title: Identification of a Lipid Oxygen Radical Defense pathway and its epigenetic control [RNA-Seq]

city: Lausanne

first: Evarist

last: Planet

state: Vaud

address: Route Cantonale

country: Switzerland

institute: EPFL

summary: Membrane phospholipids are vulnerable to oxidative radicals, and uncontrolled lipid peroxidation affects cell viability. Cells have evolved quality control and defense mechanisms, of which the genetic regulation is not fully understood. Here, we identify what we have coined the Lipid Oxygen Radical Defense (LORD) pathway. It is epigenetically repressed by a complex comprising the KRAB-zinc finger protein ZNF354A, the scaffold protein KAP1/TRIM28, the histone methyltransferase SETDB1, and the transcriptional activator ATF2. Upon lipid peroxide accumulation, p38- and JNK-dependent phosphorylation of ATF2, KAP1/TRIM28, and ZNF354A leads to disassembly of the repressive complex, releasing ZNF354A from specific DNA loci and activating a network of protective genes, including NRF2 targets. The pathway affects the cellular sensitivity to oxidative stress and ferroptosis, revealing a previously uncharacterized layer of epigenetic control in lipid quality control and damage repair. This positions the LORD pathway as a promising therapeutic target for diseases linked to chronic inflammation, neurodegeneration and cancer.

overall_design: U2OS human osteosarcoma cells were transfected with control siRNA (siCtrl) or siRNA targeting ZNF354A (siZNF354A). Four independent biological replicates were prepared per condition. Total RNA was extracted 72 hours after transfection for mRNA sequencing to identify transcriptional changes upon ZNF354A depletion. For human ZNF354A, 3 different siRNA in 3’-UTR were tested and validated, from Qiagen: ZNF354A (TACATCCTTGAGAGAGATGTA), from Santa Cruz (sc-91974), ZNF354A (GTCACGTGATCATCAGAAA), and from Origene , ZNF354A (GAGGTTTATCCATGGAATTAAATACCT). A control siRNA, a sequence targeting the viral glycoprotein VSV-G (5’-ATTGAACAAACGAAACAAGGA-3’) was used.

first: Francisco S

last: Mesquita

first: Laurence

last: Abrami

first: Charlene

last: Raclot

first: Romain

last: Forey

first: Gisou

last: van der Goot


title: Identification of a Lipid Oxygen Radical Defense pathway and its epigenetic control [ChIP-Seq]

status: Public on Dec 15 2025

type: Genome binding/occupancy profiling by high throughput sequencing

overall_design: ChIP-seq in K562 cells expressing GFP or doxycycline-induced ZNF354A-HA, with or without 50 µM H₂O₂ for 4 h. Chromatin was crosslinked, sonicated, and immunoprecipitated with anti-HA beads (Pierce: 88837). Libraries were prepared with the NEBNext Ultra II kit and sequenced on the Element Aviti platform (PE150). Four replicates per condition.

first: Béatrice

last: Kunz

first: Charlène

first: Lucie

last: Bracq

first: Danica

last: Milovanovic

first: Olga

last: Rosspopoff

first: Filipe

last: Martins

first: Didier

last: Trono


title: Neural crest cell derived DKK1 modulates Wnt signalling in the second heart field to orchestrate cardiac outflow tract development.

city: Adelaide

first: John

last: Toubia

address: Frome Rd

country: Australia

institute: Centre for Cancer Biology

type: Expression profiling by high throughput sequencing

summary: Heart morphogenesis is highly complex, and depends on the generation of diverse cell types which interact with each other in an orchestrated manner to remodel the primitive heart tube into a functional organ. Cardiac outflow tract formation critically depends on continued contribution of cardiac progenitor cells from the anterior second heart field to ensure proper growth of the outflow tract. Prior to entering the outflow tract, neural crest cells migrate in close apposition to the second heart field and may play important roles in regulating second heart field growth dynamics, however the molecular mechanisms by which neural crest cells interact with the second heart field have remained elusive. Here, we discover that neural crest cells are a primary source of Dickkopf1 (DKK1), a secreted Wnt signalling inhibitor, which modulates Wnt signalling activity in the second heart field to impose a balance between progenitor maintenance and differentiation. Further, we identify the ubiquitin ligase NEDD4 as a critical regulator of DKK1 levels, with disruption of Nedd4 activity leading to outflow tract defects. In the context of disease pathogenicity, we show a novel human congenital heart disease variant of NEDD4 has lost the ability to ubiquitinate DKK1, and is associated with heart defects in a mouse model of the genetic variant. Our findings point to an unexpected role for neural crest cells acting as a rheostat of Wnt signalling activity in cardiac progenitors, identifying a new molecular pathway underpinning correct outflow tract morphogenesis, and a new causative factor of congenital heart disease.

overall_design: Tissue sections from embryonic day E9.5 mouse embryos were acquired and subjected to laser capture microdissection. A region containing the anterior second heart field and associated neural crest and pharyngeal endoderm tissue was laser dissected, and total RNA prepared. This was done from 3 independent wildtype and Wnt1-Cre;Nedd4fl/fl embryos. Prepared RNA was subjected to bulk RNAseq.

first: Sophie

last: Wiszniak

first: Quenten

last: Schwarz


title: Spec-seq data for human and macaque ZNF626 with replicates

city: Palo Alto

first: Zheng

last: Zuo

state: CA

address: Stanford

department: Genetics

country: USA

institute: Stanford university

summary: Systematic analysis of available ChIP-exo data of human zinc finger proteins identified ZNF626 as a prominent binder to the protein-coding region of the HERVK9 gag gene, with a pronounced enrichment peak within a ~950-1000 bp window within HERVK9-int consensus site. Since conventional motif prediction algorithms failed to pinpoint the exact binding site with high confidence, we employed Spec-seq—a high-resolution, sequencing-based biophysical assay—to comprehensively map ZNF626's binding specificity. Alongside with human ZNF626, we ran Spec-seq on macaque ortholog of ZNF626, and compared it with human ZNF626. The resulting motifs differences can be directly attributed to their contact residues differences in fingers 1, 5, 9, and 10.

sample_organism:
Macaca
Homo sapiens

platform_organism:
Macaca
Homo sapiens

overall_design: We designed a DNA binding sites library containing 18 triply randomized sequences spanning the entire 54 bp region of interest (positions 942-995), alongside 64 completely dissimilar sequences as a Non-specific background control. The resulting energy profiles and sequence logos for human ZNF626 unambiguously identified a 30 bp specific binding site (positions 967-995), which aligns perfectly with the expected motif length for a ten-finger protein.

first: Zuo

last: Zheng


title: Opposing functions of distinct regulatory T cell subsets in colorectal cancer

city: New York

first: Sneha

last: Mitra

state: NY

address: 1275 York Avenue

institute: Memorial Sloan Kettering Cancer Center

summary: Regulatory T (Treg) cells are considered major contributors to the growth of solid organ tumors, with the notable exception of colorectal cancer (CRC), despite the documented abundance of Treg cells in CRC. Here, we demonstrate that IL-10⁺ and IL-10⁻ Treg cells constitute two distinct subsets with opposing functions: IL-10⁺ Treg cells counteract, while IL-10⁻ Treg cells promote, the growth of genetically engineered CRC tumors. Our findings suggest that the tumor-suppressive function of IL-10⁺ Treg cells is mediated by their suppression of effector CD4⁺ T cell production of IL-17, a cytokine that directly stimulates CRC tumor cell proliferation. Consistently, IL-10⁺ Treg cells were more abundant in both mouse and human CRC tumors than in tumor-adjacent normal colon tissue, whereas IL-10⁻ Treg cells exhibited the opposite distribution. Furthermore, gene expression signatures associated with IL-10⁺ and IL-10⁻ Treg cells correlated with better and worse disease prognoses in human CRC, respectively. This functional dichotomy between Treg subsets provides a rationale for therapeutic strategies aimed at selectively targeting pro-tumoral Treg cells while preserving their anti-tumoral counterparts across various barrier tissue cancers that harbor both subsets.

overall_design: Mouse T cells and myeloid cells were sort purified from transcplanted AKP tumor, liver metastases, and lymph node metastases ten weeks after tumor transplantation.

first: Xiao

last: Huang

first: Dan

last: Feng

first: Emma

middle: S

last: Andretta

first: Nima

middle: B

last: Hooshdaran

first: Aazam

middle: P

last: Ghelani

first: Eric

middle: Y

last: Wang

first: Joe

middle: N

last: Frost

first: Victoria

last: Lawless

first: Aparna

last: Vancheswaran

first: Qingwen

last: Jiang

first: Christina

last: Leslie

first: Alexander

last: Rudensky


title: The Conserved Small Protein SniP3 is Involved in Nitrogen Stress Response and Metabolic Regulation among Cyanobacteria

city: Freiburg

first: Wolfgang

middle: R.

last: Hess

state: Baden-Würtemberg

address: Schänzlestrasse 1

department: Institute of Biology III

country: Germany

institute: University of Freiburg

summary:
Cyanobacteria are oxygenic phototrophs that are regularly exposed to fluctuating environmental conditions. Particularly the availability of CO2 and combined nitrogen sources vary in many aquatic habitats. In the present study we investigated the role of SniP3, a well conserved, 3 kDa nitrogen starvation-induced protein, which is mainly localized on thylakoid membranes.
Deletion of sniP3 resulted in a non-bleaching phenotype and reduced glycogen accumulation under N-limited conditions. Many more distinct metabolic alterations were observed, whereas the signaling molecule 2-oxoglutarate and the overall transcriptome remained comparable in mutant and wild-type cells under N-replete and N-deplete conditions. The SniP3 interactome revealed glyceraldehyde-3-phosphate dehydrogenase (Gap2) as main specific interaction partner. Gap2 activity was lowered in cells of the sniP3 mutant, while the activity of recombinant Gap2 was stimulated in the presence of the soluble N-terminal part of SniP3. Corresponding to the interaction of SniP3 and Gap2, the sniP3 mutant was unable to grow heterotrophically with glucose in the dark. Collectively, our result imply that the SniP3 protein plays an important role in the regulation of the cyanobacterial metabolism under fluctuation C/N conditions.

overall_design: Total RNA (5 µg) was extracted from the WT and ∆sniP3 mutant grown at 100 μmol photons m-2 s-1 in BG11 medium and after a shift to nitrogen-free BG1-N medium for 48 h. Total RNA was DNase-treated using TURBO DNase (Thermo Fisher Scientific). 2 µg DNase-treated RNA were labeled with Cy3 using the ULS Fluorescent Labeling Kit for Agilent arrays (Kreatech). A high-resolution microarray manufactured by Agilent (Design ID 075764, format 8 × 60 K, slide Layout = IS-62976-8-V2) was used. 600 ng labeled RNA were used per array for hybridization for 17 h at 65 °C.

first: Luna

last: Alvarenga-Lucius

first: Sandra

last: Maaß

first: Viktoria

last: Reimann

middle: R

first: Martin

last: Hagemann


title: Effect of CDK8/19 kinase inhibitor SNX 631 on OV90 parental and anoikis resistant isogenic lines

city: COLUMBIA

first: HAO

last: JI

state: SC

address: 715 SUMTER ST CLS RM 109

department: College of Pharmacy

institute: University of South Carolina

summary: Anoikis resistance or evasion of cell death triggered by matrix detachment is a hallmark of cancer cell survival and metastasis. We show that repeated exposure to suspension stress followed by recovery under attached conditions leads to development of anoikis resistance. The acquisition of anoikis resistance is associated with enhanced invasion, chemoresistance, and immune evasion in vitro and distant metastasis in vivo. This acquired anoikis resistance is not genetic, persisting for a finite duration without detachment stress, but is sensitive to CDK8/19 Mediator kinase inhibition that can also reverse anoikis resistance. Transcriptomic analysis reveals that CDK8/19 kinase inhibition induces bidirectional transcriptional regulation in both sensitive and resistant cells disrupting the balanced reprogramming required for anoikis adaptation and resistance and reversing some resistance associated pathways while enhancing others. Both anoikis resistance and in vivo metastatic growth of ovarian cancers are sensitive to CDK8/19 inhibition, thereby providing a therapeutic opportunity to both prevent and suppress ovarian cancer metastasis

overall_design: Transcriptomic profiling of the effect of CDK8/19 inhibition using SNX631 on OV90 parental ( anoikis sensitive) and adapted isogenic OV90 resistant lines in attached or suspension cultures with either pretreatment in 2D culture alone or co- treatment with suspension culture in adapted resistant lines.

first: Hao

last: Ji

first: Mengqian

last: Chen

first: Mythreye

last: Karthikeyan

first: Resha

last: Rajkarnikar


title: HIV-1 Tat protein exposure alters the morphological characteristics and gene expression in the primary mouse cortex endothelial cells and human brain microvascular endothelial cells

city: Kodaira

first: Lili

last: Quan

state: Tokyo

address: 4-1-1 Ogawa-Higashi

department: National Institute of Neuroscience

country: Japan

institute: National Center of Neurology and Psychiatry

summary: HIV-1 viral proteins can disrupt tight junctions within the blood-brain barrier (BBB), directly compromising its structural and functional integrity. In this study, we investigated the effects of HIV-1 transactivator of transcription (Tat) protein on mouse prefrontal cortex endothelial cells (ECs) morphological alterations and gene expression profiles. Our results demonstrated that Tat-exposed ECs showed downregulated CD31 expression, increased phalloidin staining intensity, and activation of inflammatory response pathways as revealed by RNA-seq analysis, which is similar to the effects of Tat on human brain microvascular endothelial cells (HBMVECs).

overall_design: Mouse ECs were cultured in the culture medium [DF12 (120 mg/mL), NaHCO3 (12 mg/mL), 1% Penicillin/Streptomycin, 10% FBS, Heparin (5 µg/mL), Insulin (5 µg/mL), Sodium selenite (5 ng/mL), 100 nM Hydrocortisone, Apo-transferrin (5 µg/mL), bFGF (1 µg/mL), and Puromycin (1 µg/mL), pH 7.2] and maintained at 37 °C in a 5% CO2 incubator for 10 days before Tat exposure. Cells were seeded on 96-well plate at a density of 10,000 cells/well and exposed to control medium containing 0.1% protease inhibitor cocktail for preventing Tat protein degradation or the same medium containing recombinant Tat1-86 at a 12.5 nM final concentration and incubated at 37°C, 5% CO2 incubator for 48 hours before RNA isolation.

first: Rieko

last: Muramatsu

first: Jun

last: Zhu


title: Bridging the gap of late-gestation nephrogenesis using a non-human primate model

city: Cincinnati

first: Kairavee

last: Thakkar

address: 3333 Burnet Ave

institute: Cincinnati Children's Hospital Medical Center

summary:
Background: Prematurity is associated with low nephron endowment and an increased risk of chronic kidney disease. Human nephrogenesis is complete at 34-36 weeks gestation, with 60% of nephrons forming during the third trimester through lateral branch nephrogenesis (LBN). We hypothesized that a differentiated but dividing population of nephron progenitor cells (NPCs) would contribute to the amplification of nephrons in late gestation.
Results: scRNA-Seq of 64,782 rhesus cells revealed 37 transcriptionally distinct cell populations, including 7,879 rhesus NPCs. Pseudotime analyses identified a late gestation-specific lineage branch of differentiated NPC in rhesus that was not observed in mid-gestation humans. Differential expression analyses identified increased SFRP1, FZD4, and TLE2 and decreased FZD7, SHISA2, SHISA3, and TLE4 within the late-gestation rhesus NPC compared to mid-gestation human NPC and increased SEMA3D within the rhesus UB tip, suggesting a compositional shift in WNT and SEMA signaling components within the naive NPC population during LBN.
Conclusion: The rhesus macaque uniquely enables molecular studies of late-gestation primate nephrogenesis.
Our study suggests the hypothesis that a transitional state of self-renewing NPC supported by compositional shifts in key pathways may underlie the switch from branching phase nephrogenesis to lateral branch nephrogenesis and support ongoing nephron formation in late gestation.

overall_design: Single-cell RNA-sequencing (scRNA-Seq) was performed on cortically-enriched fetal rhesus kidneys (n=9) from late second trimester and third trimester during LBN. This data was integrated with publicly available human scRNA-seq datasets from 8-18 weeks gestation kidneys (n=8) using state-of-the-art bioinformatics pipelines. Differentially expressed genes and ligand-receptor interactions were assessed and validated using RNAScopeTM on human and rhesus archival tissue.

first: Meredith

last: Schuh


title: Comparison of Multi-MiRNA Models from Donor-Matched Plasma and Cerebrospinal Fluid as Candidate Biomarkers for Alzheimer’s Disease Prediction

city: Portland

first: Julie

middle: A

last: Saugstad

state: Oregon

address: 3181 SW Sam Jackson Park Road

department: Anesthesiology & Perioperative Medicine

institute: Oregon Health & Science University

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

summary:
To validate candidate miRNA biomarkers for AD in donor and date matched CSF and plasma from a cohort of healthy control and AD donors;
We obtained 320 donor- and date-matched normal control (NC) and AD CSF and plasma samples from the AD Neuroimaging Initiative 2, then analyzed 57 potential AD miRNA biomarkers in both biofluids by RT-qPCR.

overall_design:
CSF and plasma samples from 160 participants (80 AD and 80 NC; 320 total) were obtained from ADNI 2.
We used CSF and plasma samples from living donors obtained from the Alzheimer's Disease Neuroimaging Initiative 2 to validate the expression of 57 candidate Alzheimer's Disease vs Healthy control miRNA biomarkers. The miRNAs were quantified on custom Taqman Low Density miRNA Arrays from ThermoFisher Scientific.
Please note that 32 samples are excluded from the final analysis as detailed in the sample data processing field (i.e. normalized_data.txt contains

first: Ursula

last: Sandau

first: Joseph

last: Quinn

first: Jack

last: Wiedrick

first: Siting

first: Trevor

last: McFarland


title: Chronic compression recapitulates transcriptional, metabolic, and functional state changes in macrophages associated with pro-tumor phenotypes

city: Notre Dame

first: Alice

last: Burchett

state: IN

address: 2030 McCourtney Hall East

institute: University of Notre Dame

summary: Macrophages comprise a significant portion of the glioblastoma tumor microenvironment and are essential in promoting immunosuppression and tumor progression. Solid tumors such as glioblastoma generate solid stress as they expand, creating a compressive microenvironment for mechanosensitive immune cells including macrophages. Macrophages are known to respond to various mechanical stimuli but have not yet been studied in the context of chronic compression observed in growing tumors. Here, we used a custom in vitro compression system to elucidate the effects of compressive solid stress on murine macrophages. We found that macrophages have significant morphological, transcriptional, metabolic, and functional responses to compression. These changes corresponded to both canonical pro- and anti-inflammatory macrophage states. The gene expression signatures of compressed macrophages more closely resembled those of glioma-associated macrophages known to be associated with worse patient outcomes. These results indicate that compression alone, independent from tumor cell-derived biochemical factors, may contribute to the pathological tumor-associated macrophage phenotype. This could represent a vicious cycle of tumor immunomechanics and mechano-immunology. Targeting solid stress in tumors or the response to solid stress by macrophages may interrupt this feedback loop to help normalize the tumor immune microenvironment and improve glioblastoma response to immunotherapy.

overall_design: RNA-seq profiling of RAW264.7 macrophages under 24 hours of statioc compressive solid stress (weighted) compared to uncompressed controls (agarose).

first: Julian

last: Najera

first: Sc ott

last: Howard

first: Meenal

last: Datta


title: Glc7/PP1 triggers Paf1 complex dissociation from RNA polymerase II to enable transcription termination

city: Pittsburgh

first: Karen

middle: M.

last: Arndt

state: Pennsylvania

address: 4249 Fifth Avenue

department: Biological Sciences

institute: University of Pittsburgh

summary: The mechanisms that control the dynamic composition of RNAPII elongation complexes govern major transitions in the transcription cycle yet are poorly understood. Here, we show that the transcription elongation factor Spt5 determines elongation complex composition to promote productive elongation and the transition to termination. Using an unbiased genetic screen and genomic approaches in Saccharomyces cerevisiae, we provide evidence that dephosphorylation of the Spt5 C-terminal repeat domain (CTR) by Glc7/PP1 is required to dislodge the Paf1 complex (Paf1C) from RNAPII near the cleavage and polyadenylation site (CPS). Mutations in Paf1C or the Spt5 CTR that dissociate Paf1C from RNAPII bypass the requirement for two critical regulators of Glc7 in the cleavage and polyadenylation factor that promote Glc7 enrichment at the 3’ ends of genes. Depletion of Glc7 causes aberrant retention of Paf1C past the CPS and a dramatic increase in readthrough transcription, which is fully suppressed by Paf1C mutations. Our results demonstrate that Paf1C retention antagonizes transcription termination and that Glc7-mediated restructuring of the RNAPII elongation complex to evict Paf1C at the CPS is a critical step in the transition from elongation to termination.

overall_design: 4tU-seq to investigate nascent transcription in mAID-Spt5 depletion strains, spt5 mutants, HA-Glc7 mutants and Glc7-anchor-away strains.

first: Sanchirmaa

last: Namjilsuren

middle: M


overall_design: Saturated Transposon Analysis in Yeast using WT, spt5∆5, spt5∆CTR and spt5-S1-15A mutant S. cerevisiae strains. After transposition induction over 18-20 days, the genomic DNA regions flanking the transposon insertion site were sequenced.


overall_design: ChIP-seq to probe occupancies of RNA polymerase II and elongation factors in mAID-Spt5 depletion strains, spt5 mutants, HA-Glc7 mutants and Glc7-anchor-away strains.


title: Extremes on the benign-malignant tumour spectrum: Distinct transcriptomic landscapes between two common canine perianal neoplasms based on the hallmarks of cancer [nCounter®]

city: Berlin

first: Alexander

middle: F. H.

last: Haake

address: Robert-von-Ostertag-Straße 15

department: Department of Veterinary Medicine

institute: Freie Universität Berlin

summary: The hallmarks of cancer describe stepwise changes in cellular functions and molecular pathways of tumour cells that undergo malignant progression. Although 14 hallmarks have been proposed, their manifestation at the molecular level across different tumour types remains incompletely understood. Here, we hypothesised that hallmark-associated transcriptomic signatures reflect the contrasting biological behaviours of a pair of a benign and a malignant tumour. To this end, we compared the two most common canine perianal tumours, the highly malignant apocrine gland anal sac adenocarcinoma (AGASAC) and the benign hepatoid gland adenoma (HGA). Spontaneous primary tumours were analysed using RNA-Seq (AGASAC n = 11, HGA n = 10) and a direct mRNA hybridisation panel (AGASAC n = 28, HGA n = 12). Large transcriptomic differences were observed in both the number of differentially expressed genes and enriched pathways. Surprisingly, transcriptomic differences based on the hallmarks of cancer were less pronounced that anticipated, particularly for “activating invasion and metastasis” and “inducing or accessing vasculature”. Only the HGAs showed a significant enrichment for “deregulating cellular energetics”. However, our data point towards a contrasting immune landscape between the compared tumours and a subgrouping of AGASACs associated with the inflammatory landscape. Overall, the transcriptomes of these two benign-malignant tumour extremes on the tumour spectrum contrasted each other in select aspects of the hallmarks. Still, their malignant or benign behaviours could not generally be mirrored on the mRNA level.

overall_design:
FFPE tissue samples of AGASACs (n = 33; 28 primary tumours, 5 nodal metastases) and HGAs (n = 12) from pet dogs were retrospectively obtained from the archive (spanning the years 2006 to 2021) and total RNA was extracted. The RNA was hybridised to the nCounter® Canine IO Panel XT CodeSets and a 30 probe Panel Plus. Differential gene expression (DGE) was implemented in the nSolverTM 4.0 Analysis Software with the R 3.3.2-based Advanced Analysis 2.0 plug-in (version 2.0.134) for AGASAC primary tumour (PT) versus baseline of HGA and AGASAC PT versus AGASAC nodal metastases (NM).

Attention: This GEO submission only includes the nCounter® data. The 3' RNA-Seq data used in the publication were submitted separately to GEO (GSE302161). 25 samples in this submission are re-used/re-analyzed (see GSE261791), please see "Description" under each sample for the cross-reference.

middle: F

first: Alina

middle: K

last: Loriani Fard

first: Vladimir

last: Jovanovic

first: Michael

last: Hummel

first: Sandro

last: Andreotti

first: Achim

middle: D

last: Gruber


title: Heterogeneity in  populations from subcutaneous abdominal and thigh adipose tissues of women

first: Yifei

last: Sun

address: 1468 Madison Ave

institute: Icahn School of Medicine at Mount Sinai

summary: Women with a ‘pear-shaped’ fat distribution are at lower risk to develop insulin resistance and type 2 diabetes, but the mechanisms involved remain poorly understood. The objective of this study was to  determine if the  adipose cell populations differed between abdominal subcutaneous adipose tissue  (ASAT) and lower body thigh (TSAT) adipose tissue of clinically healthy,  premenopausal women with body mass indices (BMI) in the non-obese to obese range.  Paired samples of A- and T- SAT were taken under local lidocaine anaesthesia and quick frozen in lquid nitrogen.  Single nuclei RNA sequencing (snRNASeq) was used to identify and major cell populations and their subsclusters. We found differences in subcluster cell numbers and identified  differentially expressed genestthat may contribute to depot differences in metabolism as well as  expansion and homeostatic remodeling capacities through cell cell communucation.  These variations likely contribute to protective effect of lower body fat on metabolic health and T2D risk.

overall_design: Paired samples of Subcutaneous Adipose tissues were collected  from same subject's  Abd and Thi were collected  for snRNAseq

first: Gregory

last: Smith

first: Jeanine

last: Albu

first: Kalyspo

last: Karastergiou

first: Bingyao

last: Wang

first: Ofer

last: Modlavski

first: Martin

last: Walsh

first: Susan

last: Fried


title: Molecular Signatures and Signaling Interactions of the Hair Follicle Stem Cell Niche

last: Rendl

address: 1428 Madison Avenue

institute: Mount Sinai School of Medicine

summary: Signaling interactions between the dermal papilla (DP) and neighboring stem cells in the hair germ (HGSCs) and bulge (BuSCs) regulate new follicle growth in the hair cycle. To study these interactions, the three populations had been profiled together by now-outdated microarrays or separately by bulk RNA-sequencing. Recent single-cell transcriptomics established signatures of DP, BuSCs and HGSCs, but low detection sensitivity limited the depth of gene expression discovery. Here, we define the transcriptomes of DP, BuSCs, HGSCs, epidermal, follicle and dermal fibroblast cells – after flow-sorting each population from four neighboring mouse back skin regions – to gain deeper insights into the unique gene expression programs of SCs and their instructive niche. With cross-comparisons of 56 whole-transcriptome measurements, we classify cell type-specific molecular signatures of enriched genes with unprecedented sensitivity. Joint analysis with signatures from 15 leading studies published in the last 20 years revealed many previously undescribed DP, BuSC and HGSC genes in mouse and human counterparts. With ligand-receptor mapping and CellChat analyses we then uncover comprehensive cell-cell communication insights. Finally, we provide a new installment of our Hair-Gel repository along with numerous signature and other gene tables for easy exploration of gene expression in hair follicle SCs and their niche.

overall_design: Bulk RNA-seq was performed on FACS isolated Dermal Papilla (DP), Dermal Fibroblasts (DF), Epidermis (Epi), Follicle Cells (Foll), Bulge Stem Cells (BuSC) and Hair Germ Stem Cells (HGSC), and a negative fraction (Neg) from 4 individual back skin sections Z1-Z4 at P22 of 2 biological replicates using the IlluminaHiSeq 2000 platform.

first: Sangeeta

last: Ghuwalewala

first: Jenny

last: Cao

first: Amelie

last: Rezza

first: Martina

last: Rangl

first: Alexia

last: Brown

first: Laura

last: Grisanti

first: Nicholas

last: Heitman

first: Markus

last: Schober

first: Zichen

first: Avi

last: Ma’ayan

first: Axel

last: Almet

first: Maksim

last: Plikus


title: Comprehensive interrogation of chemo-genomic interactions between DNA damaging agents and DNA damage repair genes with targeted CRISPR knockout screening

city: West Haven

first: Francesc

last: Lopez

state: CT

department: Department of Genetics

institute: Yale University

summary: Genetic instability is a hallmark of cancer, often arising from mutations to DNA damage repair and response (DDR) genes. Classical genetic, biochemical and structural approaches have elucidated the foundational mechanisms of DDR pathways and provided a scientific understanding of their involvement in repair of broad classes of DNA-damaging agents (DDAs). However, given the chemical diversity of DDAs and resultant DNA lesions, along with the multitude of interconnected DDR factors, the chemogenomic landscape of DDA–DDR interactions remains incompletely mapped. To this end, we developed a DDR-targeted, CRISPR knockout screening approach and assessed relationships among 353 DNA repair genes and 15 DDAs in LN229 glioma cells. Within this dataset of 5295 DDR-related chemogenomic interactions, we identified many established interactions and discovered novel ones. For example, we observed a specific role of TC-NER in the repair of adducts generated by monofunctional alkylating agents, a role for the FA pathway in addressing methyl lesions, overt differences in DSB repair following treatment with topoisomerase I and II poisons, and repair dependencies associated with imidazotetrazine derivatives temozolomide, mitozolomide, and KL-50. Future directions will continue to investigate the mechanisms of novel chemogenomic interactions that we have uncovered as well as work to identify chemogenomic interactions amenable to clinical translation.

overall_design: NOT PROVIDED; REQUESTED

first: Collin

last: Heer

first: James

middle: L

last: Elia

first: Spenser

last: Johnson

first: Vijay

last: Menon

first: Sofia

last: Arbelaez

first: Sam

last: Friedman

last: Lopez-Giraldez

first: Seth

last: Herzon

first: Ranjit

last: Bindra

middle: E

last: Gueble


title: Dual Epigenetic and Nuclear Export Inhibition by Chidamide and Selinexor in High Grade B-Cell Lymphomas via Survivin and PI3K/AKT Inhibition

city: Xiamen

first: jiazhen

last: lin

state: Fujian

address: No. 2000 of Xiang an East road

country: China

institute: Xiamen uniersity

summary: Double hit lymphoma (DHL) is a highly aggressive disease thatis resistant to conventional first-line immunochemotherapeutic regimens. This resistance necessitates the exploration of innovative therapeutic strategies. In this study, the combination of chidamide and selinexor showed significant synergistic antilymphoma effects in the treatment of DHL. The synergistic effects were evidenced by the inhibition of cell proliferation, induction of apoptosis, and perturbation of the cell cycle in cell lines, as assessed by Cell Counting Kit-8, Annexin V/PI staining, and PI staining assays. Furthermore, in a xenograft mouse model of DHL, this combination therapy  markedly reduced the tumor burden without causing lethal toxicity. At the mechanistic level, the combination of chidamide and selinexor resulted in the synergistic downregulation of survivin and the PI3K/AKT signaling pathway. This dual inhibition was attributed to the interactive effects of the two drugs. The downregulation of key downstream targets of the PI3K/AKT pathway, including c-Myc, MCL1, BCL-XL, cyclin A2, and survivin, was synergistic and aligned with the phenotypic outcomes. Notably, survivin, an antiapoptotic protein and a direct target of HDAC3 and XPO1, was synergistically downregulated at the nuclear, cytoplasmic, and total protein levels by the combination of chidamide and selinexor, thereby inducing apoptosis. Our preclinical data highlighted the potential synergistic efficacy of chidamide and selinexor in targeting DHL, providing a rationale for further clinical investigation of this therapeutic combination for the treatment of this refractory disease.

overall_design: TMD8 cell were treated with DMSO, chidamide, selinexor and chidamide combined with selinexor for 24h.The RNA from four groups of cells was extracted and subjected to RNA sequencing.

first: Jizhen

last: Lin


title: RNA sequence for RMG1 cells with HIF2A knockdown under 1.5% O₂ for 24 hours

city: kyoto

first: MENGXIN

last: JIANG

address: 54 Kawahara-cho, Shogoin, Sakyo-ku

department: Department of Gynecology and Obstetrics

institute: Kyoto University

summary: We performed RNA-seq analysis on ovarian clear cell carcinoma (OCCC) cell lines to investigate the transcriptional effects of HIF2A knockdown. Hypoxia-inducible factor 2 alpha (HIF2A) plays a key role in cellular adaptation to low oxygen environments and has been implicated in tumor progression. In this study.A human OCCC cell line,  RMG1, was cultured under hypoxic conditions following lentiviral transduction with shHIF2A or a control vector. This dataset provides insights into the downstream gene expression changes and mitochondrial function modulated by HIF2A suppression.

overall_design: RMG1 cell line was transduced with lentiviral vectors expressing either shHIF2A or shcontrol. After selection, the cells were cultured in F-12 with 10% FBS under 1.5% O₂ for 24 hours. Total RNA was extracted and subjected to paired-end sequencing using the Illumina NovaSeq 6000 platform. Each condition included biological replicates.

first: Mengxin

last: Jiang

first: Ken

last: Yamaguchi

first: Kohei

last: Hamada

first: Yuko

last: Hosoe

first: Yuka

last: Mise

first: Sachiko

last: Kitamura

first: Zhihong

last: Liu

first: Hairong

last: Wei

first: Zhenhai

last: Gao

first: Mana

last: Taki

first: Koji

last: Yamanoi

first: Ryusuke

last: Murakami

first: Rin

last: Mizuno

first: Taito

last: Miyamoto

first: Junzo

last: Hamanishi

first: Masaki

last: Mandai


title: Deletion of neuronal Idol ameliorates Alzheimer's disease–related pathologies via APOE receptors

city: Indianapolis

first: Luke

middle: Child

last: Dabin

state: Indiana

department: Medical and Molecular Genetics

institute: Indiana University

summary:
Overexpression of the low-density lipoprotein receptor (LDLR) is known to decrease apolipoprotein E (APOE) levels and alleviate amyloid beta (Aβ) pathology. We hypothesized that inhibiting the Inducible Degrader of LDLR (IDOL), an enzyme that ubiquitinates LDLR for degradation, would increase endogenous LDLR levels and attenuate amyloid pathology.

To investigate the cell-type–specific role of IDOL, we generated Idol conditional knockout mice on an Aβ-amyloidosis mouse model and performed biochemical, histological, and multi-omics analyses.

We demonstrated that neuronal, but not microglial, Idol deletion reduced amyloid accumulation and altered brain LDLR and APOE levels, indicating the critical role of neuronal IDOL-LDLR in amyloid pathology. In addition, neuronal Idol deletion increased the levels of Reelin receptors important for synaptic function, and single-nuclei RNA sequencing revealed significant changes associated with synaptic organization.

overall_design: Posterior cortices of 2 female 5XFAD mice and 2 female 5XFAD mice with a neuron-specific loss of IDOL were processed for single nucleus RNA sequencing

first: Karahan

last: Hande

first: Hartigan

last: Kelly

first: Al-Amin

last: Md Mamun

first: John

last: Sutha K.

first: McCord

last: Brianne

first: Wijeratne

last: H.R. Sagara

first: Acri

last: Dominic J.

first: Smith

last: Daniel C.

first: Dabin

last: Luke C.

first: Cordero

last: Hannah M. Rondon

first: Kim

last: Byungwook

first: Lee

last: Do-Hun

last: Jungsu


title: RNA-seq of human ovarian clear cell carcinoma PDX tumors

first: MENGXIN

last: JIANG

summary: This dataset contains RNA-seq profiles of patient-derived xenograft (PDX) tumors established from human ovarian clear cell carcinoma (OCCC). Tumor tissues were collected without any therapeutic treatment and used to generate xenografts in immunodeficient mice. Total RNA was extracted from the PDX tumors and sequenced on the Illumina NovaSeq 6000 platform using 150 bp paired-end reads. These data represent the baseline transcriptomic landscape of OCCC tumors and may serve as a resource for studying tumor biology and preclinical model characterization.

overall_design: This experiment was designed to profile the baseline transcriptomic landscape of human ovarian clear cell carcinoma (OCCC) using patient-derived xenograft (PDX) models. Tumor tissues from OCCC patients were implanted into immunodeficient mice to establish PDX models. No therapeutic treatment was applied. Tumors were harvested at the baseline timepoint, and total RNA was extracted and sequenced using paired-end 150 bp reads on the Illumina NovaSeq 6000 platform. The resulting data provide a reference transcriptomic map for human OCCC in PDX models.

first: Mengxin

last: Jiang

first: Ken

last: Yamaguchi

first: Kohei

last: Hamada

first: Yuko

last: Hosoe

first: Yuka

last: Mise

first: Sachiko

last: Kitamura

first: Zhihong

last: Liu

first: Hairong

last: Wei

first: Zhenhai

last: Gao

first: Mana

last: Taki

first: Koji

last: Yamanoi

first: Ryusuke

last: Murakami

first: Rin

last: Mizuno

last: Miyamoto


title: Decoding the heterogeneity of human undifferentiated spermatogonia reveals RAS-dependent regulation of stem cell fate

city: La Jolla

first: Kun

last: Tan

address: 9500 Gilman Drive # 0695

department: Dept. of Obstetrics, Gynecology & Reproductive Science

institute: UCSD

summary: Undifferentiated spermatogonia (uSPG), including spermatogonial stem cells, are essential for long-term spermatogenesis and hold therapeutic potential for treating male infertility. While rodent uSPG are well characterized, human uSPG remain poorly understood. Here, we screened antibodies against 21 proteins encoded by genes enriched in human uSPG subsets identified from single-cell RNA sequencing analysis. Among them, we identified FSD1 as a pan-uSPG cell-surface marker suitable for purifying the entire uSPG population. Transcriptomic profiling of FSD1+ uSPG uncovered candidate signaling pathways and transcriptional networks involved in uSPG development. PIWIL4 and EGR4 label primitive, quiescent uSPG, whereas PPP1R36 and NANOS3 mark proliferative uSPG subsets. PIWIL4+ and NANOS3+ cells do not overlap, highlighting distinct uSPG developmental states. We provide empirical evidence that RAS signaling regulates the transition between these states. Together, our study provides valuable tools for studying uSPG development and facilitating future translational advances.

overall_design: FSD1+ cells were purified from testis biopsies from four individuals by FACS and performed bulk RNAseq. For comparison, bulk RNAseq analysis was performed in parallel on unfractionated human testicular cells from the same donors.

first: Chiara

last: Capponi


title: Free-floating Long-term Vascularized Mesenchymal Organoids [RNA-seq]

city: Atlanta

first: Alejandro

last: de Janon

state: GA

address: 225 North Ave NW

department: Biomedical Engineering

institute: Georgia Institute of Technology

summary: The vasculature is essential for tissue function and pathology. Spheroid co-cultures of endothelial and mesenchymal stem/stromal cells (MSCs) form consistent structures, but the vascular components are short-lived. iPSC-derived  vascular organoids can establish complex vasculature but often have cell heterogeneous variable maturation and low reproducibility. We presents consistently-formed, free-floating, long-term Vascularized Mesenchymal Organoids (VMOs), formed by co-culturing human umbilical vein endothelial cells (HUVECs) and MSCs in a pre-gelled minimal Matrigel scaffold.  We studied organoid dynamics over 60 days culture time and used spheroids cultured for 14 days as control.

overall_design: RNA was extracted from pooled samples (organoids) cultured for 3, 14, 28  and 60 days respectively. RNA was also extracted from day 14 spheroids (HUVEC + MSC no ECM scaffold)

first: Shuichi

last: Takayama


title: A role for the cholinergic neuron circadian clock in RNA metabolism and mediating neurodegeneration [Bmal1-KO]

city: Chicago

first: Sharon

last: Tam

state: IL

address: 303 E Chicago Ave, Rm 13-270

institute: Northwestern University

summary: Circadian clocks are encoded by a transcription-translation feedback loop that aligns physiological processes with the solar cycle. Previous work linking the circadian clock to the regulation of RNA-binding proteins (RBPs) provides a foundation for the vital examination of their mechanistic connections in the context of amyotrophic lateral sclerosis (ALS)—a fatal neurodegenerative disease commonly marked by disrupted RBP function. Here, we reveal that the spinal cord cholinergic neuron rhythmic transcriptome is enriched for genes associated with ALS and other neurodegenerative diseases. We show that there is time-of-day-dependent expression of ALS-linked RBP transcripts and rhythmic alternative splicing of genes involved in microtubule cytoskeleton organization, intracellular trafficking, and synaptic function. Through in silico analysis of RNA sequencing data from sporadic ALS patients, we find that gene expression profiles altered in disease correspond with rhythmic gene networks. Finally, we report that clock disruption through cholinergic neuron-specific deletion of clock activator BMAL1 increases neurodegeneration and drives time-of-day-dependent alternative splicing of RNA processing genes. Our results establish a role for the cholinergic neuron circadian clock in RNA metabolism and mediating neurodegeneration.

overall_design: RNA-seq profiling of Bmal1 knockout mouse spinal cord cholinergic neurons and controls collected at ZT8 and ZT20

first: Nathan

middle: J

last: Waldeck

first: Matthew

last: Wright

first: Erin

last: Baker

first: Evangelos

last: Kiskinis

first: Joseph

last: Bass

first: Robert

middle: G

last: Kalb

first: Jelena

last: Mojsilovic-Petrovic


title: Role of CTGF-LRP1 in Impaired Healing of Cesarean Section Incisions

city: Shanghai

first: Chuqing

last: He

address: No. 910, Hengshan Rd

department: Department of Obstetrics and Gynecology

institute: School of Medicine, Shanghai Jiaotong University

summary: Niche is a long-term complication following a cesarean section, which may increase the risk during subsequent pregnancies, and its pathogenesis remains incompletely understood. In this study, we utilized single-cell RNA sequencing to analyze cells obtained from adjacent myometrium tissue of the niche (Adjacent), well-healed cesarean scar tissue (Control), and niche tissue (Niche). Our analysis revealed that fibroblasts (FB) could be classified into three subgroups. FB3 in the Niche exhibited a reduced capacity for extracellular matrix synthesis. Cell communication analysis has identified that the secretion of CTGF by Endo3 and the expression of LRP1 by FB3 are involved in the receptor-ligand interaction of uterine fibroblasts during wound healing. Our research findings were supported by tissue staining and in vitro experiments, indicating that silencing LRP1 may prevent CTGF from promoting the synthesis of extracellular matrix by primary uterine fibroblasts. Furthermore, animal models have indicated that CTGF can promote the healing of uterine scars. These results provide insights into the cellular environment and mechanistic background underlying the formation of the uterine niche and poor wound healing following a cesarean section, laying the groundwork for future investigations into therapeutic approaches for the uterine niche.

overall_design: In this study, we utilized single-cell RNA sequencing to analyze  individual cells obtained from adjacent myometrium tissue of niche (Adjacent), well-healed cesarean scar tissue (Control), and niche tissue (Niche).

first: Xiaoya

last: Zhao

first: Qian

last: Zhu

first: Jian

last: Zhang


title: Mitochondrial protease Afg3l2 regulates the homeostasis of mouse hematopoietic stem and progenitor cells through Mmadhc [proteomics]

first: Bin

last: Guo

address: 950 W. Walnut Street

department: Microbiology & Immunology

summary: In order to study how mitochondrial proteases function on mouse hematopoietic stem cells, the article constructed Afg3l2 knockout mice. When Afg3l2 was knocked out, mouse hematopoietic stem cells showed HSC depletion accompanied by amino acid accumulation and TCA overactivation. We performed transcriptome and proteome sequencing analysis on cells from wild-type (WT) and Afg3l2-KO mice, and differential gene analysis revealed the Mmadhc protein. Further verification showed that Mmadhc is a substrate of the mitochondrial protease Afg3l2, and overexpression of Mmadhc can mimic the amino acid accumulation and TCA overactivation caused by Afg3l2 knockout. Overall, the mitochondrial protease Afg3l2 affects the homeostasis of hematopoietic stem cells by regulating the level of Mmadhc.

overall_design: Comparative gene expression profiles were performed using proteomics data from WT and Afg3l2 KO LK(Lin−c-Kit+) cells.

first: Meng

first: Xiashiyao

first: Jun

last: Wan


title: Mitochondrial protease Afg3l2 regulates the homeostasis of mouse hematopoietic stem and progenitor cells through Mmadhc [RNA-Seq]

overall_design: Comparative gene expression profiling was performed using RNA-seq data from WT and Afg3l2 KO LSK(Lin−Sca-1+c-Kit+) cells.


title: Ancient co-option of LTR retrotransposons as yeast centromeres

city: Dortmund

first: Max

last: Haase

address: Otto-Hahn-Str. 11

institute: Max Planck Institute of Molecular Physiology

summary:
Centromeres ensure accurate chromosome segregation, yet their DNA evolves rapidly across eukaryotes, leaving the origin of new centromere architectures unresolved. In the brewer’s yeast Saccharomyces cerevisiae (order Saccharomycetales), compact, genetically defined “point” centromeres replaced large, repeat-rich, epigenetically specified centromeres, but how this transition occurred has been unclear. Competing models have proposed either descent with modification from ancestral epigenetic centromeres or acquisition from selfish plasmid DNA.

Here we map and characterize centromeres in the sister order Saccharomycodales and identify evolutionarily related “proto-point” centromeres that bridge repeat-rich and point centromeres. Proto-point centromeres contain a single centromeric nucleosome positioned over an AT-rich core, but retain relaxed organization and sequence variability in flanking cis-elements. In two species, including Saccharomycodes ludwigii, proto-point centromeres are embedded within clusters of Ty5 long terminal repeat (LTR) retrotransposons, and their core CDEII and flanking motifs share sequence similarity to Ty5 LTR sequence.

Comparative genomics, synteny, and phylogenetic analyses across multiple yeast orders show that Ty5-cluster centromeres are ancient genomic features and support a model in which proto-point and point centromeres evolved by co-option of Ty5 LTR sequences in an ancestor with retrotransposon-rich centromeres, rather than by horizontal transfer from the 2-µm plasmid. These results indicate that yeast point centromeres are direct descendants of retrotransposons and illustrate how transposable elements can be repurposed to create genetically encoded centromeres.

overall_design: Cse4 was mapped by MNase-ChIP-sequencing of a mNeonGreen-tagged Cse4 variant.

first: Luciana

last: Lazar-Stefanita

first: Lyam

last: Baudry

first: Aleksandra

last: Wudzinska

first: Xiaofan

last: Zhou

first: Antonis

last: Rokas

first: Chris

last: Hittinger

first: Andrea

last: Musacchio

first: Jef

last: Boeke


title: Circular ZBTB44-encoded ZBTB44-342aa alleviates osteogenic differentiation of human aortic valve interstitial cells by inhibiting the cGAS-STING pathway

city: Guangzhou

first: Qingchun

last: Zeng

state: Guangdong Province

address: 1838 Northern Guangzhou Ave

department: State Key Laboratory of Organ Failure Research, Department of Cardiology

institute: Nanfang Hospital, Southern Medical University

type: Non-coding RNA profiling by high throughput sequencing

summary:
Background
Circular RNAs (circRNAs) are implicated in various physiopathologic activities and play a crucial role in calcific aortic valve disease (CAVD) progression. However, the role of coding circRNAs in CAVD remains unclear. In this study, we aimed to characterize coding circRNAs and explore their functions in CAVD.
Methods
We identified circular Zinc finger and BTB domain-containing protein 44 (CircZBTB44) through transcriptome sequencing and characterized its coding capacity through a series of experiments. We investigated the biological role of circZBTB44 and its encoded peptide, ZBTB44-342aa, in CAVD, as well as its mechanism of action through in vivo and in vitro experiments.
Results
We found that circZBTB44 promotes the translation of ZBTB44-342aa through N6-methyladenosine modifications. Functionally, ZBTB44-342aa alleviates mitochondrial DNA (mtDNA) release into the cytoplasm, suppresses the activation of the Cyclic GMP-AMP synthase-stimulator of interferon genes (cGAS-STING) pathway, and alleviates the osteogenic differentiation of human aortic valve interstitial cells (HAVICs). In vivo, circZBTB44 adeno-associated virus and STING deprivation alleviated the aortic valve lesions induced in an aortic valve injury (AVI) model. In vitro, the overexpression of circZBTB44 or the exogenous addition of the ZBTB44-342aa recombinant protein inhibited the osteogenic response of HAVICs. Conversely, the siRNA-mediated knockdown of circZBTB44 enhanced this response. Furthermore, H-151-induced inhibition of STING alleviated the osteogenic response of HAVICs, whereas STING activation by dimeric amidobenzimidazole exacerbated this response. ZBTB44-342aa inhibits mitochondrial fission by binding to Insulin-like growth factor 2 mRNA-binding protein 3, which reduces the leakage of mtDNA into the cytoplasm and promotes the stability of calcium/calmodulin-dependent protein kinase 1. This reduction inhibited STING activation and suppressed downstream osteogenic responses.
Conclusion
This study demonstrates that circZBTB44 encodes a novel peptide, ZBTB44-342, which alleviates CAVD progression by inhibiting the activation of the cGAS-STING signaling pathway. Therefore, circZBTB44 and STING may be potential targets for future therapeutic interventions in CAVD.

overall_design: Aortic valve tissues were collected from six patients with calcific aortic valve disease and five patients with non-calcific aortic valves. These were then subjected to high-throughput sequencing in order to screen for differentially expressed genes.

first: Dongtu

last: Hu

first: Yingwen

last: Lin

first: Gaopen

last: xian

first: Hailun

last: Huang

first: Songlin

last: Du

first: Tianyu

last: Xu


title: Free-floating Long-term Vascularized Mesenchymal Organoids [scRNA-seq]

first: Alejandro

last: de Janon

summary: The vasculature is essential for tissue function and pathology. Spheroid co-cultures of endothelial and mesenchymal stem/stromal cells (MSCs) form consistent structures, but the vascular components are short-lived. iPSC-derived  vascular organoids can establish complex vasculature but often have cell heterogeneous variable maturation and low reproducibility. We presents consistently-formed, free-floating, long-term Vascularized Mesenchymal Organoids (VMOs), formed by co-culturing human umbilical vein endothelial cells (HUVECs) and MSCs in a pre-gelled minimal Matrigel scaffold.  We studied, MSC heterogeneity, vascular biology and system dynamics.

overall_design: Samples are cells extracted and pooled from digested organoids cultured for 3 days and 14 days respectively

first: Shuichi

last: Takayama


title: A role for the cholinergic neuron circadian clock in RNA metabolism and mediating neurodegeneration [timecourse]

first: Sharon

last: Tam

overall_design: Timecourse RNA-seq profiling of wild-type mouse spinal cord cholinergic neurons collected every 4 hours across the 24-hr day

middle: B

first: Nathan

last: Waldeck

first: Matthew

last: Wright

first: Erin

last: Baker

first: Evangelos

last: Kiskinis

first: Joseph

last: Bass

first: Robert

last: Kalb

first: Jelena

last: Mojsilovic-Petrovic


title: The hegemonic EWSR1::ETS oncogene overrules core regulatory circuitry mechanisms in Ewing sarcoma [ChIP-seq]

city: Paris

first: Olivier

last: Delattre

address: 26 rue d'ulm

country: France

institute: Institut Curie

summary: Ewing sarcoma is an aggressive bone tumor of adolescence characterized by a hallmark EWSR1::FLI1 fusion oncogene. This chimeric protein drives tumorigenesis by reshaping transcriptional and epigenetic landscapes. However, how it is transcriptionally regulated and whether additional master transcription factors (MTFs) form a core regulatory circuit (CRC) in Ewing sarcoma remain unclear. Here, using an extensive panel of Ewing sarcoma cell lines and primary tumors, we mapped super-enhancers and identified strong enrichment of GGAA microsatellites, confirming their specificity to Ewing sarcoma as compared to other pediatric cancers and normal tissues. Integrating transcriptomic, epigenetic, 3D chromatin conformation, and dependency data, we predicted a set of MTFs potentially comprising a CRC. However, functional validation demonstrated that these MTFs neither establish auto-regulatory loops nor confer proliferative dependencies typical of CRC s in other pediatric tumors. Instead, EWSR1::FLI1 emerged as an “hegemonic” oncogene, regulating the expression of these MTFs without reciprocal regulation. Knockdown (KD) of EWSR1::FLI1 strongly reduced Ewing sarcoma cell proliferation and shifted H3K27ac profiles toward mesenchymal states, whereas silencing of individual or combined MTFs did not alter cell growth or EWSR1::FLI1 expression. These findings highlight the absence of a classical CRC in Ewing sarcoma and emphasize EWSR1::FLI1 as the dominant oncogene but also as a major vulnerability in this disease.

overall_design: ChIP-seq (H3K27ac and FLI1) were performed to complete the actual catalog of Ewing sarcoma cell line. We also generate H3K27ac profile after inhibiting the oncogene EWSR1::ETS and in some MSC models. Some transcription factors defined as CRC candidates were also sequenced (IKZF2, POU3F2, SOX6)

first: Sandrine

last: Grossetete

first: Sakina

last: Zaidi

first: Karine

last: Laud-Duval

first: Didier

last: Surdez


title: The hegemonic EWSR1::ETS oncogene overrules core regulatory circuitry mechanisms in Ewing sarcoma [RNA-seq]

overall_design: Expression data (RNA-seq) in Ewing sarcoma cell line expressing or not the oncogene EWSR1::ETS


title: RNAseq analysis of human neutrophils from healthy controls and from people with systemic lupus eruthematosus

city: Liverpool

first: Helen

middle: Louise

address: 6 West Derby Street

department: Institute of Life Course and Medical Sciences

country: United Kingdom

institute: University of Liverpool

summary: We report gene expression profiles of blood neutrophils from (1) healthy adults (2) healthy pediatrics, (3) adults with SLE, (4) pediatrics with SLE. Participants were recruited from two sites, Liverpool UK or Bangkok, Thailand.

overall_design: Neutrophils (purity >97%) were isolated from peripheral blood by separation of leukocytes from erythrocytes using HetaSep, then centrifugation through density-gradient reagent (HetaSep). Remaining erythrocytes were removed by hypotonic lysis using ammonium chloride buffer. RNA was isolated using Trizol:chloroform, cleaned up by Qiagen RNeasy kit including on-column DNase digest and sequenced on the DNBseq platform.

first: Grace

last: Filbertine

first: Marie

last: Phelan

first: Direkrit

last: Chiewchengchol

first: Nattiya

last: Hirankarn


title: Derepression of transposable elements in mouse prefrontal cortex disrupts social behavior

city: Richmond

first: Corinne

last: Smith

state: Virginia

address: 1217 E. Marshall St.

department: Anatomy and Neurobiology

institute: Virginia Commonwealth University

summary: The neurobiological origins of social behaviors are poorly understood. Previous studies have linked the function of a single Krüppel-associated box zinc finger protein (KZFP), ZFP189, in the mouse prefrontal cortex (PFC), with the regulation of transposable elements (TEs), immune genes, and social behaviors. Here, we expand the scope of inquiry to explore the relationship between collective PFC KZFP function and social behaviors by altering the function of the cognate KZFP interacting protein TRIM28 within the PFC of male and female mice. We reprogrammed natural TRIM28WT by replacing the endogenous, transcriptionally repressive domain with a synthetic, enhanced transcriptional activation domain VP64-p65-Rta (TRIM28VPR), or by excising the transcriptional regulatory domain (TRIM28NFD). Upon intra-PFC viral-mediated delivery of TRIM28 variants, we observed that inversion of TRIM28 transcriptional control via HSV-TRIM28VPR selectively produced deficits in social behaviors, without affecting non-social behaviors. RNA-sequencing of manipulated PFC revealed that HSV-TRIM28VPR drove transcriptional escape of all classes of TEs, particularly those located within intronic and enhancer regions proximal to downregulated immune genes. HSV-TRIM28VPR-mediated social deficits were reversible by intra-PFC repletion of interferon cytokines. These data point to PFC KZFP-TRIM28 interactions as necessary to stabilize genomic TEs to enable cis-regulation of key immune gene expression, which enhances organismal capacity for complex, pro-social behaviors.

overall_design: RNA-seq profiling of mouse prefrontal cortex tissue 4 days post-viral-introduction of TRIM28WT, TRIM28NFD, TRIM28VPR, or GFP transgenes.

first: R

last: Kim

first: Natalie

last: Truby

first: Shelbey

last: Strandberg

first: Jessica

last: Bell

first: Nic

last: Carwile

first: Gabriella

last: Silva

first: Rachael

last: Neve

first: Theingi

last: Aung

first: Xiaohong

last: Cui

first: Peter

last: Hamilton


title: The hegemonic EWSR1::ETS oncogene overrules core regulatory circuitry mechanisms in Ewing sarcoma [ChIP-seq 2]

first: Olivier

last: Delattre

overall_design:
ChIP-seq (H3K27ac and FLI1) were performed to complete the actual catalog of Ewing sarcoma cell line. We also generate H3K27ac profile after inhibiting the oncogene EWSR1::ETS and in some MSC models. Some transcription factors defined as CRC candidates were also sequenced (IKZF2, POU3F2, SOX6)

***************************************************************
Raw files for human/patient samples were not submitted to GEO due to concerns about submitting personally identifiable sequence data for open access.
***************************************************************

first: Sandrine

last: Grossetete

first: Sakina

last: Zaidi

first: Karine

last: Laud-Duval

first: Didier


title: Mismatch repair deficiency drives malignant progression and alters the tumor immune microenvironment of glioblastoma

city: NEW YORK

first: Alexander

last: Tsankov

state: New York

address: 1425 Madison Ave

department: Genetics and Genomic Sciences

summary: Mutations in DNA mismatch repair (MMR) pathway genes (MSH2, MSH6, MLH1, and PMS2) have been associated with acquired resistance to temozolomide and with a high tumor mutation burden (TMB) in high-grade gliomas (HGG), including glioblastoma (GBM). However, the specific roles of individual MMR genes in the initiation, progression, increased TMB and microsatellite instability (MSI), and resistance to temozolomide (TMZ) in glioma have not been extensively studied. In the present study, we developed de novo mouse models of germline and somatic MMR-deficient (MMRd) HGG. Surprisingly, we found that loss of Msh2 or Msh6 does not lead to a high TMB and MSI nor confer response to anti-PD-1 in GBM. Our data also revealed that germline MMRd leads to the transition from low to high-grade glioma. Additionally, we show that mice with germline MMRd had reduced survival compared to MMR-proficient (MMRp) tumor-bearing mice. This effect was tumor cell intrinsic but was associated with MMRd in the tumor immune microenvironment, which led to increased immunosuppressive pathways in myeloid cells, reduced lymphoid infiltration, and increased exhaustion of CD8+ T cells. We have found that both MMR-reduced (MMRr) and MMRd GBM are resistant to TMZ, unlike MMRp tumors. Our study shows that KL-50, a new imidazotetrazine-based DNA targeting agent designed to induce DNA interstrand cross-links, is effective in treating both germline and somatic MMRr and MMRd GBM-bearing mice. These findings present a new treatment option for HGG patients with inherited or acquired MMRd who are resistant to TMZ.

overall_design: Using immunocompetent de novo mouse models of GBM, we generated PDGFB+shp53-driven tumors in the right-frontal striatum of Msh2 WT, HET and KO mice. At human endpoint of these mice, tumor cells were isolated and sc-RNA seq was conducted to compare the difference between genotypes.

first: Dolores

last: Hambardzumyan

first: Puigdelloses Vallcorba

last: Montserrat

first: Nishant

last: Soni


title: Profling of breast cancer cells using single-cell triomics analysis of DNA, RNA, and proteins with Tapestri chemistry

city: Göteborg

first: Emma

last: Jonasson

address: Medicinaregatan 1F, floor 5

department: Sahlgrenska Center for Cancer Research

country: Sweden

institute: University of Gothenburg

summary: Cancer displays large heterogeneity both between and within tumors where intratumor heterogeneity can occur at genetical, transcriptional and translational level. The variability results in tumor cell subpopulations with distinct molecular and cellular characteristics, some of which can have therapy-resistant properties. To date it has been experimentally challenging to study tumor cell heterogeneity. Here, we present a single-cell triomics approach to assess DNA, RNA and protein simultaneously in thousands of single cells using microfluidics. First, we profiled two breast cancer cell lines, MCF7 and T-47D, cultured in monolayer with and without 5-fluorouracil treatment. Next, we generated cell-free patient-derived scaffolds (PDS) that we repopulated with MCF7 cells for 21 days. This experimental model system mimics in vivo tumor cell growth and is suitable for drug testing, such as chemotherapies. We profiled individual MCF7 cells cultured in different PDSs with and without 5-fluorouracil treatment using triomics. Our approach and data demonstrate that single-cell triomics is feasible, which opens up new means to decipher tumor heterogeneity at different layers.

overall_design: Single-cell triomics using Tapestri chemistry (MissionBio) assessing DNA, RNA and protein in MCF7 and T-47D breast cancer cells grown in monolayer culture and MCF7 cells grown in patient-derived scaffold, with or without 5-fluorouracil.

first: Stefan

last: Filges

first: Anna

last: Gustafsson

first: Anders

last: Ståhlberg


title: HIF-1α+ CD4 T cells coordinate a tissue resident immune cell network in the lung [Xenium]

city: Basel

first: Carolyn

last: King

address: Hebelstrasse 28

department: Department of Biomedicine

institute: University of Basel

summary: Pulmonary infection leads to the development of CD4+ T resident helper (TRH) cells in the lung. TRH cells generated after influenza infection support local humoral responses and exhibit differentiation plasticity following infectious challenge. A subset of TRH cells is enriched for expression of the transcription factor HIF-1α which has unknown function in the lung. Here we find that inducible deletion of HIF-1α in CD4 T cells leads to decreased numbers of CXCR6+ tissue resident T cells with minimal impact on peripheral lymphoid responses. At the same time, HIF-1α deletion impairs the replenishment of tissue resident macrophages and NK cells, as well as influenza specific IgA titers. These seemingly disparate responses converge upon a requirement for IL-21 produced by HIF-1α+ CD4 T cells. A similar HIF-1α dependent network is engaged in a lung adenocarcinoma model, highlighting novel roles for HIF-1α+ CD4 T cells in coordinating protective immunity during infection and cancer.

overall_design: WT mouse infected with influenza strain PR8. Harvest at day 14 and day 30 post infection


title: KRAS-Dependent Metabolic Reprogramming of Endothelial Cells as a Therapeutic Vulnerability in Brain Arteriovenous Malformations

city: Toronto

first: Ruilin

last: Wu

state: Ontario

address: 101 College Street

country: Canada

institute: University of Toronto

summary: Brain arteriovenous malformation (bAVM) is a disease characterized by the formation of tangled blood vessels with direct connections between arteries and veins. There is currently limited understanding of the underlying molecular mechanisms that are responsible for the development of bAVM. Our previous work identified the presence of somatic activating KRAS mutations in endothelial cells (ECs) isolated from clinical bAVM tissue. Given the association between mutant KRAS, metabolic reprogramming and EC angiogenesis, we aimed to investigate if altered endothelial metabolism is involved in the pathogenesis of bAVM to drive abnormal vascular remodeling. As such, we engineered a human EC model with inducible expression of mutant KRAS G12V. We observed a significant increase in glucose metabolism upon mutant KRAS expression. This elevation in EC glycolysis contributes to the angiogenic behaviours that are observed in our mutant KRAS EC models. To further explore the transcriptional alterations underlying these changes, we performed this RNA-sequencing study with various glycolysis inhibitors and a MEK inhibitor (the major signaling pathway altered in mutant KRAS ECs) to profile the impact of glucose metabolism changes on endothelial biology.

overall_design: Bulk mRNA sequencing of telomerase-immortalized human umbilical vein endothelial cells (Telo-HUVECs) with stable expression of KRAS G12V proteins. The mutant KRAS cells were treated with various glycolysis inhibitors and a MEK inhibitor to examine the effects of glycolytic and MAPK/ERK pathway suppression on the transcriptional landscape of mutant KRAS endothelial cells.

first: Negar

last: Khosraviani

first: Ann

last: Mansur

first: Emilie

last: Boudreau

first: Dakota

last: Gustafson

first: Sneha

last: Raju

first: Crizza

last: Ching

first: Amira

last: Klip

first: Thomas

last: Wälchli

first: Kathryn

last: Howe

first: Ivan

last: Radovanovic

first: Joshua

last: Wythe

first: Jason

last: Fish


title: Chromatin accessibility profiling of HSPC-derived erythroblast differentiation [ATAC-seq]

city: Vienna

first: Florian

last: Halbritter

address: Zimmermannplatz 10

country: Austria

institute: St. Anna Children's Cancer Research Institute (CCRI)

summary: A prime goal of regenerative medicine is to replace dysfunctional cells in the body. To design protocols for producing target cells in the laboratory, one may need to consider exponentially large combinations of culture components. Here, we tested the potential of iteratively approximating the target phenotype by quantifying the distance between chromatin profiles (ATAC-seq) of differentiating cells in vitro and their in-vivo counterparts. We tested this approach on the well-studied generation of erythroblasts from haematopoietic stem cells, evaluating a fixed number of components over two sequential differentiation rounds (8x8 protocols). We found that the most erythroblast-like cells upon the first round yielded the most erythroblast-like cells at the second round, suggesting that greedy selection by chromatin approximation can be a viable optimisation strategy. Furthermore, by targeting transcriptional regulators linked to chromatin regions that were incompletely reprogrammed even after two rounds of differentiation, we could make a data-driven selection of additions to the protocol that further improved erythropoiesis. In future, our methodology can help craft notoriously difficult cells in vitro, such as B cells.

overall_design: 165 ATAC-seq datasets from two round of differentiation with different treatment combinations: 16 from pilot experiment (8 combinations round 1 only) + 135 full experiment (8 combinations round 1; 8x8 combinations round 2) + 15 validation experiment with adapted treatment protocols (+TGFB or IL4);  2-3 replicates per condition, low-quality samples removed

first: Luis

last: Montano-Gutierrez

first: Sophie

last: Müller

first: Ana

middle: P

last: Kutschat

first: Igor

last: Adameyko

first: Davide

last: Seruggia


title: Opposing functions of distinct regulatory T cell subsets in colorectal cancer

city: New York

last: Mitra

address: 1275 York Avenue

institute: Memorial Sloan Kettering Cancer Center

summary: Regulatory T (Treg) cells are considered major contributors to the growth of solid organ tumors, with the notable exception of colorectal cancer (CRC), despite the documented abundance of Treg cells in CRC. Here, we demonstrate that IL-10⁺ and IL-10⁻ Treg cells constitute two distinct subsets with opposing functions: IL-10⁺ Treg cells counteract, while IL-10⁻ Treg cells promote, the growth of genetically engineered CRC tumors. Our findings suggest that the tumor-suppressive function of IL-10⁺ Treg cells is mediated by their suppression of effector CD4⁺ T cell production of IL-17, a cytokine that directly stimulates CRC tumor cell proliferation. Consistently, IL-10⁺ Treg cells were more abundant in both mouse and human CRC tumors than in tumor-adjacent normal colon tissue, whereas IL-10⁻ Treg cells exhibited the opposite distribution. Furthermore, gene expression signatures associated with IL-10⁺ and IL-10⁻ Treg cells correlated with better and worse disease prognoses in human CRC, respectively. This functional dichotomy between Treg subsets provides a rationale for therapeutic strategies aimed at selectively targeting pro-tumoral Treg cells while preserving their anti-tumoral counterparts across various barrier tissue cancers that harbor both subsets.

overall_design: Mouse T cells were sort purified from transcplanted AKP tumor, adjacent cecal tissue, and the draining C1 lymph node two, four, or six week after tumor transplantation.

first: Xiao

last: Huang

first: Dan

last: Feng

first: Emma

middle: S

last: Andretta

first: Nima

last: Hooshdaran

first: Aazam

last: Ghelani

first: Eric

middle: Y

last: Wang

first: Joe

middle: N

last: Frost

first: Victoria

last: Lawless

first: Aparna

last: Vancheswaran

first: Qingwen

last: Jiang

first: Christina

last: Leslie

first: Alexander

last: Rudensky


first: Sneha

overall_design: RNA-seq profiling of tumors from locally advanced rectal cancer (stage II-III) patients.


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

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Refer to individual Series

middle: R

first: Cheryl

last: Mai

first: Karuna

last: Ganesh


overall_design: Mouse T cells and myelod cells were sort purified from transcplanted AKP tumor and the adjacent cecal tissue. Tissues were harvested from IL-10cre Foxp3 LSL-DTR mice treated with DT to deplete IL-10+ Treg cells, or bDT as control two weeks after tumor inoculation and DT/bDT treatment. DT/bDT treatments were done on Day 0, 1, 4, 7, 10, and 13.


overall_design: Single-cell multi-ome data (ATAC-seq and nuclear RNA-seq) of T cells sorted from mouse AKP tumor and normal adjacent tissue samples 4 weeks after transplantation.


title: TBP regulates transposable element expression in early mouse embryos

city: Munich

first: Clara

last: Hermant

address: Feodor-Lynen-Straße 21

department: Institute of Epigenetics and Stem Cells (IES)

institute: Helmholtz Zentrum Munich

summary: Activation of the embryonic genome is a crucial step in development. In addition to thousands of genes, many transposable elements (TEs) are robustly transcribed during early mammalian development. However, their transcriptional regulators remain largely unexplored. Here, we set out to identify transcription factors regulating the expression of highly expressed TEs from the LINE, SINE and ERVL families during mouse preimplantation development. In particular, the MaLR family are the most abundant ERVL in the mouse genome and are also the most abundant constituent of the transcriptome in early mouse embryos. Surprisingly, we find the general transcription factor TBP directly and specifically activates MaLRs. Loss-of-function of TBP in mouse embryos leads to downregulation of MaLRs, specifically the ORR1A family, which is the youngest ORR subclass and contributes a significant portion of major zygotic genome activation transcripts. We demonstrate that TBP regulates MaLRs through direct binding. Our work contributes to the identification of regulators of TE expression in vivo and highlights a previously unrecognised role for a general transcription factor in regulating an exquisitely specific TE transcriptional programme.

overall_design: Late 2-cell stage embryos were collected and Cut&Tag was performed using TBP antibody and IGG as control.

first: Maria-Elena

last: Torres-Padilla


last: Mitra

overall_design: Single-cell multi-ome data (ATAC-seq and nuclear RNA-seq) of T cells sorted from human colorectal cancer tumor and normal adjacent tissue samples. T cells are sorted as live MHC-II- CD3+TCRb+.

first: Xiao


title: TBP regulates transposable element expression in early mouse embryos [RNA-seq]

overall_design: Mouse embryos (zygotes) were microinjected with antibodies against TBP (TBP LOF) and mIGG (CONTROL for TBP LOF) or FOXJ3 (FOXJ3 LOF) and rIGG (CONTROL for FOXJ3 LOF), cultured for 4h and microinjected with Trim21 mRNA, to finally be cultured until the late 2-cell stage and collected for Smart-Seq+5', an adaptation protocol of Smart-Seq2


title: A critical role of GABAergic neurons in driving CGG repeat toxicity associated with Fragile X Premutation Carriers [scRNA-Seq]

first: Yulin

last: Jin

address: 615 Micheal Street

department: Human Genetics

institute: Emory University

summary: Fragile X-associated tremor/ataxia syndrome (FXTAS) is a neurodegenerative disorder caused by a CGG repeat expansion (55-200) in the 5′ UTR of the fragile X messenger ribonucleoprotein 1 (FMR1) gene. The molecular mechanisms of FXTAS involve the toxic effects of the expanded CGG repeat RNA and the repeat-associated non-AUG (RAN) translation product, the FMRpolyG peptide. To gain deeper insights into the pathogenesis of FXTAS, we performed comprehensive transcriptomic profiling of the brain of an FXTAS mouse model at both bulk and single-cell levels, which revealed pronounced dysregulation in inhibitory neurons of FXTAS animals. Further, conditional FXTAS mouse models demonstrated that the expression of expanded CGG repeats in GABAergic neurons alone was sufficient to induce key features associated with the expression of FMR1 premutation CGG repeats. Many of the dysregulated mRNAs in GABAergic neurons were found to be bound by hnRNPA2/B1, an RNA-binding protein sequestered by CGG repeat RNA.

overall_design: To gain deeper insights into the cellular perturbations associated with the pathogenesis of FXTAS, we applied snRNA-seq to the prefrontal cortex of FXTAS animals and littermate controls at the onset of the phenotype stage (3-4m).

first: Yiqu

last: Cao

first: Wenjing

last: Ma

first: Peng


title: A critical role of GABAergic neurons in driving CGG repeat toxicity associated with Fragile X Premutation Carriers

overall_design: To investigate the molecular signatures associated with the expression of expanded CGG repeats, we performed RNA-Seq on brain tissues from the FXTAS mouse model, which expresses both FMRpolyG and CGG repeat RNA. Samples were collected from the cortex, cerebellum, and hippocampus of FXTAS mice and control animals at two key stages: the onset of phenotypic presentation (3–4 months) and the full phenotype stage (6–7 months). Additionally, to gain deeper insights into the molecular signatures of GABAergic neurons during disease progression, we performed anti-HA Targeted TRAP (Translating Ribosome Affinity Purification), followed by RNA sequencing (TRAP-seq). This analysis was conducted on the cortex and cerebellum of FMRpolyG+RNA(+/-);Gad2-Tagger mice and their littermate controls.


title: Endoplasmic reticulum proteostasis in senescent cells is a vulnerability to senolytic vanadyl sulfate

first: Jiacheng

last: Li

address: 900 E 57th Street,

department: Medicine

institute: University of Chicago

summary: Lifestyles including dietary choices have a major impact on shaping of human physiology and health, including aging. However, the underlying mechanisms remain largely unknown. Here, we use a blood nutrient compound library-based screening approach to demonstrate that vanadyl sulfate (VS), a mineral dietary supplement, selectively induces cell death in senescent cells, thus extends lifespan and ameliorates phenotypes of aging and age-related disorders in vivo. Integrated functional omics strategies including activity-based protein profiling reveal that senescent cells accumulate glutathione disulfide, leading to overall reduced cysteine reactivity of the proteome but elevated cysteine reactivity of proteins that are crucial for endoplasmic reticulum (ER) proteostasis, which licenses VS to effectuate senolysis. VS inactivates reactive cysteines on proteins to form incorrect disulfide bonds, causing aberrant protein misfolding and aggregation that sabotage ER proteostasis, leading to senescent cell death. Our findings uncover a unique vulnerability of senescent cells for therapeutic exploration to treat aging and age-related disorders.

overall_design: RNA seq profiling of BJ cells treated with GSSG liposome or Control liposome for 48 hour. (n=3)

first: Rong

last: Wu

first: Fan

last: Yang

first: Ruitu

last: Lyu

first: Chuan

last: He

first: Jing

last: Chen


title: Feed-forward regulatory loops by NR5A2 ensure robust gene activation during pre-implantation development

city: Martinsried

first: Siwat

last: Ruangroengkulrith

state: Bavaria

address: Am Klopferspitz 18

department: Totipotency

institute: Max Planck Institute of Biochemistry


title: KDM3A catalysed formation of hydroxyacetyl-lysine on histone H3K9

city: Oxford

first: David

middle: Robert

last: Mole

address: Roosevelt Drive, Headington

department: Nuffield Department of Medicine

institute: University of Oxford

summary: Histone modifications, including Ne-lysine acetylation and methylation, play critical roles in eukaryotic transcription. The addition of acetyl- and methyl-groups and removal of acetyl-groups involves redox neutral reactions, whereas demethylation is O2 dependent, a reaction with potential to enable O2 availability mediated regulation, as occurs for reactions catalysed by the 2-oxoglutarate dependent hypoxia-inducible factor (HIF) hydroxylases. A screen for substrates of the HIF-regulated Jumonji-C (JmjC) lysine demethylase KDM3A led to the finding that purified recombinant KDM3A catalyses oxidation of the Ne-acetyl group of Lys-9 of histone H3 giving an Ne-hydroxyacetyl-lysine residue. Studies employing a Ne-hydroxyacetyl-lysine selective antibody and mass spectrometry support the cellular relevance of Ne-hydroxyacetyl-lysine. The combined biochemical and cellular results reveal evidence for an unanticipated O2-mediated link between histone lysine Ne-acetylation and JmjC catalysis. Future work can explore the biological significance of Ne-hydroxyacetylation, including in the hypoxic response where KDM3A plays a role in regulating HIF target genes.

overall_design: ChIP-seq analysis of histone modifications in HEK293T cells normalised to a Drosphila chromatin spike in, immunopreciitated in same reaction using a secondary antibody

first: Jessica

last: Kindrick


title: PUF60-Mediated Splicing Is a Key Driver of Triple Negative Breast Cancer

city: La Jolla

first: Brian

last: Yee

state: CA

address: 9500 Gilman Dr

department: Health

institute: UCSD


title: Somatosensory nerves induce the pathologic response to physis injury

city: Baltimore

first: Aaron

last: James

state: Maryland

address: 720 Rutland Avenue, Ross Bldg., Room 529

department: Pathology

institute: Johns Hopkins University

summary: Somatosensory neurons are crucial for detecting a diverse repertoire of stimuli, including in skeletal tissues in which sensations of pain, pressure, and position are transmitted. Trauma to the physis often lead to pathologic "bony bar" formation, impeding normal growth of the skeleton. The role of somatosensory nerves in detecting and in turn regulating such an event is entirely unknown.  Here, we observed prominent sensory nerve growth within the injured physis in mouse and human tissues corresponding to domains of NGF expression.  Two independent methods to inhibit TrkA (Tropomyosin Receptor Kinase A) signaling significantly reduced nerve sprouting and attenuated pathologic bony bar formation, while converse experiments using neural conditioned medium treated organ cultures showed the opposing effects.  Somatosensory nerve retrograde tracing and single cell RNA sequencing (scRNA-Seq) identified the precise neural transcriptome evoked by physis injury.  Corresponding scRNA-Seq of injured growth plates implicated PTN neuroregulatory signaling in growth plate pathology.  Finally, FDA-approved long acting bupivicaine may be used to prevent pathological innervation and ossification of the growth plate after injury.  These findings suggest that targeting somatosensory nerves could offer new strategies to treat pain and optimize patient outcomes after growth plate injury.

overall_design: The samples comprised two distinct tissue types: Dorsal Root Ganglia (DRGs) and growth plates. The experimental design included two groups: a control group and a treatment group. The control group received intraperitoneal injections of the vehicle (DMSO). The treatment group, designated as the 1NMPP1 group, received intraperitoneal injections of 1NMPP1 (5 mM solution, 17 μg/g body weight) at two time points: 48 hours and 2 hours prior to surgery. Following the procedure, animals in the 1NMPP1 group were maintained on drinking water containing 40 μM 1NMPP1 dissolved in ddH2O with 1% PBS-Tween 20.

first: Zhao

first: Xin

last: Xing


title: 5' extended gRNA increases the cleavage activity of SuperFi-Cas9

city: Hangzhou

first: Lijia

state: Zhejiang

address: Shilongshan Road No.18, Cloud Town, Xihu District

department: School of Life Sciences

institute: Westlake university

summary: High-fidelity Cas enzyme is pivotal to the safety of CRISPR technology in translational research and clinics. However, increasing editing fidelity often comes at the cost of a significant decrease in nuclease editing efficiency, e.g., the rationale designed SuperFi-Cas9 based on the structural basis for mismatch surveillance of wildtype Cas9. By conducting high-throughput profiling on the editing efficiency of 91,603 gRNAs, we found that SuperFi-Cas9/gRNA showed a strong nucleotide preference at the 21st position of the PAM-distal region of the target. A deep-learning model trained from the profiling data further provided a mutation map, which implied that a gRNA-target duplex with twenty-one matched nucleotides positively contributed to the editing efficiency of SuperFi-Cas9/gRNA. By further measuring the cleavage activities at twenty endogenous genomic locations, we demonstrated that a gRNA with extended 5' nucleotides significantly increased the editing activity of SuperFi-Cas9 while remaining its high fidelity, which makes SuperFi-Cas9 a valuable nuclease of the CRISPR toolbox. Together, by deep-learning modeling on high throughput profiling data, we reported that gRNAs with extended 5' nucleotides can rescue the impaired cleavage activities of SuperFi-Cas9.

overall_design: We chose twenty targets to verify the editing efficiency of different sgRNAs at their endogenous sites. The sgRNAs were cloned into a V6-opt-test1 backbone vector. A tRNA was placed between the U6 promoter and gRNA, allowing the incorporation of the designed leading nucleotide at the 5' of the expressed gRNA.


title: PUF60-Mediated Splicing Is a Key Driver of Triple Negative Breast Cancer [RNA-seq]

summary: RNA-binding proteins (RBPs) regulate post-transcriptional gene expression, influencing key cancer pathways. Understanding how RBPs control these processes can uncover new treatment strategies, which is crucial for cancers that lack effective targeted therapies such as triple negative breast cancer (TNBC). Here, we employ dual in vitro and in vivo CRISPR/Cas9 screens to investigate the role of RBPs in TNBC, revealing the poly(U)-binding splicing factor 60 (PUF60) as a key modulator of TNBC cell survival. Integrated eCLIP and RNA-sequencing analysis identify that PUF60 drives exon inclusion within transcripts associated with cell cycle and genomic stability, thereby promoting proliferation. Furthermore, disrupting PUF60 splice site activity via a substitution in its RNA-binding domain causes widespread exon skipping, leading to the downregulation of proliferation-associated mRNAs and inducing apoptosis in TNBC cells. We demonstrate that this RNA-binding disruption inhibits TNBC cell proliferation and shrinks tumor xenografts, highlighting the molecular mechanism through which PUF60 supports cancer progression. Our work demonstrates functional in vivo screening of RBPs as an effective strategy for identifying unexpected cancer regulators. Here, we reveal a crucial role for PUF60-mediated splicing activity in supporting oncogenic proliferation rates and highlight its potential as a therapeutic target in triple negative breast cancer.

overall_design: RNA-seq of MCF10A, MDA-MB-231, MDA-MB-436, and SUM-149 cells following PUF60 knockdown; RNA-seq of MDA-MB-231 cells expressing an L140P PUF60 mutant

first: Alexandra

middle: T

last: Tankka

first: Jaclyn

middle: M

last: Einstein

first: Catherine

last: Zhou

first: Vivian

last: Pham

first: Yuhan

last: Zhang

first: Jack

last: Naritomi

first: Grady

last: Nguyen

first: Orel

last: Mizrahi

first: Mark

last: Perelis

first: Joseph

last: Sarsam

first: Fred

last: Tan

first: Dan

last: Kaufman

first: Corina

last: Antal

first: Gene

middle: W

last: Yeo


title: PUF60-Mediated Splicing Is a Key Driver of Triple Negative Breast Cancer [eCLIP]

first: Brian

last: Yee

overall_design: eCLIP of PUF60 in MDA-MB-231, MDA-MB-436, SUM-149, and MCF10A cells and eCLIP of V5-tagged L140P mutant PUF60 in MDA-MB-231 cells, in duplicate


title: Role of CTGF-LRP1 in Impaired Healing of Cesarean Section Incisions

city: Shanghai

first: Chuqing

last: He

address: No. 910, Hengshan Rd

institute: School of Medicine, Shanghai Jiaotong University

summary: Uterine niche (niche) is a long-term complication following a cesarean section, which may increase the risk during subsequent pregnancies, and its pathogenesis remains incompletely understood. In this study, we utilized single-cell RNA sequencing to analyze 43,384 individual cells obtained from adjacent myometrium tissue of niche (Adjacent), well-healed cesarean scar tissue (Control), and niche tissue (Niche). Our analysis revealed that fibroblasts (FB) could be classified into three subgroups. FB3 in the Niche exhibited a reduced capacity for extracellular matrix synthesis. Cell communication analysis has identified that the secretion of CTGF by Endo3 and the expression of LRP1 by FB3 are involved in the receptor-ligand interaction of uterine fibroblasts during wound healing. Our research findings were supported by tissue staining and in vitro experiments, indicating that silencing LRP1 may prevent CTGF from promoting the synthesis of extracellular matrix by primary uterine fibroblasts. Furthermore, animal models have indicated that CTGF can promote the healing of uterine scars.These results provide insights into the cellular environment and mechanistic background underlying the formation of the uterine niche and poor wound healing following a cesarean section, laying the groundwork for future investigations into therapeutic approaches for the uterine niche.

overall_design: LRP1 siRNA (5'-GTTTCGAGGTGGTGATTCA-3') and a negative control siRNA (5'- TTCTCCGAACGTGTCACGT-3') were sourced from Tsingke Biological Technology in China. The siRNA reagent was prepared in a stock solution at a concentration of 20 uM. JetPRIME (Polyplus) was utilized as an efficient transfection reagent. The transfection complex, comprising 5.5 ul of siRNA stock solution, 200 ul of jetPRIME Buffer, and 4 ul of jetPRIME reagent, was combined, gently vortexed, and incubated for 10 minutes at room temperature. This transfection complex was then added to a 6-well plate containing 1.5×105 cells and gently mixed to achieve a final siRNA concentration of 50 nM in each well. Following transfection with LRP1 siRNA or negative-control siRNA for 48 hours, the cells were subsequently incubated with CTGF (HY-P70106, MCE) for 48 hours. RNA and protein were then extracted for subsequent analysis.

first: Xiaoya

last: Zhao

first: Qian

last: Zhu

first: Jian


title: A roadmap to human hippocampal neurogenesis in adulthood, aging and AD

city: Chicago

last: Maienschein-Cline

address: 900 S Ashland Ave, Rm 1058, M/C 937

department: Research Resources Center

institute: University of Illinois at Chicago

summary: In the rodent, hippocampal neurogenesis plays critical roles in learning and memory, is tightly regulated by inhibitory neurons and contributes to memory dysfunction in Alzheimer’s disease (AD) mouse models. In contrast, the mechanisms regulating neurogenesis in the adult human hippocampus, the dynamic shifts in the transcriptomic and epigenomic profiles in aging and AD and putative niche interactions within the cellular environment, remain largely unknown. Using single nuclei multi-omics of postmortem human hippocampi we map the molecular mechanisms of hippocampal neurogenesis across aging, cognitive decline, and AD neuropathology. Transcriptomic and epigenetic profiling of neural stem cells (NSCs), neuroblasts and immature neurons suggests that the earliest shift in the characteristics of neurogenesis takes place in NSCs in aging.  Cognitive impairment was associated with changes in neuroblast profile. In AD, there was a widespread cessation of the transcription machinery in immature neurons, with robust downregulation of genes regulating ribosomal and mitochondrial function. Further, there was substantial loss of parvalbumin+ inhibitory neurons in the hippocampus in aging. The number of the rest of inhibitory neurons were reduced as a function of age and diagnosis. Notably, a similar system-level effect was observed between immature and inhibitory neurons in the transition from aging to AD, manifested by common molecular pathways that were ultimately lost in AD. The numbers of neuroblasts, immature and GABAergic neurons inversely correlated with extent of neuropathology. Using CellChat and NeuronChat, we inferred the ligands and receptors by which neurogenic cells communicate with their cellular environment. Loss of synaptic adhesion molecules and neurotransmitters, either sent or received by neurogenic cells, was observed in AD. Together, this study delineates the molecular mechanisms and dynamics of human neurogenesis, functional association with inhibitory neurons and a mechanism of hippocampal hyperexcitability in AD.

overall_design: Nuclei isolated from the hippocampus of young adults (YA, n=2), aging with no cognitive impairments (NCI, n=4), mild cognitive impairments/early dementia (MCI, n=4) and Alzheimer’s disease dementia (AD, n=4) were sequenced for multi-ome single nuclei RNA sequencing (snRNAseq) and single nuclei Assay for Transposase-Accessible Chromatin (snATACseq) analysis. Machine learning label transfer algorithm scANVI was used to transfer labels from two scRNAseq datasets, a human developmental forebrain and an adult human hippocampal dataset to ensure unbiased cell annotation.  Single-cell trajectory analysis ( CytoTRACE, Pseudotime) were used t infer and confirm the different developmental paths through the cell populations. Differential gene and peak expression as well as Motif analysis were used to assess the alterations in transcriptomic profiles of diffenet neurogenic niches accross diagnosis. Lastly, CellChat and NeuroChat to infer putative interaction of the neurogenic niches with their cellular niche in the human brain and the ligand-receptor pathways that may play a role in a decline in cell-cell interaction as a function of pathology in immature neurons

first: Ahmed

last: Disouky

middle: A

last: Sanborn

first: Mostafa

last: Mostafa

first: K

last: Sabitha

first: Aimee

last: Schantz

first: Namhee

last: Kim

first: Szymon

last: Pawlowski

first: William

last: Honer

first: David

last: Bennett

first: Yi

first: C

middle: D

last: Keene

first: Jalees

last: Rehman

first: Orly

last: Lazarov


title: wrn-deficient zebrafish exhibit premature malnutrition

city: Sagamihara

first: Kota

last: Ujibe

address: 5-10-1, Fuchinobe, Chuo-ku

institute: Aoyama Gakuin University

summary: To establish Werner syndrome model, we established wrn knockout mutant zebrafish. We performed time-course individual RNA-Seq to elucidate cause of death of mutant.

overall_design: Wild-type and mutant zebrafish larvae were sacrificed at 13 time point between 4 and 35 dpf with 4-13 biological replicates.

first: Makoto

last: Kashima

first: Miku

last: Kataoka

first: Rintaro

last: Shimada

first: Masashige

last: Okamoto

first: Isao

last: Kobayashi

first: Seiji

last: Wada

first: Hiroki

last: Matsuda

first: Akira

last: Sakamoto

first: Hiromi

last: Hirata


title: Unravelling the molecular activation of the reparative cardiac fibroblasts after myocardial infarction

city: Thuwal

first: Laura

last: Sudupe

address: King Abdullah University of Science and Technology (KAUST)

country: Saudi Arabia

institute: KAUST

summary: Cardiac fibroblasts have a central role during the ventricular remodeling process associated to different types of cardiac injury. Recent studies have shown that fibroblasts do not respond homogeneously to heart damage, suggesting that the adult myocardium may contain specialized fibroblast subgroups with specific functions. Due to the limited set of bona fide fibroblast markers, a proper characterization of fibroblast population dynamics in response to cardiac damage is still missing. Using single-cell RNA-seq we have identified and characterized a fibroblast subpopulation that emerges in response to myocardial infarction (MI) in a murine model. These activated fibroblasts exhibit a clear pro-fibrotic signature, and localize to the wounded myocardium. Combining epigenomic profiling with functional assays, we unveil important candidate regulators mediating their response to cardiac damage.

overall_design: Time-course of study of cardiac fibroblasts after myocardial infarction. Samples include control, 3 and 5 days after myocardial infarction.

first: Silvia

middle: C

last: Hernandez

first: Marina

last: Ainciburu

first: Nuria

last: Planell

first: Amaia

last: Vilas-Zornoza

first: Maria

last: López-Moreno

first: Sarai

last: Sarvide

first: Luis

last: Díaz-Martínez

first: Jorge

last: Cobos-Figueroa

first: Patxi

last: San Martin-Uriz

first: Emma

last: Muiños-López

first: Gloria

last: Abizanda

first: Purificación

last: Ripalda-Cemboráin

first: Felipe

last: Prosper

first: Adrian

last: Ruiz-Villalba

last: Gomez-Cabrero


title: Bulk RNA-seq analysis reveals the crucial role of Collagen triplex helix repeat containing-1 (Cthrc1) cardiac fibroblasts for ventricular remodeling after myocardial infarction

city: Pamplona

first: Núria

address: Pio XII, 55

country: Spain

institute: CIMA - UNAV

summary: Cardiac fibroblasts have a central role during the ventricular remodeling process associated to different types of cardiac injury. Recent studies have shown that fibroblasts do not respond homogeneously to heart damage, suggesting that the adult myocardium may contain specialized fibroblast subgroups with specific functions. Due to the limited set of bona fide fibroblast markers, a proper characterization of fibroblast population dynamics in response to cardiac damage is still missing. Using single cell RNA-seq we have identified and characterized a fibroblast subpopulation that emerges in response to myocardial infarction (MI) in a murine model. These activated fibroblasts exhibit a clear pro-fibrotic signature, and localize to the wounded myocardium. Combining epigenomic profiling with functional assays, we unveil important candidate regulators mediating their response to cardiac damage.

overall_design: Time-course of study of cardiac fibroblasts after myocardial infarction. Samples include 1, 2, 3, 4, 5 and 6 days after myocardial infarction.

first: Juan

last: Romero

first: Elena

last: Iglesias

first: Diego

last: Alignani

first: David