title: A genome-wide screen reveals Ints7 deficiency activates endogenous retrovirus MERVL, accompanied by anastasis of embryonic stem cells [E14TG2a]

city: changsha

first: shen

last: yunfan

state: hunan

address: xiangya road

institute: central south university

summary: The murine endogenous retrovirus MERVL is closely associated with the developmental potential of totipotency, manifesting dynamic and reversible activation within a small population of in vitro-cultured mouse embryonic stem cells (mESCs). While numerous regulators have been identified to influence the on and off switches of MERVL, how its activation is linked to the cellular decision-making of mESCs remains poorly understood. In this study, our genome-wide knockout screen uncovered an intimate relationship between MERVL activation and DNA damage response pathways. Specifically, we found that deficiency of Ints7, a backbone subunit of the Integrator complex, resulted in increased DNA damage, stabilization of p53, and hence transcriptional activation of MERVL. Moreover, we revealed that MERVL activation upon Ints7 knockdown was frequently accompanied by the activation of caspase 3. Intriguingly, a proportion of the caspase-activated mESCs were able to switch on MERVL meanwhile circumvent apoptosis and evade cell death, resembling a process known as anastasis. Altogether, our study highlights an association between MERVL activation and the p53-dependent cell fate plasticity in response to DNA damage, suggesting that limited caspase activation can alter developmental potential of stem cells.

overall_design: RNAseq, ATACseq and CUT&Tag are performed in shInts7 and shNT group E14TG2a cells. CUT&Tag targets Kap1S824p.

first: Shen

last: Yunfan

first: Ni

last: Huiling

first: Li

last: Li

first: Yuan

last: Kai


title: mRNA expression changes after SAM CRISPRa activator (MCP-p65AD-HSF1AD) expression in the primary effusion lymphoma cell line BC-3

status: Public on Jul 30 2025

city: Chicago

first: Eva

middle: Henriette

last: Gottwein

state: IL

address: 320 E Superior St., Tarry building, Room 6-752

department: Microbiology-Immunology

country: USA

institute: Northwestern University

type: Expression profiling by high throughput sequencing

summary: CRISPR-based transcriptional activation (CRISPRa) has extensive research and clinical potential. In our related work, we showed that commonly used CRISPRa systems can exhibit pronounced cytotoxicity. Here, we investigated mRNA expression changes after the inducible expression of a fusion protein that is commonly used for CRISPRa, including the activation domains (ADs) of the transcription factors p65 and HSF1, components of the synergistic activation mediator (SAM) CRISPRa system.

overall_design: BC3 cells that were stably transduced with the inducible lentiviral vector pTOZdeltaWRE-MPH or -hrGFP2 were plated at 400,000 cells/ml and induced with or without 100ng/ml doxycycline. 21 hour later, cells were recovered by centrifugation and subjected to dead cell removal (Miltenyi Biotec Dead Cell Removal Kit) as instructed. Resulting cell pellets were lysed in TRIzol (Invitrogen) and RNA was purified as instructed. MPH refers to a fusion protein of the MCP coat protein and the transactivation domains of p65 and HSF1.

first: Jesus A.

last: Ortega

first: Elizabeth T.

last: Bartom


title: Promoter-independent gene activation by Sigma factor-associated Cas12f proteins

city: New York

first: Samuel

last: Sternberg

address: 701 W. 168th Street, HHSC 726

institute: Columbia University

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Bacterial transcription initiation is a tightly regulated process that canonically relies on sequence-specific promoter recognition by dedicated sigma (σ) factors, leading to functional DNA engagement by RNA polymerase (RNAP). Although the 7 σ factors in E. coli have been extensively characterized, Bacteroidetes species encode dozens of specialized σ factors whose precise functions are unknown, suggesting additional layers of regulatory potential. Here we uncover an unprecedented mechanism of RNA-guided gene activation involving the coordinated action of σE factor in complex with nuclease-dead Cas12f (dCas12f). We screened a large set of genetically linked dCas12f and σE homologs in E. coli using RIP-seq and ChIP-seq experiments, revealing systems that exhibited robust guide RNA enrichment and DNA target binding with a surprisingly minimal 5ʹ-G target-adjacent motif (TAM). Recruitment of σE was dependent on dCas12f and gRNA, suggesting a direct protein-protein complex, and co-expression experiments demonstrated that the ternary complex was competent for programmable recruitment of the RNAP holoenzyme. Remarkably, dCas12f-RNA-σE complexes drove potent gene expression in the absence of any requisite promoter motifs, with transcription start sites defined exclusively by the relative distance from the dCas12f-mediated R-loop. Our findings highlight a new paradigm of RNA-guided gene activation that embodies natural features reminiscent of CRISPRa technology developed by humans.

sample_organism:
Escherichia coli
Flagellimonas taeanensis

overall_design: E. coli ChIP-seq and RIP-seq profiling for 16 RNA-guided endonuclease-dead Cas12f proteins. ChIP-seq profiling for a subset of dCas12f-associated σE proteins. E. coli and F. taeanensis RNA-seq profiling.

first: Florian

middle: T

last: Hoffmann

first: Tanner

last: Wiegand

first: Adriana

middle: I

last: Palmieri

first: Stephen

last: Tang

first: Juniper

last: Glass-Klaiber

first: Renjian

last: Xiao

first: George

middle: D

last: Lampe

first: Hoang

middle: C

last: Le

first: Chance

last: Meers

first: Leifu

last: Chang

middle: H


title: Drought Recovery in Plants Triggers a Cell-State Specific Immune Activation [MERFISH]

status: Public on Jul 29 2025

city: La Jolla

first: Joseph

middle: R

last: Ecker

state: CA

address: 10010 North Torrey Pines Road

department: Genomic Analysis Laboratory

institute: HHMI-Salk-Institute

summary: Rain-fed plants are subjected to cycles of drought and re-watering. Thus, efficient recovery from drought may be among the key determinants in the success of these plants, yet the mechanisms by which plant recover from drought have been historically understudied. To parse out the specific molecular processes leading to stress recovery, we performed a fine-scale time course of bulk RNA sequencing starting just 15 minutes after rehydration. We revealed that transcriptional drought recovery is an active and rapid process involving activating thousands of recovery-specific genes. To map the immediate recovery responses in diverse leaf-cell-types, we performed a single-nucleus transcriptome analysis of the onset of recovery from long-term moderate drought. Here we show the formation of unique cell states initiated rapidly following plant rehydration. We reveal the activation of a microbial-autonomic induction of the immune system and demonstrate that the onset of recovery leads to increased resistance against pathogens in Arabidopsis (Arabidopsis thaliana), wild tomato (Solanum pennellii) and domesticated tomato (Solanum lycopersicum cv. M82). Our findings uncover that the preventive immune activation was preserved despite extensive selection during tomato domestication. Since rehydration increases microbial proliferation and, thus, the risk for infection, pre-activating immunity may be crucial for plants survival in natural environments. This work lays the foundation for unraveling the complexed processes actively facilitating stress recovery within plant leaves.

overall_design: Spatial transcriptomics was performed using the MERSCOPE platform (Vizgen) to visualize gene expression across Arabidopsis leaf tissue under four water conditions: well-watered (time 0 and 15 minutes), drought, and 15 minutes post-rehydration.

first: Natanella

last: Illouz-Eliaz

first: Joseph R.


title: Transcriptomic profiling of prolonged-culture iPSC-derived kidney organoids and X-linked Alport syndrome organoid models carrying COL4A5 deep-intronic variants

city: Paris

first: Hassan

last: Saei

address: 24 boulevard de Montparnasse

department: Imagine Institute of Genetic Diseases

country: France

institute: Universite Paris Cite

summary: Kidney organoids serve as an invaluable platform for modeling hereditary renal diseases and developing therapeutic interventions. While various kidney organoid differentiation protocols have been developed, the protocol introduced by Morizane et al. was among the first to generate kidney organoids from induced pluripotent stem cells (iPSCs). By using a modified version of this protocol, we successfully generated kidney organoids in 21 days. Most studies on kidney organoids focus on early stages, typically between days 21 and 29, leaving the gene expression dynamics during prolonged culture less explored. In this study, we cultured healthy iPSC-derived kidney organoids for 22, 32, and 42 days and performed bulk RNA sequencing to investigate gene expression regulation during organoid culture. We also developed X-linked Alport syndrome (XLAS) organoid models carrying deep-intronic variants (severe and moderate models) to develop Antisense oligonucleotide (ASO) therapeutic approaches. Kidney organoids contain over 15 distinct cell types, making them a complex model for studying cell-type-specific maturation. By comparing organoids at early (day 22), mid (day 32), and late (day 42) time points, we observed significant changes in gene expression, particularly in genes related to the extracellular matrix, alongside the downregulation of podocyte-specific genes. Notably, we confirmed the upregulation of podocyte-specific collagen IV genes (COL4A3 and COL4A4), which are critical for forming the glomerular basement membrane (GBM). Overall, these findings underscore the importance of maintaining organoids in culture for at least 15 days beyond the last day of differentiation (day 21) to ensure the development of a more mature GBM, essential for BM disease modeling such as Alport syndrome.

overall_design: To investigate gene expression profile of kidney organoids at prolonged culture we harvested them at three time points early (day 22), middle (day 32), and late (day 42). Then we performed bulk RNA sequencing on organoids (three replicated in each condition) and compared conditions. To compare organoid models of X-linked Alport syndrome with isogenic controls and to study ASO impact in gene expression organoids treated with ASO for three weeks and harvested at day 38 of culture and bulk RNA sequencing was performed.

first: Corinne

last: Antignac

first: Geraldine

last: Mollet

first: Guillaume

last: Dorval


title: Mechanism and cellular actions of the potent AMPK inhibitor BAY-3827

status: Public on Jul 28 2025

city: Copenhagen

first: Lars

middle: Roed

last: Ingerslev

address: Blegdamsvej 3B

department: NNF Center for Basic Metabolic Research

country: Denmark

institute: Copenhagen University

summary: Inhibition of AMP-activated protein kinase (AMPK) is increasingly being explored for its therapeutic potential in some diseases, including certain types of cancers. However, AMPK-inhibitory tool compounds have largely been limited to compound C/dorsomorphin and SBI-0206965, both of which display numerous off-target effects and blocking AMPK-independent metabolic processes. Here we describe molecular insights and cellular actions/utility of a recently identified potent AMPK inhibitor BAY-3827. Sequence analysis of highly/lowly-inhibited kinases by BAY-3827, based on in vitro kinase selectivity profiling, predicted key conserved residues involved in the compound-inhibitory effect. A co-crystal structure of the AMPK kinase domain (KD)-BAY-3827 complex resolved at 2.5 Å in comparison with previously reported KD-inhibitor structures, revealed an overlapping binding site in the ATP-binding pocket and common αC helix-out conformations. We identified distinct features of BAY-3827-bound structure which involve a disulfide bridge between αD helix Cys106 and activation loop residue Cys174. This may help to stabilize AMPK conformation upon BAY-3827 binding, where the position of activation loop Asn162 leads the DFG motif Phe158 to adopt a conformation facing the C-terminal kinase lobe displacing His137, leading to a broken regulatory spine and an inactive kinase state. BAY-3827 at 2.5-5 μM, but not structurally resembling inactive BAY-974, fully blocked AMPK activator (MK-8722)-mediated phosphorylation of ACC1 and inhibition of lipogenesis in hepatocytes. Unbiased transcriptome analysis in MK-8722-treated wild-type and AMPK-null hepatocytes revealed that >30% of MK-8722-stimulated AMPK-dependent genes could be downregulated by 5 μM BAY-3827. Based on its greater selectivity and potency substantiated by comprehensive molecular/cellular investigations. BAY-3827 is a powerful tool to delineate AMPK functions.

overall_design: RNA-seq profiling of wildtype and AMPKa1a2 knock-out treated with DMSO, MK-8722, BAY-3827 or BAY-3827 + MK-8722

first: Conchita

last: Fraguas Bringas

first: Mohd

middle: S

last: Ahangar

first: Joyceline

last: Cuenco

first: Hongling

last: Liu

first: Alex

middle: B

last: Addinsall

first: Maria

last: Lindahl

first: Ashley

middle: J

last: Ovens

first: Mark

middle: A

last: Febbraio

first: Marc

last: Foretz

first: Olga

last: Göransson

first: John

middle: W

last: Scott

first: Elton

last: Zeqiraj

first: Kei

last: Sakamoto


title: Growth signaling promotes H2A.Z deposition by SRCAP to sustain somatic tissue regeneration [ChIP-Seq]

status: Public on Aug 01 2025

city: Evanston

first: Stephenie

last: Droll

address: Hogan 2-100 2205 Tech Drive

department: Molecular Biosciences

summary: In this study, we find that H2A.Z expression and chromatin occupancy dramatically decrease during normal epidermal differentiation. While the chromatin remodeler SRCAP is necessary and sufficient to maintain H2A.Z deposition in epidermal progenitors, EP400 is dispensable. Growth factor signaling mediated by the MAPK and PI3K signaling pathways is necessary to maintain both the expression and deposition of H2A.Z. Loss of SRCAP, H2AZ1, or H2AZ2 induces nuclear deformation and cytoplasmic DNA in progenitor keratinocytes which impairs DNA repair and proliferation.

overall_design: Additional H2A.Z ChIP-sequencing data performed in the progenitor or differentiated state with H2AZ1, H2AZ2, or SRCAP knockdown. We also include wild-type progenitor H3K9ac Cut&Run samples.

first: Nicholas

last: Chamberlain

first: Xiaomin

last: Bao


title: SRCAP mutation promotes cutatneous squamous cell carcinoma invasion [ChIP-Seq]

summary: In this study, we identify a mutational hotspot affecting SRCAP in cancer and that SRCAP is recurrently mutated in keratinocyte carcinomas. When introduced to primary human epidermal keratinocytes, the SRCAP hotspot mutation dysregulates gene expression in the differentiated state far more than the progenitor state. Gene dysregulation occurs without a change in H2A.Z chromatin occupancy. Finall, the presence of the SRCAP mutation in a RAS-CDK4 driven model of cSCC increases invasion significantly.

overall_design: This is H2A.Z ChIP-seq data in primary human keratinocytes expressing the identified SRCAP mutation in progenitor and differentiated states.


title: Enalapril mitigates senescence and aging-related phenotypes by targeting antioxidative genes via phosphorylated Smad1/5/9 [CUT&Tag]

status: Public on Jul 25 2025

city: Beijing

first: Haochen

last: Wang

address: Peking University, No.5 Yiheyuan Road Haidian District, Beijing, P.R.China

country: China

institute: Peking University

summary: Aging increases the risk of a myriad of chronic diseases, which are expensive and difficult to treat owing to their various risk factors. Repurposing existing medications has accelerated the development of therapies aimed at slowing aging. In this study, we revealed that enalapril, a drug widely prescribed for hypertension, can improve both cellular senescence and individual health. Mechanistically, phosphorylated Smad1/5/9 act as pivotal mediators of the anti-senescence properties of enalapril. It stimulates downstream genes involved in cell cycle regulation and antioxidative defense, facilitating cell proliferation and diminishing the production of reactive oxygen species (ROS), thus increasing the antioxidative ability of enalapril. At the organismal level, enalapril has been shown to bolster the physiological performance of various organs; it notably enhances memory capacity and renal function and relieves lipid accumulation. Our work highlights the potential of enalapril to augment antioxidant defenses and combat the effects of aging, thereby indicating its promise as a treatment strategy for aging-associated diseases and its use for healthy aging.

overall_design: CUT&Tag for transcription factor pSmad1/5/9 was used to detect the enrichment of pSmad1/5/9 with enalapril treatment


title: Enalapril mitigates senescence and aging-related phenotypes by targeting antioxidative genes via phosphorylated Smad1/5/9 [RNA-seq]

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design:
To analyze the effects of enalapril on the transcriptome expression of senescent cells, RNA-seq was performed in IMR90 cell line with different enalapril concentration treatment.
We performed RNA-seq to explore the effect of BMP receptor inhibitor LDN193189 (LDN) on enalaprill treatment.
To examine the effects of enalapril on the transcriptome of individual organs, after the feeding period, six organs—brain, kidney, liver, heart, muscle, and spleen—were collected for bulk RNA-seq analysis.


title: Human fetal kidney organoids model early human nephrogenesis and Notch-driven cell fate

city: Ramat Gan

first: Benjamine

last: Dekel

address: Tel Hashomer

country: Israel

institute: heba Medical Center: Tel Hashomer, Tel Aviv, IL

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

overall_design: Refer to individual Series


title: Transcriptome of macrolide resistant emm92 invasive group A Streptococcus in response to erythromycin exposure

status: Public on Jul 24 2025

city: Morgantown

first: Lei

state: West Virginia

address: 64 Medical Center Drive

department: Department of Microbiology, Immunology and Cell Biology

institute: west virginia university

summary: Since 2010 a macrolide resistant emm92-type strain of invasive group A Streptococcus has emerged in the United States, posing the question of whether acquired erm(T)-encoded resistance may have contributed to the rise in infections. Here, we present RNA sequencing of the emm92 transcriptome in response to erythromycin exposure.

overall_design:
RNA-seq profiling of invasive group A Streptococcus strains of emm-type 92 from patients in West Virginia. Two isolates of different MLSB sub-phenotypes (cMLSB or iMLSB) were characterized with or without exposure to 10 µg/mL of erythromycin for 1 hour.
Raw data were uploaded to SRA under BioProject PRJNA1182229. SRA Accessions are listed in Series supplementary file (SRA_Run_accessions.xlsx, SRR contains raw data for three replicates).

first: Powell

last: Lillie M.

first: Choi

last: Soo Jeon

first: Damron

last: F. Heath

first: Gangqing

last: Hu

first: Lukomski

last: Slawomir


title: Non-catalytic role of SETD1A promotes  gastric cancer cell proliferation through the E2F4-TAF6 axis in the cell cycle

city: Chiba

first: Masaki

last: Fukuyo

address: 1-8-1 Inohana, Chuo-ku

department: Department of Molecular Oncology

country: Japan

institute: Chiba University


title: Non-catalytic role of SETD1A promotes  gastric cancer cell proliferation through the E2F4-TAF6 axis in the cell cycle (RNA-seq)

summary: SETD1A catalyzes methylation of histone 3 lysine 4 (H3K4) and plays a critical role in development of multiple cancers. The abrrent expression of SETD1A relates to poor prognosis of gastric cancer (GC) patients. Here, we revealed that SETD1A is required for GC cell proliferation. Rescue experiment showed catalytic function is not necessary for GC cell growth and gene expression, whereas a novel non-catalytic FLOS domain is indispensable. The CRISPR screening targeting SETD1A targets identified that TAF6 acts as a downstream target of SETD1A which is essential for GC cell survival. Both SETD1A and TAF6 are required for G1/S cell cycle progression in GC cells. Non-catalytic component of SETD1A regulating expression of TAF6 through coactivating with E2F4s. These results demonstrate that non-canonical roles of SETD1A in GC cell, which supplys a potential therapeutic opportunity for GC.

overall_design: To investigate SETD1A regulated genes in gastric cancer cells, we established SETD1A wt or knockout cell lines (AGS, SNU719 and MKN45). Then RNA-seq was performed in these cells.

first: Meng

last: Ning

first: Takayuki

last: Hoshii

first: Bahityar

last: Rahmutulla

first: Atsushi

last: Kaneda


title: Non-catalytic role of SETD1A promotes  gastric cancer cell proliferation through the E2F4-TAF6 axis in the cell cycle (ChIP-seq)

overall_design: Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for SETD1A, H3K4me1/2/3, RNAP2, Ser2P, H3K36me3 in SETD1A wildtype or knockout AGS cells, and SETD1A, E2F4 ChIP-seq in wildtype AGS cells.


title: Chromatin accessibility analysis of human monocyte derived macrophages (hMDM) in response to H2O2 or DMNQ stimulation

city: Vienna

first: Laura

last: Boccuni

address: Währinger Gürtel 18-20

department: Cardiology

country: Austria

institute: Ludwig Boltzmann Institute for Cardiovascular Research

summary: Atherosclerosis is closely associated with oxidative stress, characterized by the generation of reactive oxygen species (ROS). ROS can serve as a polarization queue depending on localization. We observed that predominantly foam cells elicit high mitochondrial ROS burden in human and mouse atherosclerotic lesions. Our results show changes in chromatin accessibility with DMNQ, a mitochondrial-specific ROS inducer, compared to H202 treatment for induction of general ROS.

overall_design: Human monocyte-derived macrophages (hMDM) chromatin of untreated, as well as H2O2 or DMNQ. Monocyte-to-macrophage differentiation was carried out with M-CSF (macrophage-colony stimulating factor) in presence of H2O2 or DMNQ for 7 days and then cells have been seeded without M-CSF in presence of H2O2 or DMNQ for 24h. Data were generated by deep sequencing, with cells deriving from four indepedent donors, using Illumina sequencing.

first: Philipp

last: Hohensinner


title: Plasma exosomes from individuals with type 2 diabetes drive breast cancer aggression in patient-derived organoids

city: Boston

first: Christina

last: Ennis

state: Massachusetts

address: 72 E Concord St, K520, K520

institute: Boston University

summary: Women with obesity-driven type 2 diabetes (T2D) face worse breast cancer outcomes, yet metabolic status does not fully inform current standards of care. We previously identified plasma exosomes as key drivers of tumor progression; however, their effect on immune cells within the tumor microenvironment (TME) remains unclear. Using a novel patient-derived organoid (PDO) system that preserves native tumor-infiltrating lymphocytes (TILs), we show that T2D plasma exosomes induce a 13.6-fold expansion of immunosuppressive TILs relative to nondiabetic controls. This immune dysfunction may promote micrometastatic survival and resistance to checkpoint blockade, a known issue in T2D cancer patients. Tumor-intrinsic analysis revealed a 1.5-fold increase in intratumoral heterogeneity and 2.3-fold upregulation of aggressive signaling networks. These findings reveal how T2D-associated metabolic dysregulation alters tumor–immune crosstalk through previously underappreciated exosomal signaling, impairing antitumor immunity and accelerating progression. Understanding these dynamics could inform tailored therapies for this high-risk, underserved patient population.

overall_design: Patient-derived organoids (PDOs) were generated from resected breast tumor tissues of three untreated ER+ breast cancer patients. PDOs were treated with plasma-derived exosomes from normoglycemic (ND) and type 2 diabetic (T2D) donors. Following 3-day treatment, PDOs were enzymatically dissociated into single-cell suspensions, viability-selected by FACS, and processed using the 10X Chromium platform for scRNA sequencing.

first: Michael

last: Seen

first: Andrew

last: Chen

first: Heejoo

last: Jang

first: Adrian

last: Ilinski

first: Kiana

last: Mahdaviani

first: Naomi

last: Ko

first: Stefano

last: Monti

first: Gerald

middle: V

last: Denis


title: Opt1 imports CoA precursors as glutathione mixed disulphides

city: Groningen

first: Hein

last: Schepers

state: Groningen

address: Hanzeplein 1

department: Internal Medicine

country: Netherlands

institute: University Medical Center Groningen

summary: Pantothenate is a key vitamin for the intracellular biosynthesis of the essential molecule coenzyme A (CoA). Pantothenate can be biosynthesised or is taken take up by cells via plasma membrane transporters. In the cell, pantothenate, ATP and cysteine are required to synthesise CoA via five enzymatic steps. This canonical CoA biosynthesis route is well-studied in various organisms. Alternative routes that begin with the uptake of pantetheine (PanSH) or 4′-phosphopantetheine (PPanSH) as initial CoA precursors also exist. These alternative routes are vital for numerous unicellular organisms and are of interest for treating human diseases caused by defects in the canonical CoA biosynthesis pathway. In contrast to the uptake mechanisms for pantothenate, the cellular uptake mechanisms for PanSH and/or PPanSH are unresolved. Through a combination of in vivo experiments, yeast genetics and the use of chemical traceable compounds, we uncovered a non-canonical CoA biosynthesis pathway. We demonstrate that extracellularly, PanSH and PPanSH form mixed disulphides with glutathione, followed by uptake via the oligopeptide transporter Opt1. Once PanSH or PPanSH are imported, they are converted into CoA. Via this route several proteins essential for the canonical pantothenate-cysteine-dependent CoA biosynthesis pathway become dispensable. Additionally, we show that yeast strains cultured on PanSH or PPanSH have a growth advantage under conditions of decreased cysteine biosynthesis. The identified non-canonical CoA biosynthesis route provides a framework to treat CoA-linked diseases and to manipulate the growth of pathogenic or beneficial organisms that grow on PanSH or PPanSH.

overall_design: RNA‐sequencing of  Saccharomyces cerevisiae ecm31D cells with ecm31D cho2D cells.

first: Jouke

last: J. Wedman

first: Lotte

last: de Vries

first: Bart

last: van Lingen

first: Marianne

last: van der Zwaag

first: Rubén

last: Gómez-Sánchez

first: Ralph

last: Hardenberg

first: Wim

last: Huibers

first: Hjalmar

last: Permentier

first: Erick

last: Strauss

last: Chang

first: Fulvio

last: Reggiori

first: Anton I

last: de Kroon

first: Ody

last: Sibon


title: Drought Recovery in Plants Triggers a Cell-State Specific Immune Activation [RNA-seq]

status: Public on Jul 23 2025

first: Joseph

last: Ecker

summary: All organisms experience stress as an inevitable part of life, from single-celled microorganisms to complex multicellular beings. The ability to recover from stress is a fundamental trait that determines the overall resilience of an organism, yet stress recovery is understudied. To understand the stress recovery process, we studied the recovery from drought stress in Arabidopsis thaliana. We performed a fine-scale time series of bulk RNA sequencing starting 15 minutes after rehydration following moderate drought. We reveal that drought recovery is a rapid process involving the activation of thousands of recovery-specific genes. To capture these rapid recovery responses in different leaf cell types, we performed single-nucleus transcriptome analysis at the onset of post-drought recovery, identifying a cell type-specific transcriptional state developing independently across cell types. Furthermore, we reveal a recovery-induced activation of the immune system that occurs autonomously, and which enhances pathogen resistance in vivo in A. thaliana, wild tomato (Solanum pennellii) and domesticated tomato (Solanum lycopersicum cv. M82). Since rehydration promotes microbial proliferation and thereby increases the risk of infection, the activation of drought recovery-induced immunity may be crucial for plant survival in natural environments. These findings indicate that drought recovery coincides with a preventive defense response, unraveling the complex regulatory mechanisms that facilitate stress recovery in different plant cell types.

overall_design: RNA-seq profilling of Col and camta1 mutants samples at low water, well-watered, moderate stress and stress recovey stages.

first: Natanella

last: Illouz-Eliaz

first: Joseph R.


title: Genome-wide in vivo CRISPR screens identify GATOR1 as a potent tumor suppressor in MYC-driven lymphomagenesis

city: Melbourne

first: Marco

last: Herold

state: VIC

address: 145 Studley Rd

department: School of Cancer Medicine, La Trobe University

country: Australia

institute: Olivia Newton-John Cancer Research Institute

summary: Identifying tumor suppressor genes is predicted to inform on the development of novel strategies for cancer therapy. To identify new lymphoma driving processes that cooperate with oncogenic MYC (which is abnormally highly expressed in ~70% of human cancers) we have used a genome-wide CRISPR knockout screen in Eµ-Myc;Cas9 transgenic hematopoietic stem and progenitor cells in vivo. We discovered that loss of any of the GATOR1 complex components - NPRL3, DEPDC5, NPRL2 - significantly accelerated c-MYC-driven lymphoma development in mice. Low expression of the GATOR1 complex genes correlated with poor survival outcomes for human patients with high MYC-expressing cancers. Murine lymphomas lacking GATOR1 were highly sensitive to mTOR inhibitors as a single agent therapy, both in vitro and in vivo. These findings identify inhibition of mTORC1 as a potent tumor suppressive mechanism in c-MYC-driven lymphomagenesis and suggest a new avenue for therapeutic intervention in GATOR1-deficient lymphomas through mTOR inhibition.

overall_design: RNA-seq

first: Giner

last: Goknur

first: Potts

last: Maggie

first: Herold

last: Marco


title: Drought Recovery in Plants Triggers a Cell-State Specific Immune Activation

sample_organism:
Arabidopsis
Arabidopsis thaliana

platform_organism:
Arabidopsis
Arabidopsis thaliana
synthetic construct


title: sc-RNAseq analysis of whole aorta from ApoE-/- atherosclerotic mice after 4 weeks, 10 weeks and 16 weeks of high-fat-diet (HFD) and STAT5-in-1 (S5i) administration

first: Laura

last: Boccuni

summary: Atherosclerosis is closely associated with oxidative stress, characterized by the generation of reactive oxygen species (ROS). ROS can serve as a polarization queue depending on localization. We observed that predominantly foam cells elicit high mitochondrial ROS burden in human and mouse atherosclerotic lesions. Our results revealed that STAT5 inhibition predominantly affected Trem2hiGpnbmhi foam cell cluster.

overall_design: Following different time points of HFD feeding, whole aortas were isolated from ApoE-/- mice and processed for sc-RNAseq.

first: Frieda

last: Marka

first: Manuel

last: Salzmann

first: Elisabeth

last: Ableitner

first: Philipp

last: Hohensinner

first: Marion

last: Mussbacher


title: AKR1B10 dictates c-Myc stability to suppress colorectal cancer metastasis via PP2A nitration

status: Public on Jul 22 2025

city: Guangzhou

first: Mei

last: Song

address: Zhongshan No.2 Road 1th, Nonglin Street

institute: Sun Yat-sen University

summary: To elucidate the mechanisms underlying AKR1B10 deficiency-enhanced metastasis, we performed transcriptome profiling by RNA-seq to explore the signaling pathways affected by AKR1B10.

overall_design: Colorectal cancer cell HCT116 stably expressing shAKR1B10 or control shRNA were harvested for RNA preparation. Total RNA was extracted using TRIzol reagent and sequenced on Illumina Novaseq 6000 sequencing instrument (BGI, China). There were two experimental groups (shNC, shAKR1B10) with 3 replicates for each.


title: Drought Recovery in Plants Triggers a Cell-State Specific Immune Activations [bulk RNA-seq]

summary: To elucidate the transcriptional landscape during drought recovery we sequenced a fine-scale time-course of drought recovering plants.

overall_design: We performed gene expression profiling analysis using data obtained from RNA-seq of 9 time points from drought to 48 hour recovery, with a well-watered control for each time-point. For sterile stress-on-plates experiments, Col-0 seeds were sterilized and seeded on plates. Plates were kept in the dark at 4C for three days. After three days plates were moved to a growth chamber with short-day photoperiod light and 22 °C temperature. Samples are on 50% Low Water (LW) content for moderate stress, and 25% LW content for severe stress, each time point had a 100% LW content sample as control.

first: Jingting

last: Yu

last: Swift

first: Kathryn

last: Lande

first: Bruce

last: Jow

first: Za Khai

last: Tuang

first: Travis

last: Lee

first: Tatsuya

last: Nobori

first: Rosa Gomez

last: Castanon

last: Nery

first: Yotam

last: Zait

first: Saul

last: burdman


title: Human fetal kidney organoids model early human nephrogenesis and notch-driven cell fate

first: Benjamine

last: Dekel

summary: Pluripotent stem cell–derived kidney organoids (PSC-KOs) have been put forward as a preferred culture system in which to study human kidney development and disease. In contrast, the development of human fetal kidney tissue–derived organoids (hFKOs) that could serve as a benchmark for PSC-KOs has not advanced very far. Herein, utilizing a chemically defined serum-free medium, we established a long-term culture protocol for hFKOs that self-organize into polarized renal epithelium and faithfully recapitulate nephrogenic and ureteric bud cell lineages. Bulk transcriptomics, single-cell RNA sequencing, and pseudotime analysis along with immunostaining revealed diverse populations among hFKOs, with a preserved differentiation axis mostly at higher levels and to a greater extent than for PSC-KOs. Accordingly, compared to PSC-KOs, hFKOs were significantly enriched for the NOTCH signaling pathway genes, allowing single-cell analysis of changes following NOTCH inhibition. We observed a preferential increase of mature distal tubules at the expense of early proximal tubules and defined a new adaptive prominin 1–expressing (PROM1+, or CD133+) cell state that escapes NOTCH inhibition to generate proximal and distal tubules. Overall, the concomitant understanding of developmental processes using hFKOs is a key development for the fields of kidney stem cell biology and regenerative medicine.

overall_design: Characterization of  human fetal kidney organoids (hFKOs) at P0 and P2 via bulk RNA sequencing, along with adult kidney organoids (AKOs) and iPSC-derived kidney organoids (iPSC-KOs) for comparison. P0 (3 weeks cluture) of week 18  human fetal kidney, with 3 replicates. P2 (8 weeks of culture) of week 18  human fetal kidney, with 3 replicates. Adult kidney organoids from a single donor of an elective nephrectomy, 3 replicates from each passage. iPSC-derived kidney organoids day 16 of Freedman et al. differentiation protocol, two patient cell lines, 8GA7 and 123A9, 2 replicates from each donor.  In addtion, 3 replicates from the source tissue material of each organoid type were sequenced (Adult kidney tissue and human fetal kidney tissue).

first: Michael

last: Namestnikov

first: Osnat

last: Cohen-Zontag

first: Dorit

last: Omer

first: Benjamin


title: Deciphering the SUMO code in plants

status: Public on Jul 21 2025

city: Durham

first: Ari

last: Sadanandom

address: Stockton Rd

department: Biosciences

country: United Kingdom

institute: Durham University

summary: Post-translational modification (PTM) events generate proteoforms that orchestrate cell signalling in almost every biological process. The SUMOcode project aims to understand a critically important but understudied PTM in plants, SUMO (Small Ubiquitin-like Modifier). The rules governing specificity and function remain rudimentary for most PTMs, but the plant SUMO system provides a unique possibility to unravel the rules governing SUMOylation, as its core machinery comprises only 33 genes in Arabidopsis, compared with many hundreds for other PTMs. Our central hypothesis is that SUMO specificity is conferred through how cells are primed to respond to different stress signals, the tissue and cellular spatial distribution of SUMO machinery and substrates and control of SUMOylation modification via activation, repression and competition for PTM sites. Given the small numbers of genes involved in SUMOylation, we are in an excellent position to test our hypothesis employing state of the art multi-omics technologies to create the first SUMO Cell Atlas of any organism. Our ultimate goal is to 'enable' researchers and breeders to decipher the SUMO code in plants, enabling them to edit and rewrite the code, to develop crops that are future proofed against ongoing climate instability and change.

overall_design: Six-day old Arabidopsis seedlings were treated with 150mM of salt, 300mM of manitol, and 250nM of flagellin22. The samples were collected at 0 and 3 hours after the treatments.

last: sadanandom

first: Jason

last: Banda

first: Shraboni

last: Ghosh

first: Dipan

last: Roy


title: Time-Series Multi-Omics Identifies ZG16 as a Prognostic Immunoregulatory Driver of Colitis-associated Colon Cancer Development and Checkpoint Blockade Response

city: Shanghai

first: Lu

last: Chen

state: Xuhui

address: Fudan University, Shanghai

department: Shanghai Cancer Center

institute: Fudan University

summary: Chronic inflammation is a known driver of colitis-associated cancer (CAC), yet the trajectory of immune suppression and its reversibility remain poorly defined. Here, we performed a time-series multi-omics analysis combining paired transcriptomic and proteomic profiling, flow cytometry, and cytokine measurements in an AOM/DSS-induced model, further supported by single-cell transcriptomic and epigenomic datasets. This roadmap delineates a stepwise progressive evolution of immune suppression, capturing coordinated changes in immune cell composition, cytokine dynamics, and inflammation-associated mutagenesis throughout CAC progression. We identified ZG16 as a previously undervalued immune modulator that is progressively downregulated during CAC and CRC development. Analysis of CRC cohorts in the TCGA database confirmed that ZG16 loss correlates with PD-L1 upregulation, suppression of T cell costimulatory/proliferative programs, and both poor prognosis and resistance to immunotherapy. Functionally, ZG16 suppresses PD-L1 expression and promotes CD8+ T/NK cell infiltration, thereby reshaping the tumor immune microenvironment. In both inflammation-driven and syngeneic tumor models, recombinant ZG16 reduces tumor burden and enhances the efficacy of PD-1 blockade. These findings position ZG16 as a rheostat-like immunoregulatory factor and translationally relevant biomarker in inflammation-driven cancers.

overall_design: Eight to twelve weeks-old male C57BL/6 mice were used for the establishment of AOM/DSS murine model with mild amendments. We randomly assigned the mice into a control group (before administration, W0) and 4 experimental groups (2/5/8/11 weeks after administration, W2/5/8/11). The mutagen AOM (8.5 mg/kg body weight) was injected intraperitoneally into mice on days 1 and 8 of the modeling process. A week later, in order to cause damage to the colonic epithelium, the mice were given drinking water that contained 1.5% (w/v) DSS for seven days in a row. The mice were then provided regular drinking water for a period of 14 days. The mice thereafter received drinking water containing DSS for two or more cycles. Throughout the entire trial, the W0 group continuously was offered distilled water, and were euthanized simultaneously with the AOM/DSS-treated mice. During the trial, we selected 3 mice in each group for further RNA and protein analysis.

first: Peipei

last: Ding

first: Ling

last: Li

first: Yan

last: Hao

first: Jianfeng

first: Zening

last: Wang

first: Weiguo

last: Hu


title: Notch, β-catenin and MAPK signaling segregate endoderm and mesoderm in the diploblast Nematostella vectensis

status: Public on Jul 20 2025

first: Juan

middle: Daniel

last: Montenegro

state: Vienna

address: Djerassiplatz 1

department: Department of neurobiology and developmental biology

institute: University of Vienna

summary: Cnidaria are typically considered diploblastic (i.e. consisting of two germ layers) in contrast to their triploblastic sister clade, the Bilateria. However, a recent study suggested that sea anemones and other cnidarians have three segregated germ layer identities, corresponding to the bilaterian germ layers1. Here, we investigated, how the three germ layer identities are specified during early development of the sea anemone Nematostella vectensis. First, the mesodermal territory is specified at the animal pole at 6 hours postfertilization, followed by the specification of a ring of endodermal territory between mesoderm and ectoderm. We then assessed the role of β-catenin, MAPK and Notch signaling pathways during mesoderm and endoderm formation. Our results show that the mesoderm is initiated by MAPK signaling and simultaneously restricted to the future oral side by mutually exclusive nuclear β-catenin signaling. The mesodermal cells then express the Delta ligand, while the ectodermal cells express the Notch receptor. Inhibition of Notch signaling as well as ectopic expression of the Notch intracellular domain showed that endodermal tissue identity is induced by Notch signaling at the boundary between mesoderm and ectoderm. We propose a new model that outlines the different steps leading to the segregation of mesoderm and endoderm identities in Nematostella, confirming the presence of 3 distinct germ layer identities in this cnidarian. Notably, the observed crosstalk of MAPK, β-catenin and Notch signaling in the specification of three germ layers in Nematostella is highly reminiscent to early stage gastrulae of sea urchins suggesting that triploblasty may predate the split of cnidarians and bilaterians.

overall_design: time series of 10X genomics single-cell RNAseq libraries for 8, 10 and 12 hours post-fertilization of Nematostella embryos. Cell dissociation was done through cysteine treatment.

first: Ulrich

last: Technau

first: Emmanuel

last: Haillot

first: Tatiana

last: Lebedeva

first: Julia

last: Steger

first: Grigory

last: Genikhovich

first: Alison

last: Cole


title: Inflammatory Cytokine-Primed MSC-Derived Extracellular Vesicles

city: 달성군

first: Jun-Kook

last: Park

state: 대구광역시

address: 대구광역시 테크노북로

country: South Korea

institute: DGIST

type: Non-coding RNA profiling by high throughput sequencing

summary: Acute lung injury (ALI) is characterized by excessive inflammation and alveolar damage, arising from pathogens or systemic insults such as sepsis, and can progress to severe acute respiratory distress syndrome (ARDS). Current treatments, including mechanical ventilation, remain largely supportive, emphasizing the urgent need for the potent, cell-free therapeutic modalities. Mesenchymal stem cell-derived extracellular vesicles (MSC-EVs) have emerged as promising candidates for lung repair, but insufficient immunosuppressive capacity often limits their efficacy. Human adipose-derived mesenchymal stem cells (hADMSCs) were primed with IFN-γ and TNF-α to enhance the immunomodulatory properties of their secreted EVs. We characterized primed MSC-EVs (P-MEVs) and unprimed control MSC-EVs (C-MEVs)  by transmission electron microscopy, nanoparticle tracking analysis, and western blotting for EV markers. Functional assays in THP-1 and A549 cells examined anti-inflammatory potency and barrier regeneration against lipopolysaccharide (LPS)-induced damage. A preclinical mouse model of LPS-induced ALI was used to evaluate inflammatory cytokine expression, immune cell infiltration, pulmonary edema, and vascular leakage. Finally, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-infected Vero E6 cells were tested to explore the antiviral and anti-inflammatory potential of P-MEVs. Primed hADMSCs exhibited elevated expression of immunosuppressive molecules (e.g., COX-2, IDO, TSG-6), without changing EV morphology or yield. P-MEVs mitigated LPS-induced inflammation more effectively than C-MEVs in THP-1 and A549 cells. In vivo , P-MEVs more robustly attenuated inflammatory cytokines, immune cell recruitment, and lung injury markers in mice challenged with LPS. In SARS-CoV-2-infected Vero E6 cells, P-MEVs suppressed cytopathic effects and inflammatory responses more potently than C-MEVs. Mechanistic analyses revealed that these enhancements were associated with elevated miRNA levels, involved in inhibiting inflammatory pathways.

overall_design: we compared overall miRNA profiles between C-MEVs and P-MEVs using miRNA sequencing (miRseq) to investigate the potential contributing factors that enhanced the efficacy of P-MEVs We isolated human MSCs, stimulated them with IFN-γ and TNF-α, then extracted extracellular vesicles for miRNA sequencing.

first: Jongwon

last: Jeong

first: Moon-Chang

last: Baek

first: Kyungmoo

last: Yea


title: Drought Recovery in Plants Triggers a Cell-State Specific Immune Activation [snRNA-Seq]

first: Joseph

last: Ecker

address: 10010 North Torrey Pines Road

summary: Rain-fed plants are subjected to cycles of drought and re-watering. Thus, efficient recovery from drought may be among the key determinants in the success of these plants, yet the mechanisms by which plant recover from drought have been historically understudied. To parse out the specific molecular processes leading to stress recovery, we performed a fine-scale time course of bulk RNA sequencing starting just 15 minutes after rehydration. We revealed that transcriptional drought recovery is an active and rapid process involving activating thousands of recovery-specific genes. To map the immediate recovery responses in diverse leaf-cell-types, we performed a single-nucleus transcriptome analysis of the onset of recovery from long-term moderate drought. Here we show the formation of unique cell states initiated rapidly following plant rehydration. We reveal the activation of a microbial-autonomic induction of the immune system and demonstrate that the onset of recovery leads to increased resistance against pathogens in Arabidopsis (Arabidopsis thaliana), wild tomato (Solanum pennellii) and domesticated tomato (Solanum lycopersicum cv. M82). Our findings uncover that the preventive immune activation was preserved despite extensive selection during tomato domestication. Since rehydration increases microbial proliferation and, thus, the risk for infection, pre-activating immunity may be crucial for plants survival in natural environments. This work lays the foundation for unraveling the complexed processes actively facilitating stress recovery within plant leaves.

overall_design: snRNA-seq was performed on vermiculite-grown plants with two independent samples of four conditions: long-term moderate drought, recovery after 15 mins of rehydration, and two control groups: a well-watered control for drought, and a well-watered control after 15 mins of rehydration.

first: Natanella

last: Illouz-Eliaz

first: Jingting

last: Yu

last: Swift

first: Kathryn

last: Lande

first: Bruce

last: Jow

first: Za Khai

last: Tuang

first: Travis

last: Lee

first: Tatsuya

last: Nobori

first: Rosa Gomez

last: Castanon

last: Nery

first: Yotam

last: Zait

first: Saul

last: Burdman


title: data from Pregnancy exacerbates neutrophil responses in murine lungs and alters gut microbiota composition after cigarette smoke exposure

city: Utrecht

first: Ali

last: dehghani

state: - None -

address: Jonkvrouw Sanderijndreef 1113

institute: utrecht university

summary: Air pollution, particularly environmental tobacco smoke, poses significant health risks, especially to pregnant women and their infants. This study explores the difference in response to cigarette smoke (CS) exposure between pregnant and non-pregnant mice by examining lung transcriptomic profiles, neutrophil numbers, key mediators of neutrophil chemotaxis, and gut microbiota composition. Pregnant and non-pregnant mice were exposed to either air or CS. Bronchoalveolar lavage fluid (BALF) was analyzed for inflammatory cells and mediators. RNA sequencing was conducted on lung tissue to identify transcriptomic alterations.

overall_design:
in this study female mice were divided into four groups, pregnant and non-pregnant  mice exposed to air and pregnant and non-pregnant mice exposed to CS.

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last: Dehghani

first: Saskia

last: Braber

first: Gert

last: Folkerts


title: Integrated time-series analysis and high-content CRISPR screening delineate the dynamics of macrophage immune regulation [CROP-seq KO15]

first: Christoph

last: Bock

address: Lazarettgasse 14

institute: CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences

summary: Macrophages are innate immune cells involved in host defense. Dissecting the regulatory landscape that enables their swift and specific response to pathogens, we performed time-series analysis of gene expression and chromatin accessibility in murine macrophages exposed to various immune stimuli, and we functionally evaluated gene knockouts at scale using a combined CROP-seq and CITE-seq assay. We identified new roles of transcription regulators such as Spi1/PU.1 and JAK-STAT pathway members in immune cell homeostasis and response to pathogens. Macrophage activity was modulated by splicing proteins SFPQ and SF3B1, histone acetyltransferase EP300, cohesin subunit SMC1A, and mediator complex proteins MED8 and MED14. We further observed crosstalk among immune signaling pathways and identified molecular drivers of pathogen-induced dynamics. In summary, this study establishes a time-resolved regulatory map of pathogen response in macrophages, and it describes a broadly applicable method for dissecting immune-regulatory programs through integrative time-series analysis and high-content CRISPR screening.

overall_design:
To complement our time series analysis of macrophage stimulation with a functional dissection of macrophage regulons, we utilized our CROP-seq method for pooled CRISPR screening with single-cell transcriptome readout to genetically perturb putative key regulators involved in the macrophage response. Furthermore, we augmented CROP-seq with a CITE-seq readout for 11 macrophage surface marker proteins. We validated this integrated CROP-seq and CITE-seq assay in a targeted screen of 15 regulatory proteins over the time course of Listeria infection.
CROP-seq screening was performed in RAW 264.7 macrophages engineered to express Cas9. We delivered the CRISPR guide RNAs with a lentiviral vector at a multiplicity of infection of 0.1, in order to minimize the fraction of cells that may express more than one guide RNA. Moreover, to mitigate potential autocrine and paracrine effects of individual CRISPR perturbations, the guide RNA expressing cells were co-cultured with 90% of untransduced wildtype cells. The cells were challenged with Listeria infection and collected at three time points: 0 hours (prior to Listeria treatment), 2 hours, and 6 hours. CRISPR guide RNA expressing cells were purified via FACS and subjected to single-cell RNA-seq with specific enrichment for the guide RNAs that identify the CRISPR perturbations as part of the single-cell transcriptomes.
We targeted regulatory genes involved in Listeria infection (Rela/p65), endogenous interferon response (Jak1, Tyk2, Stat1, Stat2, Irf9), macrophage differentiation (Spi1, Csf1r, Irf8), epigenetic regulation (Ep300, Hdac6, Kdm1b, Kdm6b), and cell stress response (Jun, Creb1). Each gene was targeted with four different guide RNAs, and we included 10% non-targeting guide RNAs as controls. In addition to the single-cell transcriptome readout and sequencing of expressed guide RNAs, we quantified 11 cell-surface proteins using a CITE-seq readout, which included the classical macrophage markers CD11b, CD14, CD115, CD163, and MAC2; activation markers CD64, CD69, CD80, and CD95; and immune checkpoints CD274/PD-L1 and CD172a.

first: Peter

last: Traxler

first: Stephan

last: Reichl

first: Lukas

last: Folkman

first: Lisa

last: Shaw

first: Victoria

last: Fife

first: Amelie

last: Nemc

first: Daniele

last: Barreca

first: André

middle: F

last: Rendeiro

first: Florian

last: Halbritter

first: Nikolaus

last: Fortelny

first: Wolfgang

last: Weninger

first: Thomas

last: Decker

first: Matthias

last: Farlik


title: Integrated time-series analysis and high-content CRISPR screening delineate the dynamics of macrophage immune regulation [CROP-seq KO150]

overall_design:
To complement our time series analysis of macrophage stimulation with a functional dissection of macrophage regulons, we utilized our CROP-seq method for pooled CRISPR screening with single-cell transcriptome readout to genetically perturb putative key regulators involved in the macrophage response. Furthermore, we augmented CROP-seq with a CITE-seq readout for 11 macrophage surface marker proteins. After we validated this integrated CROP-seq and CITE-seq assay in a targeted screen of 15 regulatory proteins over the time course of Listeria infection, we applied it to characterize the role of 135 transcription factors and chromatin regulators in the immune response of macrophages across several timepoints.
CROP-seq screening was performed in RAW 264.7 macrophages engineered to express Cas9. We delivered the CRISPR guide RNAs with a lentiviral vector at a multiplicity of infection of 0.1, in order to minimize the fraction of cells that may express more than one guide RNA. Moreover, to mitigate potential autocrine and paracrine effects of individual CRISPR perturbations, the guide RNA expressing cells were co-cultured with 90% of untransduced wildtype cells. The cells were challenged with Listeria infection and collected at three time points: 0 hours (prior to Listeria treatment), 6 hours and 24 hours. CRISPR guide RNA expressing cells were purified via FACS and subjected to single-cell RNA-seq with specific enrichment for the guide RNAs that identify the CRISPR perturbations as part of the single-cell transcriptomes.
We targeted 135 potential regulators of macrophage immune response over the time course of Listeria infection. Each gene was targeted with four different guide RNAs, and we included 10% non-targeting guide RNAs as controls. In addition to the single-cell transcriptome readout and sequencing of expressed guide RNAs, we quantified 11 cell-surface proteins using a CITE-seq readout, which included the classical macrophage markers CD11b, CD14, CD115, CD163, and MAC2; activation markers CD64, CD69, CD80, and CD95; and immune checkpoints CD274/PD-L1 and CD172a.


title: Integrated time-series analysis and high-content CRISPR screening delineate the dynamics of macrophage immune regulation [RNA-seq]

overall_design: To chart the immune-regulatory landscape of macrophages, we exposed primary bone-marrow derived macrophages (BMDMs) from wildtype mice to six immune stimuli, including infectious pathogens (Listeria, LCMV, Candida), pathogen-derived stimuli (LPS), and pro-inflammatory cytokines (IFN-beta, IFN-gamma). We established dense time series of gene expression by RNA-seq with time points at 0, 2, 4, 6, 8, and 24 hours, to resolve the gene-regulatory dynamics of the primary and secondary responses triggered by each stimulus.


title: Temporal transcriptomic and proteomic profiling of causal variants in combination uncovers molecular drivers of phenotypic additivity

city: Chennai

first: Himanshu

last: Sinha

email: hsinha@gmail.com, sinha@iitm.ac.in

address: Dept of Biotechnology

country: India

institute: IIT Madras

summary: Genetic interactions are fundamental to the architecture of complex traits, yet the molecular mechanisms by which variant combinations influence cellular pathways remain poorly understood. Here, we answer the question of whether interactions between genetic variants can activate unique pathways and if such pathways can be targeted to modulate phenotypic outcomes. The model organism Saccharomyces cerevisiae was used to dissect how two causal SNPs , MKT189G and TAO34477C, interact to modulate metabolic and phenotypic outcomes during sporulation. By integrating time-resolved transcriptomics, absolute proteomics, and targeted metabolomics in isogenic allele replacement yeast strains, we show that the combined presence of these SNPs uniquely activates the arginine biosynthesis pathway and suppresses ribosome biogenesis, reflecting a metabolic trade-off that enhances sporulation efficiency. Functional validation demonstrates that the arginine pathway is essential for mitochondrial activity and efficient sporulation only in the double-SNP background. Our findings show how genetic variant interactions can rewire core metabolic networks, providing a mechanistic framework for understanding polygenic trait regulation and the emergence of additive effects in complex traits.

overall_design: The wildtype Strain (SS strain) and the allele replacement strain (MMTT) were grown in sporulation medium and temporal RNA-sequencing was done  at 9 different timepoints The samples were taken at 0h, 30min, 45min, 1h10min, 1h40min, 2h30min, 3h50min, 5h40min, and 8h30min . Each timepoint has 3 replicates each.

first: Srijith

last: Sasikumar

first: Shannara

last: Parkins

first: Suresh

last: Sudarsan


title: Integrative ATAC-seq and Cut&Run Profiling Reveals T-bet/Runx3 Epigenetic Regulation in Antiviral CD8+T Cells

status: Public on Jul 19 2025

city: Suzhou

first: Qiang

last: Shan

address: No. 100, Chongwen Road, Suzhou Industrial Park

institute: Suzhou Institute of Systems Medicine

summary: Effector CD8? T cells must swiftly activate cytotoxic programs while preserving lineage identity and repressing alternative cell fates. T-bet and Runx3 serve as pivotal transcriptional regulators in this process, yet their integrated regulatory mechanisms have remained elusive owing to insufficient data on chromatin accessibility and binding dynamics. This study provides a systematic analysis of their cooperative and distinct roles in shaping the epigenetic landscape.

overall_design: Through combined ATAC-seq and Cut&Run analyses, we dissect the synergistic and unique contributions of T-bet and Runx3 to the epigenetic architecture of antiviral CD8+ T cells.

first: Xiaoyu

last: Lin


title: Integrated time-series analysis and high-content CRISPR screening delineate the dynamics of macrophage immune regulation [ATAC-seq]

first: Christoph

last: Bock

overall_design: To chart the immune-regulatory landscape of macrophages, we exposed primary bone-marrow derived macrophages (BMDMs) from wildtype mice to six immune stimuli, including infectious pathogens (Listeria, LCMV, Candida), pathogen-derived stimuli (LPS), and pro-inflammatory cytokines (IFN-beta, IFN-gamma). We established dense time series of chromatin accessibility by ATAC-seq with time points at 0, 2, 4, 6, 8, and 24 hours, to resolve the gene-regulatory dynamics of the primary and secondary responses triggered by each stimulus.

first: Peter

last: Traxler

first: Stephan

last: Reichl

first: Lukas

last: Folkman


title: Deep tissue profiling of Populus stem at single nucleus level reveals uncharacterized cell types and cell-specific gene regulatory networks

city: GAINESVILLE

first: Henry

last: Schmidt

state: FL

address: 2033 Mowry road

institute: University of Florida

summary: The vascular cambium is responsible for the production of secondary xylem, which comprises the most abundant form of biomass on earth, wood. Despite its massive global importance, the regulatory networks underlying the production of secondary xylem remain partially ambiguous. Countless studies have used whole-tissue (bulk) RNA sequencing (RNA-seq) to characterize transcriptome changes during wood development in Populus and other tree genera. These studies were limited by their resolution and regulatory inferences. In contrast, novel single-cell RNA sequencing (scRNA-seq) technologies provide opportunities to gain insight into the transcriptional program that controls cellular lineages as they differentiate to form the various cell types in wood. In this study, scRNA-seq was carried out utilizing a pool of 11,673 nuclei derived from two replicates of stem samples of Populus trichocarpa to create a single-cell transcriptome atlas with an unprecedented resolution for a tree species. Cell-type specific marker genes were utilized to annotate 20 cell clusters representing the known cell types within the sampled tissue. Reporter gene assays were performed to confirm the cluster-specific cell-type identity of vessel elements, fibers, ray parenchyma, cambial cells, and sub-cell type vessel-associated cells. The developmental trajectory of cambial cells and their xylem-specific derivates was implemented to identify putative regulators that control distinct developmental lineages involved in vascular development and xylogenesis. Finally, an integration with a protoplast derived, single-cell, whole stem dataset was conducted to compare the proficiency of each methodology to capture the transcriptome.

overall_design: We have applied single cell sequencing approach utlizing  a novel protocol to isolate nuclei from complex plant tissues (https://doi.org/10.1371/journal.pone.0251149) and 10X Genomics scRNAseq to study stem tissue of Populus trichocarpa. A growth stage in which the stem is undergoing secondary growth was chosen to enable to the high resolution profiling of each cell type as they produce the most abundant biomass material on earth, wood.

first: Henry W.

first: Daniel

last: Conde

first: Wendell J.

last: Pereira

first: Paolo

last: Triozzi

first: Kelly M.

last: Balmant

first: Christopher

last: Dervinis

first: Matias

last: Kirst


title: The GPI-anchored aspartyl proteases encoded by the YPS1 and YPS7 genes of Candidozyma auris and their role under stress conditions.

status: Public on Jul 17 2025

city: Ciudad de México

last: Clark-Flores

state: Ciudad de México

address: Prol. de Carpio y Plan de Ayala. Col. Sto. Tomás

department: Microbiología

country: Mexico

institute: Instituto Politécnico Nacional

summary: Candidozyma auris is a multidrug resistance opportunistic pathogen, distinguished because of their thermotolerance, osmotolerance, and persistence in biotic and abiotic surfaces, attributes that seem linked to the cell wall composition. Here, seven putative genes encoding yapsins, extracellular aspartyl proteases GPI-anchored to the membrane or cell wall, were identified in the genomes of C. auris strains CJ97 and 20- 1498, from clades III and IV, respectively. C. auris YPS1 gene is orthologous to the SAP9 of C. albicans. The YPS7 gene is orthologous to YPS7 of C. glabrata and S. cerevisiae, so they could coincide in roles. The CauYPS1 and CauYPS7 expression increased under nutrient starvation, 1.5 M NaCl, and at 42 o C. Transcriptomic analysis of the 20-1498 strain suggests that in addition to glucose and iron homeostasis, autophagy and cell membrane fluidity play roles in thermal stress conditions. In silico analysis showed the interaction of pepstatin A with the catalytic domain of Yps1 and Yps7. This inhibitor, together with caffeine, had a subtle effect on the growth of C. auris. However, these compounds induce alterations in the cell wall. Thereby, yapsins CauYps1 and CauYps7 may play a role in the cell wall integrity of C. auris in stress conditions, and they could be a target of new antifungal or antivirulence agents.

overall_design: RNA-seq profile of Candidozyma auris clade IV strain 20-1498 from stationary-phase YPD cultures at 37°C, subsequently incubated at 37°C and 42°C for six hours in YNB medium with 2% dextrose and 0.5% (NH4)2SO4.

first: A

last: Vidal-Montiel

first: D

first: E

last: Valentín-Gómez

first: J

middle: P

last: Luna-Arias

last: Rosales-Cruz

first: C

last: Hernandez-Rodriguez

first: L

last: Villa-Tanaca

first: M

last: Juarez-Montiel


title: Failing heart-specific cardiac fibroblasts induce heart failure via c-Myc Expansion [snRNA-seq]

status: Public on Jul 18 2025

city: Bunkyo-ku

first: Manami

last: Katoh

state: Tokyo

address: 7-3-1 Hongo

department: Department of Cardiovascular Medicine

institute: The University of Tokyo

summary: Heart failure (HF) is a major global problem with increasing numbers of patients and deaths in many countries. Existing studies on HF have focused primarily on cardiomyocytes, with few studies targeting non-cardiomyocytes. This study focused on cardiac fibroblasts (CFs) as a cause of HF. Single-cell RNA sequencing of mouse hearts revealed six distinct subclusters of CFs at various stages after pressure overload, with one subcluster being specific to the HF stage. The transcription factor c-Myc is specifically expressed in HF-specific CFs. CFs-specific deletion of c-Myc ameliorates pressure overload-induced cardiac dysfunction without affecting fibrosis. The chemokine Cxcl1 is highly expressed in HF-specific CFs and downregulated in CFs-specific c-Myc knockout mice. Chromatin immunoprecipitation analysis revealed that c-Myc binds to the promoter region of Cxcl1 in CFs. Cxcr2, the receptor for Cxcl1, is expressed in cardiomyocytes, and blockade of the Cxcl1-Cxcr2 signalling pathway prevents pressure overload-induced cardiac dysfunction. The addition of CXCL1 reduces the contractility of cardiomyocytes of neonatal rats and human iPS-derived cardiomyocyte organoids. Human CFs from failing hearts expressed c-MYC and CXCL1, while CFs from control hearts did not. These findings suggest that HF-specific CFs play an important role in inducing HF by upregulating c-Myc and Cxcl1 and that CFs could be a novel therapeutic target for HF.

overall_design: Nuclei were extracted from hearts of dilated cardiomyopathy patients and control, and single-nucleus RNA-seq was performed on each sample. Human heart tissue samples were obtained from the left ventricular apex myocardium of four heart failure patients undergoing implantation of left ventricular assist devices due to reduced cardiac function and two patients with preserved cardiac function who underwent pathological autopsy.

first: Jin

last: Komuro

first: Shinsuke

last: Yuasa

first: Masaki

last: Ieda


title: Gene expression data from SQ-diEG-treated and BBN-induced bladder cancer mouse model at　8 and 16 W

city: Tsukuba

first: Farhana

last: Ferdousi

state: Ibaraki

address: Tennodai 1-1-1

institute: University of Tsukuba

summary:
Squalene(SQ), a natural triterpene with antioxidant, anti-inflammatory, and immunostimulatory properties, holds promise for cancer therapy. We examined a previously developed, diethylene glycol derivative of squalene (SQ-diEG) and investigated its in vivo anti-carcinogenic effects in bladder cancer. C57BL/6 mice(6–8 weeks old) were treated with 0.025% N-butyl-N-(4-hydroxybutyl)nitrosamine (BBN) to induce bladder cancer, with SQ-diEG or PBS (control) administered orally from Week 0 to 8 or 16. On the last day of administration, mice was sacrificed and its bladder collect for the RNA extraction.We performed an untargeted whole-transcriptomic analysis to explore the functionality of SQ-diEG.
Squalene(SQ), a natural triterpene with antioxidant, anti-inflammatory, and immunostimulatory properties, holds promise for cancer therapy. We examined a previously developed, diethylene glycol derivative of squalene (SQ-diEG) and investigated its in vivo anti-carcinogenic effects in bladder cancer. C57BL/6 mice(6–8 weeks old) were treated with 0.025% N-butyl-N-(4-hydroxybutyl)nitrosamine (BBN) to induce bladder cancer, with SQ-diEG or PBS (control) administered orally from Week 0 to 8 or 16. On the last day of administration, mice was sacrificed and its bladder collect for the RNA extraction.We performed an untargeted whole-transcriptomic analysis to explore the functionality of SQ-diEG.

overall_design: According to the manufacturer's guide, the RNA was extracted using Isogen kit (Nippon Gene Co. Ltd., Japan). Then, RNA quantity and quality were determined using the NanoDrop 2000 spectrophotometer (ThermoScientific, USA). DNA microarray analysis was conducted on urinary bladder samples of control and SQ-diEG-treated groups using the GeneChip WT PLUS Reagent Kit (ThermoFisher Scientific) and GeneChip™ Hybridization, Wash and Stain Kit (ThermoFisher Scientific) following the manufacturer's instructions. In brief, complementary DNA (cDNA) was synthesized from 100 ng of RNA solutions. cRNA was synthesized from in vitro transcription of cDNA and then purified and reverse transcribed. Finally, single-stranded cDNA (ss-cDNA) was synthesized, purified, fragmented, and labeled following the manufacturer's instructions. Cartridge Array Hybridization was performed using the Clariom S array (Mouse; ThermoFisher Scientific) on the GeneChip™ Fluidics Station (ThermoFisher Scientific). Scanning was performed using GeneChip Scanner (ThermoFisher Scientific). The raw image data obtained after scanning were analyzed using the Transcriptome Analysis Console (TAC) software (ver. 4.0.2, ThermoFisher Scientific). The raw data were normalized following the signal space transformation robust multi-chip analysis (SST-RMA) algorithm. Further, gene-level analysis was performed using the Limma Bioconductor package. For differential expression analysis, a One-Way ANOVA followed by an empirical Bayes correction was performed. The detected above back-ground (DABG) cutoff was set to 0.05. The positive vs negative area under the curve (AUC) value was set at greater than or equal to 0.7. Finally, genes that passed the filter criteria of p value < 0.05 (one-way between-subjects ANOVA) and fold change > 2 (in linear space) were considered as differentially expressed genes (DEGs).

first: Keisuke

last: Sano


title: Ablation of Linker Histone H1.4 Reveals Focal Regulation of AP-1 Directed Enhancers

city: Research Triangle Park

first: Nolan

middle: Garrity

last: Gokey

state: North Carolina

address: 111 TW Alexander Dr, Building 101 Room C432

department: DIR

institute: NIEHS

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

overall_design: Refer to individual Series


title: Epigenetic dysregulation of energy homeostasis drives aortic valve stenosis that is treatable with metformin [RNA-Seq]

city: Worcester

first: Chinmay

last: Trivedi

state: MA

address: 368 Plantation St, AS7-1047

department: Medicine

institute: University of Massachusetts Medical School

summary: This study aimed to characterize transcriptomic alterations in murine aortic valves following conditional deletion of Hdac3 in valvular interstitial cells. By comparing gene expression profiles between control and Hdac3-deficient valves, we aimed to identify molecular pathways regulated by HDAC3 that are crucial for maintaining aortic valve structure and function. These data provide insights into the epigenetic and metabolic mechanisms underlying non-calcific aortic valve disease.

overall_design: RNA seq on Aortic valves from 12-week-old control and Hdac3-knockout mice

first: Timothy

last: Cashman

middle: M


title: Epigenetic dysregulation of energy homeostasis drives aortic valve stenosis that is treatable with metformin [Cut & Tag]

summary: This study aimed to investigate epigenetic regulation in murine aortic valves by profiling H3K27ac enrichment in control and Hdac3-deficient valves, as well as HDAC3 and phosphorylated HDAC3 (Ser424) occupancy in control valves. Using CUT&Tag, we assessed genome-wide changes in enhancer activity and HDAC3 binding to identify regulatory elements and pathways disrupted by Hdac3 deletion in valvular interstitial cells. These data provide insight into the role of HDAC3 and its phosphorylation in maintaining epigenetic homeostasis and gene expression programs critical for aortic valve integrity.

overall_design: CUT&Tag on Aortic valves from 12-week-old control and Hdac3-knockout mice

first: Sherin

last: Saheera


title: The muscle-specific MEF2D⍺2 isoform promotes muscle ketolysis and running capacity in mice

city: Portland

first: Zheng

last: Xia

state: Oregon

address: 2720 S. Moody Avenue

institute: OREGON HEALTH & SCIENCE UNIVERSITY

summary: Ketone bodies are an alternate fuel source generated by the liver in response to low carbohydrate availability in neonates and after starvation and exhausting exercise in adulthood. The postnatal alternative splicing generates a highly conserved muscle-specific MEF2D⍺2 protein isoform of the transcription factor MEF2D. Here, we discovered that compared to WT mice, MEF2D⍺2 exon knockout (Eko) mice displayed reduced running capacity and muscle expression of all three ketolytic genes, BDH1, OXCT1, and ACAT1. Consistent with reduced muscle utilization of ketone bodies, MEF2D⍺2 Eko mice also showed increased ketone body levels in a tolerance test, after exercise, and upon feeding a ketogenic diet. Lastly, using mitochondria isolated from skeletal muscle, we showed reduced ketone body utilization and respiration in Eko compared to WT mice. Thus, we identified a new role of MEF2D⍺2 protein isoform in regulating skeletal muscle ketone body oxidation, exercise capacity, and its effect on systemic ketone body levels.

overall_design: Quadrips were isolated from ~20-week old WT and Eko mice and subjected to polyA RNA sequencing.

first: Kumar

last: Sushil

first: Ji

last: Xuan

first: Iqbal

last: Hina

first: Xiangnan

last: Guan

first: Mis

last: Brittany

first: Dave

last: Devanshi

last: Suresh

first: Besler

last: Jacob

first: Dash

last: Ranjan

first: Xia

last: Zheng

first: Singh

last: Ravi K.


title: Endothelial c-Maf protects against liver fibrosis by regulating chromatin accessibility to suppress pathogenic microvascular subpopulations [scRNA-seq]

city: Mannheim

first: Volker

last: Ast

address: Theodor-Kutzer-Ufer 1-3

department: NGS Core Facility

country: Germany

institute: University Hospital Mannheim

summary: Liver sinusoidal endothelial cells (LSEC) are highly specialized within the hepatic vascular niche, controlling liver function and disease pathogenesis by angiocrine signaling. Recently, we identified GATA4 as a major transcription factor controlling LSEC development and protecting against liver fibrosis. As the transcription factor c-Maf was strongly downregulated in Gata4-deficient LSEC, we hypothesized that c-Maf might be an important downstream effector of GATA4 in LSEC differentiation and liver fibrogenesis. Clec4g-iCre/Maf fl/fl mice (Maf-LSEC-KO) mice with LSEC-specific Maf deficiency were generated and liver tissue was analyzed by histology, immunofluorescence, and in situ-hybridization at the age of three months. LSECs were isolated for RNA-, ATAC-seq, and single-cell RNA-seq (scRNA-seq) analysis. The expression of MAF and its targets were analyzed in published human scRNA-seq data. Endothelial Maf deficiency resulted in perisinusoidal liver fibrosis without affecting metabolic liver zonation, accompanied by a switch from sinusoidal to continuous endothelial differentiation. Furthermore, endothelial Maf deficiency caused hepatic endothelial proliferation and expression of profibrotic angiocrine factors such as Pdgfb, Igfbp5, Sparcl-1, and Flrt2. scRNA-seq revealed replacement of zonated LSEC subpopulations by capillarized, proliferative, sprouting and secretory endothelial cell subset promoting liver fibrogenesis and angiogenesis. This fundamental dysregulation of LSEC gene expression and differentiation was caused by changes in chromatin accessibility and transcription factor network alterations at promoter and enhancer regions following loss of Maf. Notably, endothelial MAF expression was also significantly reduced in human liver cirrhosis patients. Hepatic endothelial c-Maf protects against metabolic dysfunction-associated steatohepatitis-like liver fibrosis and regulates endothelial differentiation and zonation by controlling chromatin opening. Notably, our findings may open up new avenues to develop angiotargeted strategies for hepatic disease prevention and liver repair.

overall_design: scRNA-seq experiment was performed to compare LSECs transcriptomic in WT and MAF KO mice

first: Manuel

last: Winkler

first: Sina

last: Kürschner-Zacharias

first: Rita

last: Manco

first: Christian

last: Schmid


title: Transcriptomic differences in cochlear cells isolated from mice implanted with dexamethason-eluting electrode arrays or standard electrode arrays

city: Bethesda

first: Robert

last: Morell

state: MD

address: 35A Convent Drive, Room 1F-103

institute: National Institute on Deafness and Other Communication Disorders

summary: The inflammatory foreign body response (FBR) following cochlear implantation (CI) can negatively impact CI outcomes, including increased electrode impedances. This study aims to investigate the long-term efficacy of dexamethasone eluting cochlear implant and locally delivered dexamethasone, a potent anti-inflammatory glucocorticoid, on the intracochlear FBR and electrical impedance post-implantation in a murine model. The left ears of CX3CR1+/GFP Thy1+/YFP (macrophage-neuron dual reporter) mice were implanted with dexamethasone-eluting cochlear implants (Dex-CI) or standard implant (Standard-CI) while the right ear served as unoperated control. Another group of dual reporter mice was implanted with a standard CI electrode array followed by injection of dexamethasone in the middle ear to mimic current clinical practice (Dex-local). Mouse implants were electrically stimulated with serial measurement of electrical impedance. Dex-CI reduced electrical impedance and inflammatory FBR in the murine model for an extended period. Dex-local in the murine model is ineffective for long-term reduction of FBR and electrode impedance. Our data suggest that dexamethasone eluting arrays are more effective than the current clinical practice of locally applied dexamethasone in reducing FBR and electrical impedance.

overall_design: Following cochlear implantation (CI), mice were euthanized at 33 days post-CI, implanted and contralateral cochlea were microdissected, tissues collected separately in DMEM F-12 media and lysed with accutase at 37°C for 30 minutes on a shaker. The media was replaced with 2 mL of DMEM F-12 containing 5% FBS to stop the lysis. The tissue was triturated for 2 minutes and filtered through a 20 µm filter (pluriSelect Life Science, El Cajon, CA, United States). The filtered cells were centrifuged at 300 x g for 3 minutes and resuspended in 90 µL MACS buffer (0.5% BSA in PBS). 10 µL of CD11b microbeads (71-097-142, Miltenyi Biotec, Auburn, Ca, USA) were added to the cell suspension and incubated for 15 minutes at 4°C, washed in MACS buffer followed by centrifugation 350 x g for 5 minutes resuspended in 500 µL MACS buffer. Cell suspension was then applied to a prewashed column in a magnetic holder to collect flow-through containing Cd11b negative cells. Column was then washed 3 times with 500 µL MACS buffer with negative flow-through captured each time. The CD11b microbead-bound cells were then eluted with 1 mL MACS buffer. Samples were centrifuged at 300 x g for 3 minutes and adjusted with MACS buffer to 3 x 106 cells/mL. Samples at this concentration were used for 10x cell capture. Single-cell captures were performed following the manufacturer’s recommendations on a 10x Genomics Controller. The targeted number of captured cells ranged from 3231 to 3572 per run. Library preparation was performed according to the instructions in the 10x Genomics Chromium Single Cell c’ Chip Kit V2. Libraries were sequenced on a Nextseq 500 instrument (Illumina, San Diego, CA) and reads were subsequently processed using 10x Genomics CellRanger analytical pipeline using default settings and 10x Genomics downloadable mm10 genome as previously described.

first: Muhammad

middle: T

last: Rahman

first: Rafal

last: Olszewski

first: Shoujun

last: Gu

first: Michael

last: Hoa

first: Marlan

last: Hansen


title: HEP14-activated PKC-ERK1/2 pathway boosts empowered-hADSCs for ovarian regeneration and functional restoration [RNA-seq]

city: Shenzhen

first: Liming

last: Gui

address: 1120 Lianhua Street

department: Obstetrics and Gynaecology

institute: Peking University Shenzhen Hospital

summary: Ovarian insufficiency (OI), including both premature ovarian insufficiency (POI) and age-related natural-aging ovarian insufficiency (ARNA-OI), significantly affects women’s health and remains persistent challenge in clinical treatment. In this study, bulk and scRNA-sequencing analysis indicated that HEP14 alters gene expression patterns in hADSCs and promotes the survival, antifibrotic, and proangiogenic capabilities, as well as the transdifferentiation potential of hADSCs. The key factors involved in these processes were upregulated by the HEP14-activated PKC-ERK1/2 signaling pathway. In both doxorubicin-induced POI and age-related natural-aged ARNA-OI mouse models, the combined administration of h-hADSCs and HEP14 encapsulated in PLGA microspheres (HEP14/PLGA) substantially enhanced ovarian regeneration and restoration of gonadal hormone levels through autocrine, paracrine, or endocrine mechanisms. Importantly, this combined therapy markedly improved the fertility of POI mice. These results demonstrate that the therapeutic potential of this combined therapy for POI and ARNA-OI, opening new avenues for clinical research and treatment strategies.

overall_design: The isolation of hADSCs was carried out using the previously described method15  with modifications. hADSCs were cultured in DMEM/F-12 medium (Gibco) supplemented 10% fetal bovine serum (Gibco), 1%Glumax (Gibco), 1%NEAA(Gibco) and 1% penicillin/ streptomycin (Gibco) at 37 °C in a humidified 5% CO2 incubator. Primary hADSCs at passage 4 treated with HEP14 or vehicle for 48h  were used as smaples for RNA-sequencing.

first: Jiajia

last: Sun

first: Kan

last: Liu

first: Qili

first: Yingtong

last: Di

first: Lin

last: Bai

first: Cunan

last: Lu

first: Qin

last: Zhong

first: Bin

last: Tang

first: Xiaojiang

last: Hao

first: Helen

last: Picton


title: HEP14-activated PKC-ERK1/2 pathway boosts HEP14-empowered hADSCs for ovarian regeneration and functional restoration

overall_design: The isolation of hADSCs from human subcutaneous adipose tissue was carried out using the previously described method with modifications. hADSCs were cultured in DMEM/F-12 medium (Gibco) supplemented 10% fetal bovine serum (Gibco), 1%Glumax (Gibco), 1%NEAA(Gibco) and 1% penicillin/ streptomycin (Gibco) at 37 °C in a humidified 5% CO2 incubator.  Primary hADSCs treated with HEP14 (h-ADSCs) or vehicle (c-hADSCs) for 48h at passage 4 were used as samples for performing scRNA-Sequencing.

first: Changzhong

last: Li


title: Comprehensive evaluation of bone marrow microenvironment in patients with autoimmune hemolytic anemia: from trephine biopsy to single cell RNA sequencing.

status: Public on Jul 14 2025

city: Barcelona

first: Giancarlo

last: Castellano

address: 2 Passeig Sant Joan de Déu, Esplugues de Llobregat, Barcelona, Spain

department: PCCB

country: Spain

institute: Hospital Sant Joan de Déu Barcelona

summary: Here we investigated bone marrow (BM) features in a large cohort of 97 patients with autoimmune hemolytic anemia (AIHA) and observed a high prevalence of hypercellularity, dyserythropoiesis, reticulin fibrosis, and T-cell infiltration (65%, 29%, 76%, and 69% of patients, respectively). These findings were associated with inadequate bone marrow compensation, more severe anemia at onset, and need of multiple treatments. In a subset of warm type AIHA patients we investigated BM microenvironment by single-cell RNA sequencing. We found distinct immune cell profiles across disease stages (diagnosis, remission, relapse). In particular, upregulation of inflammatory response pathways was noted in CD8+, CD4+, and monocyte subsets during relapse compared to diagnosis and remission. Moreover, by single-cell TCR sequencing, we found small T cell clones at diagnosis that may either disappeared or expanded at remission. Disappearing clones exhibited a naive CD8+ phenotype and were more likely to respond to glucocorticoid treatment. Expanding clones showed upregulation of cytotoxic T cell markers and may play a role in the transition to a chronic/relapsing phase. Finally, cytokine gene expression differed across disease phases. At relapse, pro-inflammatory cytokines such as TNF-alpha, IL-1, and IL-6 were upregulated in CD4+ and CD8+ T cells, while TGF-beta was downregulated, potentially counteracting the transition to chronic phase. This is the largest study evaluating BM histology and clinical characteristics, and the first evaluation of BM microenvironment by single-cell RNA sequencing. We showed a complex scenario encompassing T-cell infiltration, clonality, and up/down-regulation of cytokine genes, associated with a more severe and relapsing disease.

overall_design: Retrospective series of patients with available bone marrow trephines, plus a prospective series of AIHA patients evaluated by scRNAseq on BM aspirate at AIHA onset, remission, and relapse.

first: Bruno

last: Fattizzo

first: Matteo Claudio

last: Da Vià

first: Francesca

last: Lazzaroni

first: Alfredo

last: Marchetti

first: Alessio

last: Marella

first: Akihiro

last: Maeda

first: Antonio Giovanni

last: Solimando

first: Loredana

last: Pettine

first: Francesco

last: Passamonti

first: Niccolò

last: Bolli

first: Wilma

last: Barcellini


title: Targeting NRF2 and FSP1 to Overcome Ferroptosis Resistance in TSC2-Deficient and Cancer Cells

city: Cardiff

first: Darius

last: McPhail

address: University Hospital of Wales

institute: Cardiff University

summary: Background/Objectives: Ferroptosis, an iron-dependent form of cell death, shows promise as a target for therapy-resistant cancers that exhibit increased iron metabolism. Ferroptosis is primarily characterised by lipid peroxidation within cell membranes. However, many cancers evade ferroptosis by upregulation of specialised ferroptosis defence mechanisms. This study investigates ferroptosis susceptibility in Tuberous Sclerosis Complex (TSC) cell models, and ovarian and breast cancer cell lines to identify ferroptosis resistance mechanisms and therapeutic targets. Methods: We assessed ferroptosis susceptibility using the ferroptosis inducers, RSL3 and erastin. We explored ferroptosis resistance genes using inhibitors of NRF2 (ML385) and FSP1 (iFSP1). RNA-sequencing was conducted to identify dysregulated ferroptosis resistance genes and to better characterise NRF2 target genes. RNA sequencing was also performed to identify gene expression after treatment with pro-ferroptotic iron-oxide nanoparticles. Results: TSC2-deficient cells exhibited resistance to RSL3 and erastin-induced ferroptosis, which correlated with increased ferroptosis defence gene expression, including NRF2 and downstream targets. NRF2 inhibition re-sensitised TSC2-deficient cells to ferroptosis, confirming its protective role. However, FSP1 inhibition did not re-sensitise TSC2-deficient angiomyolipoma kidney tumor cells to RSL3. In contrast, FSP1 knockdown significantly enhanced ferroptosis sensitivity in ovarian (PEO1, PEO4, OVCAR3) and breast (MDA-MB-436) cancer cell lines. Notably, in MDA-MB-436 cells, FSP1 knockdown was more effective than NRF2 inhibition in reversing ferroptosis resistance. Conclusions: Our findings highlight NRF2 and FSP1 as key regulators of ferroptosis resistance in TSC2-deficient and cancer cells. However, the differential efficacy of targeting these pathways suggests that patient stratification may be necessary for optimal therapeutic strategies. Targeting NRF2 and FSP1 could enhance ferroptosis susceptibility, offering a potential therapeutic approach for ferroptosis resistance cancers.

overall_design: MDA-MB-436 cells. Cells were cultured in DMEM with 15% (v/v) FBS, 2mM L-Glutamine, 0.01mg/mL Insulin, and 100 U/mL penicillin/streptomycin.  Cells were subject to reverse transfection with siRNA SMARTpools (DharmaconTM ON-TARGETplus siRNA) targeting NFE2L2, AIFM2, or non-targeting siRNA (all at 50 nM), using DharmaFECT-1 transfection reagent (DharmaconTM, Horizon Discovery Ltd.). Alternatively, cells were treated with 500 µM (Fe2+) of iron-oxide nanoparticles coated in DLin-MC3-DMA for 48 h, with reverse transfection of non-targeting siRNA. After 72 h target knockdown, samples were collected in RNAprotect reagent (Qiagen, West Sussex, UK) prior to total RNA extraction with an RNEasy kit (Qiagen). Total RNA was quantified using a QubitTM RNA HS Assay.

first: Andrew

last: Tee

first: Alexandre

last: Faris-Orr


title: Failing heart-specific cardiac fibroblasts induce heart failure via c-Myc

city: Tokyo

first: Jin

last: Komuro

address: 35 Shinanomachi Shinjuku-ku

institute: Keio University School of Medcine

summary: Heart failure (HF) is a global concern, marked by limited therapeutic options. While existing studies primarily focus on cardiomyocytes, there is a notable absence of drugs targeting noncardiomyocytes for HF. We focused on cardiac fibroblasts (CFs), utilising single-cell RNA-sequencing analysis. The analysis of murine hearts revealed one subcluster exclusive to the HF stage. The transcription factor c-Myc is specifically expressed in heart failure-specific fibroblasts (HF-Fibro). Cardiac fibroblast-specific deletion of c-Myc ameliorates pressure overload-induced cardiac dysfunction without affecting fibrosis. To elucidate the molecular function of c-Myc in HF-Fibro, transcriptome analysis by RNA-seq was conducted, in vivo.

overall_design: Conditional deletion of c-Myc in CF were achieved by crossing c-Myc flox/flox homozygous mice with Tcf21 iCre hemizygous mice. c-Mycflox/flox;Tcf21 iCre (Myc-Tcf21-iCre) mice were used as c-Myc cardiac fibroblast-specific knockout (c-Myc KO) mice. Tamoxifen was administered intraperitoneally at a treatment dose of 45 mg/kg for 5 consecutive days to activate the inducible MerCreMer protein, thereby inducing Cre recombinase activity. Male mice (8–12 weeks old) underwent TAC surgery to induce cardiac hypertrophy, HF, or a sham operation. For RNA-seq sample preparation, total RNA was extracted from sham, TAC control, KO-TAC.

first: Hisayuki

last: Hashimoto

first: Shinsuke

last: Yuasa


title: Integrated time-series analysis and high-content CRISPR screening delineate the dynamics of macrophage immune regulation

city: Vienna

first: Christoph

last: Bock

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

Macrophages are innate immune cells involved in host defense. Dissecting the regulatory landscape that enables their swift and specific response to pathogens, we performed time-series analysis of gene expression and chromatin accessibility in murine macrophages exposed to various immune stimuli, and we functionally evaluated gene knockouts at scale using a combined CROP-seq and CITE-seq assay. We identified new roles of transcription regulators such as Spi1/PU.1 and JAK-STAT pathway members in immune cell homeostasis and response to pathogens. Macrophage activity was modulated by splicing proteins SFPQ and SF3B1, histone acetyltransferase EP300, cohesin subunit SMC1A, and mediator complex proteins MED8 and MED14. We further observed crosstalk among immune signaling pathways and identified molecular drivers of pathogen-induced dynamics. In summary, this study establishes a time-resolved regulatory map of pathogen response in macrophages, and it describes a broadly applicable method for dissecting immune-regulatory programs through integrative time-series analysis and high-content CRISPR screening.


title: Single-cell DNA methylome and 3D genome atlas of human subcutaneous adipose tissue

city: Los Angeles

first: Asha

last: Kar

state: CA

address: 695 Charles E. Young Drive South

department: Human Genetics

institute: University of California Los Angeles

type: Methylation profiling by high throughput sequencing


title: Synergistic Organelle-Microenvironment Regulation Strategy for Intervertebral Disc Degeneration Therapy by Reinforcing Mitochondrial Outer Membrane Stability and Sequestering exDNA

status: Public on Jul 16 2025

city: hangzhou

first: Zhao

last: lan

state: Zhejiang Province

address: Dinglan Street

institute: Hangzhou Traditional Chinese Medicine Hospital

summary: Free extracellular DNA (exDNA) accumulation within the cytoplasm is closely associated with chronic inflammation in intervertebral disc degeneration (IVDD). While numerous minimally invasive treatments targeting the various etiologies of IVDD have been developed, there remains a lack of systematic approaches to address the chronic inflammation caused by both intracellular and extracellular nucleic acids. In this study, we propose a novel strategy for treating IVDD that involves dual nucleic acid clearance. This approach targets and regulates mitochondrial DNA (mtDNA) release while clearing exDNA from the microenvironment, working synergistically to block the inflammatory cascade. We first analyzed the mechanism of chronic inflammation mediated by intracellular and extracellular nucleic acids during the progression of IVDD using single-cell sequencing and clinical sample analysis. Next, we developed a composite delivery system consisting of engineered nanovesicles with mitochondrial targeting functions (BNPMT) and a nucleic acid clearance hydrogel (H-P gel). BNPMT achieves targeted delivery to nucleus pulposus cells via membrane modification with triphenylphosphonium (TPP), while encapsulated BAI1, a small molecule drug, specifically inhibits mitochondrial outer membrane permeabilization (miMOMP). This reduces mitochondrial DNA (mtDNA) leakage into the cytoplasm, thereby blocking the mtDNA-cGAS/STING signaling pathway-mediated inflammatory response in nucleus pulposus cells. The H-P gel, crosslinked by Schiff base reactions and using G3-PAMAM dendrimers as functional crosslinkers, efficiently captures exDNA from the intervertebral disc microenvironment. This inhibits cfDNA-induced M1 macrophage polarization and the release of inflammatory cytokines. In an IVDD rat model, this dual-modal system demonstrated synergistic therapeutic advantages, significantly reversing the progression of IVDD over 8 weeks. This targeted nucleic acid clearance strategy provides a novel approach for treating IVDD and offers a theoretical framework for addressing other nucleic acid-related inflammatory diseases.

overall_design: RNA-seq profiling of wildtype Nucleus pulposus cells and Doxorubicin-induced senescence of nucleus pulposus cells and BNPMT intervention in senescence of nucleus pulposus cells

first: Lan

last: Zhao

first: Kaiyu

last: Liang

first: WenQiang

last: Cheng

first: Xizhuo

last: Zhou

first: Libin

last: Yang

first: Xinning

last: Mao

first: Weihui

last: Qi

first: Hongting

last: Jin

first: Wei

last: Zhang

first: Hao

last: Pan

first: Dong

last: Wang


title: Dual mitotic bookmarking by GAF condensates and H3K27ac orchestrates differential propagation of cell fate memory in neural development [ATAC-seq]

city: Beijing

first: Yan

last: Song

state: 北京

address: No.5 Yiheyuan Road

institute: Peking University

summary: In brain development, neural stem cells (NSCs) undergo asymmetric cell divisions to replicate themselves and meanwhile produce differentiating siblings. It remains obscure how NSCs preserve their self-renewing fate across mitosis. Even less is known how cell fate memory is differentially propagated to sibling daughter cells adopting distinct cell fates. Here we found that key differentiation genes are dually bookmarked by pioneer factor GAF (GAGA factor) and H3K27ac in asymmetrically-dividing Drosophila central brain NSCs. In daughter cells adopting NSC fate, GAF promotes self-renewal through timely inhibiting differentiation genes via HDAC1-mediated H3K27 deacetylation, whereas in sibling daughter cells adopting a differentiating fate, GAF occupancy is replaced by competitor SWI/SNF complex, allowing H3K27ac deposition retained and fast activation of differentiation genes. Thus, our study unveils a new paradigm by which cell fate memory can be differentially transmitted to sibling daughter cells via dual antagonistic mitotic bookmarking and selective molecular competition mechanism.

overall_design: ATAC-seq results for chromatin accessibility in type II neural stem cells (NSCs) and progenitors in Drosophila larval central brain .

first: Rulan

first: Jie

last: Liu

first: Zimo

first: Zili

last: Chen

first: Tanpeng

first: Yuying

last: Shen

first: Zejun

last: Lan

first: Jingyi

last: Chu

first: Haoxuan

last: Tang

first: Xiyue


title: Endothelial c-Maf protects against liver fibrosis by regulating chromatin accessibility to suppress pathogenic microvascular subpopulations

first: Carsten

last: Sticht

address: Theodor-Kutzer-Ufer

department: ZMF

institute: University Heidelberg

overall_design: 4 Maf-knockout samples vs. 4 control samples

first: Christian

middle: D

last: Schmid

first: Manuel

last: Winkler


title: Dual mitotic bookmarking by GAF condensates and H3K27ac orchestrates differential propagation of cell fate memory in neural development [CUT&Tag]

overall_design:
CUT&Tag results for GAF, HDAC1, Osa as well as the histone modifications H3K27ac, H3K14ac, H3K4me in Drosophila larval central brain type II neural stem cells or neural progenitors (NSCs are default).
CUT&Tag results for HDAC1 binding in interphase and mitotic type II neural stem cells (NSCs) in Drosophila larval central brain.