetraacetic acid (EDTA). Cells were filtered through 100-μm cell strainers, and dithiothreitol (DTT) was added at a final concentration of 5 mM. After intraepithelial lymphocyte removal, LPL cells were extracted by digesting tissue in complete RPMI medium containing 1 mg ml–1 Collagenase IV at 37 °C for 45 min under agitation. Cells were filtered and subjected to density gradient centrifugation using 40% and 70% Percoll solutions. Cells were collected, sorted, and processed for scRNA-seq or collected and stained for flow cytometry analysis. Some samples were processed and frozen for later flow cytometry analysis. Control patients for the present study were undergoing abdominal surgery for colon cancer or polyposis, which had the non-involved terminal ileum removed, as part of the surgical procedure. All human protocols were approved by the Institutional Review Board (IRB) at Washington University School of Medicine.

first: Natalia

last: Jaeger

first: Alina

middle: U

last: Antonova

first: Daniel

last: Kreisel

first: Florence

last: Roan

first: Erica

last: Lantelme

first: Steven

last: Ziegler

first: Marina

last: Cella


title: Effect of overexpress of lncFABP4 on gene expression during adipogenic differentiation of buffalo intramuscular adipocytes

city: Nanning

first: Huang

last: JiePing

address: Xixiangtang District

institute: Guangxi University

summary: To investigate the function lncFABP4 in the regulation of adipogenic differentiation, we overexpressed lncFABP4 in buffalo intramuscular adipocytes

overall_design: First, intramuscular adipocytes were planted in six-well plates in triplicate. When the cells reach 80% confluence, transduction was performed. Adenovirus were added to cells at the indicated MOI. Then, The culture medium was exchanged three hours later. Next, When transduction was performed for 48 hours, cells were treated with an inducing medium. Finally, Cells were collected for RNA sequencing after 48 hours of induction.

first: Jieping

last: Huang


title: SDS22 coordinates the assembly of holoenzymes from nascent protein phosphatase-1

first: Rekin's

last: Janky

address: Herestraat 49 Box 816

department: Nucleomics Core

institute: VIB

summary: RNAseq analysis of SDS22 degron cell lines

overall_design:
Two cell types are investigated under two conditions in three independent experiments, resulting in 12 samples (i.e. 2x2x3).
The two cell types are: HCT116 Tet-OsTIR1 (parental) and HCT116 Tet-OsTIR1 SDS22 WT (SDS22).
The two conditions are (1) untreated, (2) treated with  2 µg/ml doxycycline and 500 µM IAA.

first: Sarah

last: Lemaire

first: Aleyde

last: Van Eynde

first: Margareta

last: Nikolic

first: Mathieu

last: Bollen


title: Integrated single-cell analysis defines the epigenetic basis of castration-resistant prostate luminal cells (scATAC-seq)

city: Buffalo

first: Tao

last: Liu

address: Elm and Carlton St

department: Biostatistics and Bioinformatics

institute: Roswell Park Comprehensive Cancer Institute

summary: Understanding prostate response to castration and androgen receptor signaling inhibitors (ARSI) is critical to improving long-term prostate cancer (PCa) patient survival. Here we use a multi-omics approach on 229,794 single cells to create a mouse single-cell reference atlas better suited to interpreting mouse prostate biology and castration response. Our reference atlas refines single-cell annotations and provides chromatin context, which, when coupled with mouse lineage tracing demonstrates that the castration-resistant luminal cells are distinct from the pre-existent urethra-proximal stem/progenitor cells. Molecular pathway analysis and therapeutic studies further implicate JUN/FOS, WNT/b-Catenin, FOXQ1, NFkB, and JAK/STAT pathways as the major drivers of castration-resistant luminal populations with high relevance to human PCa. Importantly, we demonstrate the utility of our datasets, which can be explored through an interactive portal (https://visportal.roswellpark.org/data/tang/), to aid in developing novel combination treatments with ARSI for advanced PCa patients.

overall_design: Prostates from C57Bl/6 wild type mice were collected from intact, castrate, and testosterone regenerated mice at 12 weeks of age.  Mice were castrated for 4 weeks and regenerated for 96 hours with testosterone pellets.  Prostates were digested into single cells and analyzed using scRNAseq and scATACseq to understand prostate cell response to castration and regeneration.

first: Jason

last: Kirk

first: Jie

last: Wang

first: Amanda

last: Tracz

first: Mark

last: Long

first: Spencer

last: Rosario

first: Yibing

last: Ji

first: Rahul

last: Kumar

first: Xiaozhuo

first: Prashant

middle: K

last: Singh

first: Igor

last: Puzanov

first: Gurkumal

last: Chatta

first: Qing

last: Cheng

first: Jiaoti

first: Jeffrey

last: Wrana

first: Jonathan

last: Lovell

first: Han

last: Yu

first: Song

first: Michael

last: Shen

first: Dean

middle: G

last: Tang


title: Integrated single-cell analysis defines the epigenetic basis of castration-resistant prostate luminal cells (scRNA-seq)

first: Tao

last: Liu

first: Jason

first: Jie

first: Amanda

last: Huang

middle: M


title: Integrated single-cell analysis defines the epigenetic basis of castration-resistant prostate luminal cells.

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

overall_design: Refer to individual Series


title: FOXC1 in triple negative breast cancer [ChIP-seq]

city: Dubai

first: Fahad

last: Ali

address: Bldg 14, Dubai Healthcare City

country: United Arab Emirates

institute: Mohammed Bin Rashid University of Medicine and Health Sciences

summary: To investigate FOXC1 chromatin binding, and the effect of FOXC1 CRISPR knockout in triple negative breast cancer cell lines MDA-MB-231, MDA-MB-468, Hs578t, and Bt-549.

overall_design: We performed RNA-seq on WT and FOXC1 KO lines, and ChIP-seq for FOXC1 on WT cell lines.

first: Revathy

last: Ramachandran

first: Laura

last: Woods


title: FOXC1 in triple negative breast cancer


title: Age-related epithelial defects limit thymic function and regeneration [single cell; Foxn1Lin]

status: Public on Jun 27 2024

city: Duarte

first: Anastasia

middle: I

last: Kousa

state: CA

address: 1500 E Duarte Rd

department: Hematology / HCT

institute: City of Hope

summary: The thymus is essential for establishing adaptive immunity yet undergoes age-related involution that leads to compromised immune responsiveness. The thymus is also extremely sensitive to acute insult and although capable of regeneration, this capacity declines with age for unknown reasons. We applied single-cell and spatial transcriptomics, lineage-tracing and advanced imaging to define age-related changes in non-hematopoietic stromal cells and discovered the emergence of two atypical thymic epithelial cell (TEC) states. These age-associated (aa)TECs formed high-density peri-medullary epithelial clusters that were devoid of thymocytes; an accretion of non-productive thymic tissue that worsened with age, exhibited features of epithelial-to-mesenchymal transition (EMT), and was associated with downregulation of FOXN1. Interaction analysis revealed that the emergence of aaTEC drew tonic signals from other functional TEC populations at baseline acting as a sink for TEC growth factors. Following acute injury, aaTEC expanded substantially, further perturbing trophic regeneration pathways and correlating with defective repair of the involuted thymus. These findings therefore define a unique feature of thymic involution linked to immune aging and could have implications for developing immune boosting therapies in older individuals.

overall_design: Single cell RNA-seq profiles of CD45- thymic cells from Foxn1 lineage trace and wild type 20-month-old mice at steady state.

first: Lorenz

last: Jahn

first: Kelin

last: Zhao

first: Angel

middle: E

last: Flores

first: Dante

last: Acenas II

first: Emma

last: Lederer

first: Kimon

last: Argyropoulos

first: Andri

last: Lemarquis

first: David

last: Granadier

first: Kirsten

last: Cooper

last: D’Andrea

first: Julie

last: Sheridan

first: Jennifer

last: Tsai

first: Lisa

last: Sikkema

first: Amina

last: Lazrak

first: Katherine

last: Nichols

first: Nichole

last: Lee

first: Romina

last: Ghale

first: Florent

last: Malard

first: Hana

last: Andrlova

first: Enrico

last: Velardi

first: Salma

last: Youssef

first: Marina

last: Burgos da Silva

first: Melissa

last: Docampo

first: Roshan

last: Sharma

first: Linas

last: Mazutis

first: Verena

middle: C

last: Wimmer

first: Kelly

last: Rogers

first: Susan

last: DeWolf

first: Brianna

last: Gipson

first: Antonio

last: Gomes

first: Manu

last: Setty

first: Dana

last: Pe'er

last: Hale

first: Nancy

last: Manley

first: Daniel

middle: H

last: Gray

first: Marcel

last: van den Brink

first: Jarrod

last: Dudakov


title: FOXC1 in triple negative breast cancer [RNA-seq]

first: Fahad

last: Ali

first: Revathy

last: Ramachandran

first: Laura

last: Woods


title: Effect of BTK and GSK3 inhibition on gene expression in primary peripheral human B cells

first: Diego

last: Espinoza

address: 3400 Civic Center Boulevard

summary:
To investigate the BTK and GSK3 in the regulation of human B cell biology, we treated B cells from peripheral blood with either vehicle or BTK inhibitor or GSK3 in presence or absence of BCR cross-linking antibodies.
We then performed gene expression profiling analysis using data obtained from RNA-seq of those samples at 24hrs.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for B cells treated with either BTK inhibitor or GSK3 inhibitor

first: Rui

last: Li

first: Amit

last: Bar-Or


title: Cell state transitions are decoupled from cell division during early embryo development [I]

status: Public on Jun 13 2024

city: Boston

first: Kalki

last: Kukreja

summary: As tissues develop, cells divide and differentiate concurrently. Conflicting evidence shows that cell division is either dispensable or required for formation of cell types. To determine the role of cell division in differentiation, we arrested the cell cycle in zebrafish embryos using two independent approaches and profiled them at single-cell resolution. We show that cell division is dispensable for differentiation of all embryonic tissues during initial cell type differentiation from early gastrulation to the end of segmentation. However, in the absence of cell division, differentiation slows down in some cell types, and cells exhibit global stress responses. While differentiation is robust to blocking cell division, the proportions of cells across cell states are not but show evidence of partial compensation. This work clarifies our understanding of the role of cell division in development and showcases the utility of combining embryo-wide perturbations with single-cell RNA sequencing to uncover the role of common biological processes across multiple tissues.

overall_design: We arrested the cell cycle in zebrafish embryos at 6 hours post fertilization (hpf) using two independent approaches: a chemical approach (hydroxyurea and aphidicolin, HUA) and a genetic approach involving a loss-of-function mutation in the gene emi1 (allele hi2648). We tracked differentiation dynamics using single-cell RNA sequencing (scRNA-seq) on embryos spanning 6–24 hpf for HUA-treated and control embryos, and at 24 hpf for emi1 mutant embryos. Overall we have collected multiple replicates for control embryos (ABs) at 6, 8, 10, 14, 18, 21 and 24 hpf, for HUA treated at 8, 10, 14 and 24 hpf and for emi1 mutants at 24 hpf.  Details about the samples can be found in the supplementary file "sample_information.txt"

first: Allon

last: Klein


title: Role of Retinol Saturase (RetSat) in the intestine

city: Berlin

first: Michael

last: Schupp

address: Hessische Str. 3-4

department: Institute of Pharmacology

institute: Charite Universitätsmedizin Berlin

summary:
This study investigates the transcriptional consequences of an inducible deletion of RetSat in epithelial cells of the intestine.
Kiefer MF et. al, American Journal of Physiology-Endocrinology and Metabolism, 2024

overall_design: Male C57BL/6J mice with floxed RetSat alleles with (Cre+) or without (Cre-)  villin-Cre-ERT2 were treated with Tamoxifen for 5 days. Afterwards, mice were fed a high-fat diet for 14 weeks. At the end of the study, jenjunal tissue was subjected to RNA  isolation. RNA was DNAse digested and profilex by RNAseq.

first: Marie

last: Kiefer

first: Sascha

last: Wulff


title: Acidity perturbs IL-2 responsiveness, mTORC1 and c-Myc in CD8+ T cells

city: Lausanne

first: Isaac

last: Crespo

state: Vaud

address: Bugnon 25A

department: Oncology

country: Switzerland

institute: Ludwig Cancer Research,UNIL

summary: CD8+T cells play a critical role in tumor control but a range of barriers in the microenvironment including low pH can suppress their function.Here, we demonstrate that acidity dampens T-cell expansion mainly due to impaired IL-2 responsiveness, blunts cytokine secretion upon re-activation, and lowers the cytolytic capacity ofCD8+T cells expressing weak affinity TCR. We further reveal decreases in both mTORC1 signaling and c-Myc levels at low pH.Mechanistically, concomitantly to nuclear/cytoplasmic acidification, mTORC1 was disrupted in a Rheb-, Akt/TSC2/PRAS40-, GATOR1- and Lkb1/AMPK -independent manner, and c-Myc levels dropped due to both decreased transcription and higher proteasome-mediated degradation. In addition, lower intracellular levels of glutamine/glutamate/aspartate and elevated proline were observed with no apparent impact on mTORC1 or c-Myc. Overall, we conclude that, due to the broad disruption caused by acidity, multiple interventions will be required to restore T-cell function unless intracellular pH can be effectively restored.

overall_design: To investigate the impact of low pH on the CTL transcriptome, OT-I CTLs from 5 biological replicates were cultured at pH7.4, pH7 or pH6.6 in the presence, or absence, of 200IU/mL mIL-2 for 24 hours.

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