title: A dual-function microcin in Acinetobacter baumannii facilitates biofilm formation and interbacterial competition

city: Austin

first: Mady

last: Telford

state: TX

address: 2506 Speedway

department: Molecular Biosciences

institute: The University of Texas at Austin

summary: Acinetobacter baumannii is a frequent cause of drug-resistant nosocomial infections. Its ability to persist in harsh environments facilities its spread and is supported by formation of robust biofilms. Biofilms are comprised of communities of bacteria encased in a matrix of exopolymeric substances. Microcins are small (<10kDa) secreted antimicrobial proteins that mediate bacterial competition but have not been associated with biofilms directly. Here we describe the first dual action microcin from A. baumannii that has both antibacterial activity and is required for biofilm formation. We show the ability of this microcin to support biofilm formation is independent of its antibacterial action, and how this microcin forms amyloid fibers in secreted and purified forms, a function that is critical to biofilm formation. Scanning electron microscopy (SEM) shows this microcin surrounding A. baumannii in fibers suggesting it may act as support for polymetric biofilm components. Finally we map the molecular pathway used by this microcin to inhibit prey cells, which uncovers a new host protein that facilities microcin mediated-membrane disruption. This study reports the first instance of a multifunctional microcin, which uncovers new roles for this understudied class of antibacterial proteins.

overall_design: RNA seq profiling of A. baumannii 17978 with and without the MccAB1 microcin gene. RNA profiles were collected from stationary phase after 24 hours of growth.

first: Bryan

last: Davies


title: Trop-2 governs anti-metastatic desmosomal integrity

status: Public on Feb 10 2026

city: Brno

first: Ondřej

last: Vacek

email: vacek@ibp.cz, ooondra.vacek@gmail.com

address: Královopolská 135

department: Department of Cytokinetics

country: Czech Republic

institute: Institute of Biophysics of the Czech Academy of Science

type: Expression profiling by high throughput sequencing

summary: Trop-2 and DSG2 are proteins critical for the formation of intercellular contacts in healthy epithelial tissue. In cancer cells, their roles have been reported to vary, acting as either tumor suppressors or promoters depending on the context. In our study, deletion of Trop-2 or DSG2 in T-47D breast cancer cell lines resulted in a similar phenotype. In vivo, xenografts derived from these knockout cell lines exhibited an increased ability to form distant metastases. The present study links the observed in vivo metastatic phenotype to transcriptomic alterations identified in both knockout cell lines and their corresponding xenografts.

overall_design: We performed a differential gene expression analysis comparing control T-47D cells to Trop-2 and DSG2 knockout (KO) clones cultured in 2D in vitro. Next, we compared Trop-2 wild-type (WT) xenografts to DSG2 KO xenografts and Trop-2 KO xenografts to investigate transcriptomic changes in vivo.

first: Ondrej

first: Terézia

last: Kurucová

first: Boris

last: Tichý

first: Karel

last: Souček


title: Non-necroptotic MLKL function damages mitochondria and promotes hematopoietic stem cell aging

city: Memphis

first: Masayuki

last: Yamashita

state: TN

address: 262 Danny Thomas Place

department: Division of Experimental Hematology, Department of Hematology

country: USA

institute: St. Jude Children's Research Hospital

type: Genome binding/occupancy profiling by high throughput sequencing

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

overall_design: Refer to individual Series


title: Non-necroptotic MLKL function damages mitochondria and promotes hematopoietic stem cell aging [ATAC-seq]

summary: Hematopoietic stem cells (HSCs) survive many types of cellular stress but often lose their regenerative and lymphopoietic capacities as a result. Such functional decline also occurs with age, and dysfunctional HSCs with impaired mitochondria accumulate during aging. However, the molecular link between HSC stress response and age-related functional decline remains poorly understood. Here we show that multiple stress responses converge on the RIPK3-MLKL axis to induce age-related changes in HSCs. The necroptosis effector MLKL is readily activated by inflammation and replication stress and accumulates in HSC mitochondria. Consequently, activated MLKL does not cause cell death but impairs HSC self-renewal and lymphoid differentiation. Such MLKL-mediated functional decline also occurs in HSCs during organismal aging, with activated MLKL primarily mediating age-related mitochondrial damage and reduced glycolytic flux. Collectively, our results establish the RIPK3-MLKL axis as a key mediator of HSC aging and identify a necroptosis-independent role of MLKL in mitochondrial damage.

overall_design: ATAC-seq profiling of HSCs (Lin-/cKit+/Sca1+/Flk2-/CD48-/CD150+) isolated from 3- and 18-month-old wildtype (WT) and Mlkl-/- mice

first: Yuta

last: Yamada


title: Genetic landscape and functional exploration of kidney cancer predisposition  causality in cross-ancestral populations [ATAC-seq]

city: Tianjin

first: Chen

last: Kexin

address: No.22, Qixiangtai Road, Hepin

country: China

institute: Tianjin Medical University

summary: To functionally explore renal cell carcinoma (RCC) susceptibility variants, we performed CUT&Tag for H3K27ac and ATAC-seq on 769P cells to assess transcriptional activity. Subsequently, we conducted a CRISPR screen and CRISPR droplet sequencing (CROP-seq), which combines pooled CRISPR screens with single-cell RNA sequencing (scRNA-seq), to identify target genes potentially regulated by likely causal SNPs. Functionally, we established a novel association between rs28684409 and the oncogene RPL4 at the complex genetic locus 15q22.31. As the next step, we performed CRISPRi on rs28684409 and conducted RNA-seq to identify differential gene expression associated with this variant.

overall_design: ATAC-seq (Assay for Transposase-Accessible Chromatin using sequencing) in renal cancer cell line 769P. This experiment focuses on identifying open chromatin regions, revealing the accessibility of regulatory elements and their potential role in transcriptional regulation.

first: Kexin

last: Chen

first: Mulin Jun

last: Li

first: Hongji

last: Dai

first: Xinlei

last: Chu

first: Han

last: Du

first: Shutong

last: Yang

first: Yanxin

last: Yao


title: Genetic landscape and functional exploration of kidney cancer predisposition  causality in cross-ancestral populations

summary: To functionally explore renal cell carcinoma (RCC) susceptibility variants, we performed CUT&Tag for H3K27ac and ATAC-seq on 786-O cells to assess transcriptional activity. Subsequently, we conducted a CRISPR screen and CRISPR droplet sequencing (CROP-seq), which combines pooled CRISPR screens with single-cell RNA sequencing (scRNA-seq), to identify target genes potentially regulated by likely causal SNPs. Functionally, we established a novel association between rs28684409 and the oncogene RPL4 at the complex genetic locus 15q22.31. As the next step, we performed CRISPRi on rs28684409 and conducted RNA-seq to identify differential gene expression associated with this variant.

overall_design: RNA Sequencing (RNA-seq) to analyze gene expression in CRISPRi-targeted rs28684409 and rs179785 cells compared to control cells. This experiment aims to identify differential expression profiles and elucidate the functional consequences of targeting rs28684409 and rs179785, providing insights into its role in cellular pathways and potential implications in disease.


title: A sympathetic-basophil axis promotes skin inflammation

city: Guangzhou

first: xie

last: xinyang

address: 58 Zhongshan Second Road, Yuexiu District, Guangzhou City, Guangdong Province

institute: The First Affiliated Hospital, Sun Yat-sen University

summary: As the largest barrier organ of the body, the skin integrates immune and peripheral sensory nervous systems to respond to environmental stimuli. While sensory neuroimmune interactions in the skin are extensively studied, the role of the sympathetic nervous system (SNS) in skin immunity remains unclear. We show that stress exacerbates atopic dermatitis (AD)-like inflammation in murine models by driving basophil infiltration through SNS activation, independent of the hypothalamic-pituitary-adrenal axis.  By elucidating the crosstalk between SNS and inflammation pathways, our study underscores the critical role of autonomic responses in chronic inflammatory skin diseases.

overall_design: Following the ablation of tyrosine hydroxylase-positive (TH⁺) nerves by 6-hydroxydopamine hydrobromide, live CD45⁺ lineage⁻ (CD3e, CD5, CD11c, CD19, NK1.1)  cells of the ear skin of WT mice were sorted by fluorescence-activated cell sorting (FACS) and analyzed using scRNA-seq.

first: Fang

last: Wang

first: Xinyang

last: Xie


title: Non-necroptotic MLKL function damages mitochondria and promotes hematopoietic stem cell aging [RNA-seq]

overall_design: RNA-seq profiling of HSCs (Lin-/cKit+/Sca1+/Flk2-/CD48-/CD150+) isolated from 3- and 18-month-old wildtype (WT) and Mlkl-/- mice


title: Genetic landscape and functional exploration of kidney cancer predisposition  causality in cross-ancestral populations [screen]

overall_design: CRISPRi/a (786-O and Caki-1) cells were transduced with the pooled sgRNA library at low MOI of 0.5.Twenty-four hours after infection, cells were selected using puromycin (2 μg/ml) for 7 days. Cells were split into triplicates every 3 days, and maintained at 300X coverage throughout the screen. Cells were harvested in replicates on day 0, day 14 and day 21 after puromycin selection. sgRNA was amplified by PCR using specific primers, the amplified products were sent to the Beijing Genomics Institute (BGI) for library construction and sequencing.


title: Genetic landscape and functional exploration of kidney cancer predisposition  causality in cross-ancestral populations [CROP-seq]

overall_design: CRISPRi/a (786-O and Caki-1) cells were transduced with the CROP-seq library at high MOI. Cells were selected using puromycin (2 ug/ml) for 3 days after transduction, after which they were expanded and harvested at day 7. The cells were further processed using the Chromium Next GEM Single Cell 3ʹ Reagent Kits v3.1 (Dual Index) (10x Genomics) according to the manufacturer’s protocol. To enrich for sgRNA transcripts, PCR was performed on 10 ng of cDNA from the 3’ single-cell RNA libraries using specific primers. The libraries were sequenced on Illumina (Novogene), using a configuration identical to that of standard 10x Genomics libraries.


title: Bypass of blocking lesions by RNAPII reveals a novel stress induced by DNA damage [ChIP-seq]

city: Miami

first: Carolina

middle: Paula

last: Banuelos

state: Florida

address: 1580 NW 10th Ave

department: Medicine

institute: University of Miami

summary: Platinum-based compounds and ultraviolet (UV) irradiation produce bulky DNA lesions that stall RNA polymerase II (RNAPII), activating transcription-coupled nucleotide excision repair (TC-NER), RNAPII degradation, and global transcriptional shutdown. However, the consequences of RNAPII bypassing such lesions remain unclear. We identified the acetyltransferase p300 as a key regulator of TC-NER-dependent RNAPII removal from damaged chromatin via a USP7-dependent mechanism. Loss of p300 permits RNAPII to bypass transcription-blocking lesions, sustaining transcription and full-length mRNA production despite DNA damage. This leads to continued translation, endoplasmic reticulum (ER) stress, and activation of the unfolded protein response (UPR), compromising cell viability. Notably, this stress response resensitizes tumors resistant to platinum-based chemotherapy. Our findings reveal a vulnerability in tumor cells that evade transcriptional shutdown and define a synthetic lethal interaction between p300 inhibition and platinum-induced DNA damage, offering a targeted strategy to overcome chemoresistance.

overall_design:
Chromatin immunoprecipitation DNA sequencing (ChIP‑seq) for RNA Polymerase II (RPB1) was performed in WT or P300 knockout (△p300) HeLa cells at 0 or 2 hours post‑UV irradiation (20 J/m²).
For RPB1ser2 ChIP‑seq experiments using DRB (5,6‑Dichloro‑1‑β‑d‑ribofuranosylbenzimidazole), WT or △p300 HeLa cells were treated with siRNA of CSB or a control pool from Dharmacon. 24 after the transfection, cells were treated with 100 µM DRB for 3 hours. Following this treatment, the cells were split into two groups: one group was exposed to UV irradiation (20 J/m²), while the other group (control) was not irradiated. Five minutes after UV exposure, DRB was removed from both groups to allow transcription to restart. Samples were collected immediately after DRB washout (0 minutes) and at 45 minutes post‑UV irradiation for ChIP‑seq analysis. For RPB1 ChIP‑seq experiments using A485, WT HeLa cells were treated with either DMSO or 1 µM A485 1 hour before UV treatment (20 J/m²). For some groups, 1 µM of the USP7 inhibitor FT671 was added 1 hour before UV irradiation. Cells were harvested 2 hours post‑UV irradiation.
For the siRNA experiments, cells were transfected with a pool of siRNAs from Dharmacon using Polyplus reagent 24 hours before treatment with either DMSO or 1 µM A485. One hour post‑treatment, cells were divided into two groups: one group was exposed to UV radiation (20 J/m²) and the other (control) was not exposed. Cells were harvested 2 hours post‑UV radiation.
Chromatin immunoprecipitation DNA sequencing (ChIP‑seq) for RNA Polymerase II (RPB1) was performed in WT or P300 knockout (△p300) HeLa cells at 0 or 2 hours post‑UV irradiation (20 J/m²), with or without 1 h of treatment with Tripolide 50 nM.  Cells were harvested 2 hours post‑UV irradiation.

middle: P

last: Bañuelos

first: Lucas

middle: D

last: Caeiro

first: Pradeepkuma

middle: R

last: Cingaram

first: Felipe

last: Beckedorff

first: Daniel

last: Bilbao

first: Ramin

last: Shiekhattar

first: Ramiro

middle: E

last: Verdun


title: Focal white matter lesions drive inflammation, neuronal dysfunction and synapse loss in the grey matter

city: St Louis

first: Sabine

last: Dietmann

state: MO

address: 4565 McKinley Avenue

institute: Washington University School of Medicine

summary: Focal white matter lesions occur in most neurodegenerative disorders1-4. They are well characterised in multiple sclerosis (MS) but much less understood in other conditions. Despite occurring early in disease, white matter lesions are considered either independent of or secondary to grey matter neuroinflammation, synaptic loss and altered neuronal activity5-8. Notably their functional impact on neuronal circuits has been overlooked. To address this, we performed a focal white matter lesion in an anatomically well-defined circuit, in which white matter lesions occur in many neurodegenerative disorders. Here we show that focal white matter lesions evoke transient neuronal activity changes and microgliosis, synaptic loss and increased engulfment in the grey matter, which resolves by the time myelin regeneration completes. Synaptic loss and microgliosis are often considered detrimental but we show they are instead an integral part of the myelin regenerative process. When we prevent these transient changes in the grey matter, myelin regeneration is blocked in the white matter. Conversely, inducing myelin regeneration failure leads to chronic neuroinflammation in the grey matter, suggesting that myelin regeneration failure drives sustained microglial activation. This recapitulates the low-grade inflammation considered to be a dominant mechanism underlying neurodegeneration8-11. Hence, we present a novel mechanism that may underlie multiple conditions and highlight the potential of targeting myelin regeneration to prevent chronic neuroinflammation.

overall_design: To test for accompanying transcriptional changes in demyelinated neurons within the IO, we injected fast blue into the CCP along with EB and sorted and performed RNA-seq on labelled cells. We performed transcriptional profiling for 4 time points including Saline samples as a reference.

first: Omar

last: de Faria Jr

first: Stavros

last: Vagionitis

first: Ragnhildur

middle: T

last: Karadottir


title: Gene expression + ATAC multiome profiling of mPFC engram cells after OSK partial reprogramming

city: Lausanne

first: Cesar

last: Sierra

address: SV 2513 (Bâtiment SV)

department: SV

country: Switzerland

institute: EPFL

summary: Counteracting cognitive decline is an important goal in regenerative medicine. Recently, partial cellular reprogramming has emerged as a promising strategy to restore cellular function and promote tissue regeneration, but whether this approach can also reverse cognitive frailty is not known. In old mice and mouse models of Alzheimer’s Disease (AD) engram cells responsible for memory formation are functionally impaired, which prompted us to test the potential of their reprogramming to recover cognitive capacities. Here, by performing single nucleis multiome sequencing on the mPFC of the APP/PS1 AD mouse model, we found that partial reprogramming of its engram cells re-established aberrant epigenetic and transcriptional alterations.

overall_design: mPFC nuclei from 10 month old WT, APP/PS1 and APP/PS1+OSK mice were isolated by  Fluorescence-activated nuclear sorting (FANS)  and analyzed using 10X multiome sequencing (snRNAseq + snATACseq)

first: Gabriel

last: Berdugo-Vega

first: Johannes

last: Gräff


title: Genetic landscape and functional exploration of kidney cancer predisposition  causality in cross-ancestral populations [CUTnTag]

first: Chen

last: Kexin

overall_design: CUT&Tag (Cleavage Under Targets and Tagmentation) for histone modification H3K27ac (Histone H3 Lysine 27 Acetylation) in renal cancer cell line 769P. This experiment aims to map the distribution of H3K27ac, providing insights into active regulatory regions associated with gene expression.

first: Kexin

last: Chen

first: Mulin Jun

last: Li

first: Hongji

last: Dai

first: Xinlei

last: Chu

first: Han

last: Du

first: Shutong

last: Yang

first: Yanxin

last: Yao


title: Dynamics of phage-host interactions in Bacteroides fragilis resolved by single-cell transcriptomics [bulk RNA-Seq data]

status: Public on Feb 09 2026

city: Seattle

first: Dmitry

last: Sutormin

state: WA

address: 401 Terry Ave N

institute: Institute for Systems Biology

summary: Success of phage therapies is limited by bacterial defenses against phages. While a variety of anti-phage defense mechanisms has been characterized, how expression of these systems is distributed across individual cells and how their combined activities translate into protection from phages has not been studied. Using bacterial single-cell RNA sequencing, we profiled the transcriptomes of ~50,000 cells from cultures of a human pathobiont, Bacteroides fragilis, infected with a lytic bacteriophage. We quantified the asynchronous progression of phage infection in single bacterial cells and reconstructed the infection timeline, characterizing both host and phage transcriptomic changes as infection unfolded. We discovered subpopulations of bacteria that remained uninfected and heterogeneously expressed protective factors. Each cell’s vulnerability to phage infection was defined by combinatorial expression of multiple genetic loci, including phase-variable capsular polysaccharide (CPS) biosynthesis pathways, restriction-modification systems (RM), and a novel operon predicted to encode fimbrial genes. Acting in concert, these heterogeneously expressed anti-phage defense mechanisms create a phenotypic landscape where distinct protective combinations enable the survival and re-growth of bacteria expressing these phenotypes without acquiring additional mutations. The emerging model of complementary action of multiple protective mechanisms heterogeneously expressed across an isogenic bacterial population showcases the potent role of phase variation and stochasticity in bacterial anti-phage defenses.

overall_design: Exponentialy growing Bacteroides fragilis NCTC 9343 culture was cultivated in chopped meat carbohydrate (CMC) media and either infected or not with Bf12P1 phage. Total RNA was extracted and sequenced.

first: Norma

last: Morella

first: Neelendu

last: Dey

first: Anika

last: Gupta

first: Anna

last: Kuchina


title: Sialylated CD43 forms a glyco-immune barrier that restrains anti-leukemic immunity

city: Cambridge

first: Jooho

last: Chung

state: MA

address: 300 Binney Street

institute: Broad Institute//Massachusetts General Hospital

summary: Macrophages exert anti-tumorigenic activity through phagocytosis, but phagocytosis-enhancing therapeutics have not improved acute myeloid leukemia (AML) outcomes. To identify phagocytosis regulators, we performed CRISPR knockout screens in human AML cells co-cultured with human macrophages. We found that the “don’t eat me” signal CD47 inhibited mouse but not human macrophage phagocytosis. However, O-linked glycosylation and sialylation were strong negative regulators of phagocytosis. In AML, the cell surface mucin-like glycoprotein CD43 was the major effector of these pathways. Inhibition of phagocytosis by CD43 was dependent on the length of its ectodomain and independent of the macrophage sialic acid receptors SIGLEC-1, SIGLEC-7, and SIGLEC-9. The inhibitory effects of CD43 extended beyond human macrophages to natural killer and T cells. Thus, CD43 forms a glyco-immune barrier that restrains both innate and adaptive anti-leukemic immunity.

overall_design: Human monocyte-derived macrophages were stimulated with PBS or IFNg and subjected to RNA-Seq analysis

first: Mounica

last: Vallurupalli

first: Ashwin

middle: V

last: Kammula

first: Todd

last: Golub

first: Robert

last: Manguso


title: Effects of Short-Wavelength Light on Retinal Cell Development

city: hangzhou

first: xuhong

last: zhang

address: qingchun road, no. 79. shangcheng strait, hangzhou

institute: the first affiliated hospital, school of medicine, Zhejiang university

summary: Changes in the light environment exert a profound impact on vision; however, the specific mechanisms underlying their effects on retinal development remain poorly understood. In this study, we exposed mice to short-wavelength light from eye opening and performed single-cell RNA sequencing on their retinas two weeks later. Our results revealed that short-wavelength light significantly perturbs retinal cell development in mice, with the most pronounced differential impact observed on amacrine cells.

overall_design: Mice were exposed to either short-wavelength light or full-spectrum white light starting from postnatal day 14 (P14). After two weeks of exposure, retinal tissues were collected for single-cell RNA sequencing. Subsequent analyses included identification of differentially expressed genes, functional clustering, ligand-receptor interaction analysis, and RNA velocity assessment.


title: Core immunometabolic reprogramming of Mycobacterium tuberculosis-infected mouse lungs and stage-specific effects by glutamine supplementation

city: Newark

first: Ryan

last: Dikdan

state: NJ

address: 225 Warren St

department: Dept. of Medicine

institute: NJMS

summary: An emerging field of immunometabolism in tuberculosis (TB), an infectious disease caused by Mycobacterium tuberculosis, offers an opportunity for the development of urgently needed host-directed therapies (HDTs). Combining hypothesis-generating widely targeted metabolomics screening with transcriptome profiling in a mouse model of TB, we identify core metabolic programs and induction of signaling pathway mediated by mTORC1, the central regulator for cell metabolism, during immune cell activation. In addition to expected enhanced glycolysis and arginine biosynthesis and catabolism, induction of glutaminolysis-associated pathways, as well as serine synthesis and its catabolism by mitochondrial branch of folate cycle underscores the pivotal role of amino acids and mitochondria metabolism in orchestrating host immunity against M. tuberculosis. A further test of the hypothesis reveals that glutamine supplementation of M. tuberculosis-infected mice not only reduces lung bacillary load, but also ameliorates progression of lung pathology. Pathway analysis of transcriptomic data further unveils infection stage-specific effects of glutamine on the immunometabolic response of immune cells. Glutamine bolsters host protective immunity during the chronic phase of infection by promoting core metabolic pathways, including glycolysis, while attenuating the overall host immune response during the acute phase. Collectively, the data emphasize the central role of amino acid and mitochondrial metabolism, in concert with other signaling and metabolic pathways, including glycolysis, in meeting the demand of activating immune cells for energy, signaling molecules, and anabolic precursors. These novel observations will provide therapeutic strategies for the development of novel HDTs for TB treatment by targeting specific metabolic pathways.

overall_design:
To investigate the effects of glutamine supplmention on the immunometabolic response in lungs of Mtb-infected mice at different stages of the infection.
C57BL/6 mice were subjected to infection with approximately 100 Colony Forming Units (CFU) of Mycobacterium tuberculosis strain H37Rv in a low-dose aerosol model of Tuberculosis. Following infection, the mice were arbitrarily assigned to one of two groups: (1) phosphate-buffered saline (PBS) or (2) glutamine supplementation. Glutamine was administered orally via gavage at a dosage of 500 mg/kg/day throughout the duration of the experiments. On days 12, 25, and 55 post-infection, both infected mice and uninfected control mice were humanely sacrificed, and their lung tissues were collected and rapidly frozen in liquid nitrogen for subsequent RNA extraction and RNA-seq analysis.
Frozen lung samples were disrupted in RNAzol in a beadbeater and RNA was extracted using RNAzol® RT Column Kit (Molecular research Center, Cincinnati, OH)
The library construction was performed using NEBNext® Ultra™ II Directional RNA Library Prep Kit for Illumina® (New England BioLabs Inc., Massachusetts, USA) and RNA sequencing by a NovaSeq S4 V1.5 (Illumina, California, USA).

first: Qingkui

last: Jiang

middle: J

first: Yunping

last: Qiu

first: Irwin

last: Kurland

first: Karl

last: Drlica

first: Selvakumar

last: Subbian

first: Sanjay

last: Tyagi

first: Lanbo

last: Shi


title: Bypass of blocking lesions by RNAPII reveals a novel stress induced by DNA damage [RNA-seq]

first: Carolina

last: Banuelos

summary: Ultraviolet (UV) irradiation or platinum-based drugs generate bulky DNA lesions that impede transcription elongation by RNA Polymerase II (RNAPII). This unscheduled transcription elongation block triggers a stress response, which includes initiating transcription-coupled DNA repair, removing and degrading the halted RNAPII, and a global transcriptional shutdown. However, the molecular and cellular consequences of RNAPII successfully bypassing these blocking lesions have remained unknown. Here, we have identified the acetyltransferase p300 as a regulator of the stress response when RNAPII stalls at blocking lesions. Indeed, p300 interacts with the halted RNAPII and controls its removal and degradation in a USP7-dependent mechanism. Notably, in p300-deficient cells, transcription persists despite DNA damage, as RNAPII bypasses blocking lesions, leading to mutant mRNAs and splicing variants accumulation. This persistent transcription activity is particularly detrimental post DNA damage as it results in toxic translation activity and higher DNA damage levels. These observations provide the first evidence that bypass of blocking lesions by RNAPII inhibits DNA repair as well as the cellular stress response. Our findings offer essential insights into the mechanisms of resistance to DNA damage and suggest strategies to enhance the efficacy of platinum-based drugs, such as cisplatin, in anti-tumor therapies.

overall_design:
To characterize the impact of p300 deficiency on transcription control post DNA damage, we determined the number of genes affected in WT and p300-deficient cells using RNAseq from Chromatin or polysome-enriched fractions
RNAseq of Polysomes-enriched fractions (Poly), 5-EU labeled RNA purified from Polysome-enriched fractions (Poly_Purif), 5-EU labeled and purified RNA from DRB treated cells (DRB_5EU_Nonfrag) or total RNA samples (5EU) were collected and analyzed in HeLa WT and Δp300 (p300KO) cells at different time points after UV irradiation (20 J/m2). In some cases, the RNA was fragmented prior to the 5-EU purification step (DRB_5EU_Frag).
RNAseq of 5-EU labeled RNA purified from Polysome-enriched fractions (Poly_Purif) were collected and analyzed in HeLa WT and Δp300 (p300KO) cells 2 hs after UV irradiation (20 J/m2) or treatment with actinomycin D (2 µg/ml)


title: Islet versus enterochromaffin lineage allocation of hPSC-derived pancreatic progenitors can be modulated at both pre-progenitor and post-progenitor stages

city: Toronto

first: Maria

middle: Cristina

last: Nostro

address: 101 College Street

country: Canada

institute: McEwen Stem Cell Institute

summary: Little is known about the molecular mechanisms that underlie the allocation of human pluripotent stem cell-derived pancreatic progenitors (hPSC-PPs) into specific endocrine lineages. By systematically interrogating the components of pancreatic differentiation media, we identify methods to reliably generate hPSC-PPs with high-grade NKX6-1 expression, but find that this alone has a minimal effect on modulating the endocrine composition of hPSC-derived islets. We then test the effect of altering the signaling cues provided after progenitor specification and find that the BMP and FGF/MEK pathways can be manipulated to shift hPSC-PPs between islet and non-islet endocrine lineages. We further find that the method of hPSC-PP specification integrates with these later cues in a way that can be leveraged to selectively increase the frequencies of specific islet lineages or enterochromaffin (EC)-like cells. Using a model of disrupted murine islet development that gives rise to pancreatic EC-like cells, we identify prolonged NEUROG3 expression as a conserved feature associated with both human and murine pancreatic EC-like cell differentiation. In addition to expanding our understanding of pancreatic endocrine lineage allocation, these findings provide some of the first tools to fine-tune composition of hPSC-derived islets for research and therapeutic applications.

overall_design: Day 13 human pluripotent stem cell-derived pancreatic progenitors with enhanced NKX6-1 expression were differentiated towards either islet or EC lineages. At day 26, cells were dissociated, viability-sorted, and fixed. 24 hours later, they were sent for fixed RNA profiling.

first: Paraish

middle: S

last: Misra

middle: C


title: Immunosuppressive monocytes are enriched in hepatocellular carcinoma patients with liver dysfunction in a phase II trial of combination sorafenib and nivolumab [scRNA-Seq]

city: San Francisco

first: Bridget

last: Keenan

state: CA

address: 505 PARNASSUS AVE

institute: University of California San Francisco

summary: Background and aims: Immune checkpoint inhibition (ICI) and anti-angiogenic therapies are active in hepatocellular carcinoma (HCC), although patients with impaired hepatic function have worse outcomes.   Methods: We conducted a multi-center, open-label clinical trial to assess the safety and efficacy of the multikinase inhibitor, sorafenib, combined with nivolumab, in patients with advanced or unresectable HCC and varying liver function. In Part 1, we investigated the primary endpoint of the maximum-tolerated dose (MTD) of the combination in patients with Child-Pugh A or B7 HCC. In Part 2, we enrolled patients with Child-Pugh B HCC with the primary endpoint of grade >/=3 treatment-related adverse events (TRAE) incidence. Secondary and exploratory endpoints included immunologic biomarkers.   Results: Overall, 25 patients were consented and 16 eligible patients enrolled. In Part 1, dose-limiting toxicity occurred in 1 of 6 patients in Dose Level -1, and 2 of 5 patients in Dose Level 1; Dose Level -1 was determined to be the MTD. In total, 69% of patients experienced a Grade >/=3 TRAE, with similar distribution for patients with Child-Pugh A and B liver function (70%, 95% CI: 0.35, 0.93 vs 66.7%, 95% CI: 0.22, 0.96). The objective response rate was 6%. We found that patients with Child-Pugh B harbored more circulating suppressive monocytes at baseline.  Conclusions: While the combination of sorafenib and nivolumab demonstrated acceptable safety at the MTD in both Child-Pugh subgroups, the objective response rate was below the pre-specified threshold to be declared worthy of further exploration. A distinct immune cell profile in Child-Pugh B patients warrants further exploration to better define mechanisms of resistance and potential therapeutic targets in this population with unmet clinical need.

overall_design: Tumor samples were collected from patients undergoing standard-of-care resections. Samples were digested in RPMI containing Collagenase I & II (0.1 mg/ml, Sigma Aldrich) and DNAse I, minced, and digested for one hour using the GentleMACS system (Miltenyi Biotec). Isolation of live and CD45+ cells was performed using MACS LS columns (Miltenyi Biotec). The samples were then used immediately for single cell RNA sequencing. Droplet-based single cell RNA sequencing (scRNAseq) was performed using the 10x Genomics 5ʹ Reagent Kits version 1, according to manufacturer instructions. All sequencing was performed on an Illumina NovaSeq S4 sequencer with paired end 200 base pair read length and 25,000 reads per droplet for gene expression.

first: Zenghua

last: Fan

first: Bryan

last: Khuong Le

first: Quincy

last: Harris

first: Jocelin

last: Chen

first: Matthew

last: Clark

first: Averey

last: Lea

first: Li

last: Zhang

first: Alexander

last: Cheung

first: Frances

last: Lara

first: John

last: Gordan

first: Paige

last: Bracci

first: Spencer

last: Behr

first: Lawrence

last: Fong

first: Alan

last: Venook

first: Edward

last: Kim

first: Robin

middle: K

last: Kelley


title: Sunitinib Synergizes with Cisplatin by Suppressing DNA Repair Pathways in High-Grade Serous Ovarian Carcinoma Cells

city: Shihezi

first: Yeernaer

last: Hazaisihan

state: Xinjiang

address: Beierlu 121

department: School of Medicine

institute: Shihezi University

summary: Abstract Background: High-grade serous ovarian carcinoma (HGSOC) frequently develops resistance to platinum-based chemotherapy such as cisplatin, leading to high mortality. Sunitinib, a multi-target Receptor tyrosine kinase inhibitor, has shown potential in the treatment of ovarian carcinoma (OC). However, the synergistic effects between cisplatin and sunitinib, and the sensitization mechanism of sunitinib in HGSOC are not yet understood. Methods: The anti-tumor effects of sunitinib on HGSOC cells were assessed using CCK-8, EdU, colony formation, wound healing, Transwell, and flow cytometry assays. RNA sequencing was performed on sunitinib-treated cells, followed by differential expression, enrichment, and protein-protein interaction network (PPI) analyses. Genes on cisplatin sensitivity were predicted using the CellMiner and GEPIA3 databases. Synergy between sunitinib and cisplatin was evaluated using SynergyFinder 3.0, and DNA damage was assessed by γ-H2AX expression. Results: Sunitinib significantly inhibited proliferation, migration, and invasion in HGSOC cells, while further inducing cell cycle arrest, promoting necrosis. Besides, sunitinib downregulated critical DNA damage repair pathways, including homologous recombination, fanconi anemia, and base excision repair. Furthermore, sunitinib synergizes with cisplatin in HGSOC cells, enhancing DNA damage compared to monotherapy. Additionally, we screened out 25 sunitinib-downregulated core genes (SDCs). Drug-sensitivity analyses showed that higher SDCs expression was significantly associated with cisplatin resistance in OC. Notably, in cisplatin-resistant HEY-A8/DDP cells, sunitinib displayed stronger cytotoxicity than cisplatin. Conclusion: Sunitinib induces cell-cycle arrest and necrosis. In addition, sunitinib synergized with cisplatin and enhanced cisplatin sensitivity by impairing DNA repair pathways. Drug-sensitivity analyses showed that SDCs are associated with cisplatin resistance in OC, suggesting that sunitinib may help overcome cisplatin chemoresistance. Notably, sunitinib retains substantial cytotoxic activity in cisplatin-resistant cells. Together, these findings suggest that the sunitinib–cisplatin combination is a promising strategy to overcome cisplatin resistance in HGSOC.

overall_design: In this study, HEYA8 cells were treated with either sunitinib (treatment group) or an equivalent volume of DMSO (control group), with three biological replicates established for each condition. Total RNA was extracted from these samples and subjected to RNA sequencing. Differential expression analysis was subsequently performed to compare the transcriptomic profiles, thereby evaluating the impact of sunitinib on HEYA8 cells and high-grade serous ovarian cancer (HGSOC). Additionally, we included RNA-seq data from HEYA8 cells treated with Compound 32 for 24 hours to evaluate its regulatory effects on the HGSOC transcriptome. Total RNA was extracted from these samples and subjected to RNA sequencing. Differential expression analysis was subsequently performed to compare the transcriptomic profiles, thereby assessing the impact of both sunitinib and Compound 32 on HGSOC.

first: Yankun

last: Yu

last: Wei

first: Xiaoning

last: Li

first: Di

last: Zhou

first: Shaojie

last: Wang

first: Xiangting

last: Gao

first: Lin

last: Tao

first: Liang

last: Guo

first: Wei

last: Jia


title: Single-cell RNA sequencing of E13.5 and E15.5 epicardium and epicardium-derived cells from Wt1CreERT2;Srsf3+/+ and Wt1CreERT2; Srsf3fl/fl embryonic hearts

city: Oxford

first: Andia

middle: Nicole

last: Redpath

address: IDRM IMS-Tetsuya Nakamura Building, Roosevelt Dr, Headington

country: United Kingdom

institute: University of Oxford

summary:
Background:  The epicardium is a fundamental regulator of cardiac development, functioning to secrete essential growth factors and to produce epicardium-derived cells (EPDCs) that contribute most coronary vascular smooth muscle cells and cardiac fibroblasts. The molecular mechanisms that control epicardial formation and proliferation have not been fully elucidated. In this study, we found that the RNA-binding protein SRSF3 is highly expressed in the proepicardium and later in the epicardial layer during heart development.
Methods: We performed scRNA-seq analyses on epicardial and epicardial-derived cells from E13.5 Wt1CreERT2;Srsf3+/+ (control), E13.5 Wt1CreERT2;Srsf3fl/fl (mutant) and E15.5 Wt1CreERT2;Srsf3fl/fl (mutant) embryonic hearts in order to elucidate the role of SRSF3 in the epicardial lineage.
Results: Induction of SRSF3 deletion using Wt1CreERT2 resulted in impaired epicardial proliferation and EPDC formation at E13.5. Single-cell RNA-sequencing showed SRSF3-depleted epicardial cells were eliminated by E15.5 and the remaining non-recombined cells up-regulated Srsf3 to become hyperproliferative and compensate for the early deficit. This research identifies SRSF3 as a master regulator of epicardial cell proliferation.
E15.5 control raw data already available under GSE145832

overall_design: 10x Genomics sequencing of E13.5 hearts containing tdTomato positive cells labelled with tamoxifen induction at E9.5 (1mg) and E11.5 (1mg) of Wt1CreERT2; Srsf3+/+ and Wt1CreERT2;Srsf3fl/fl, from 3 pooled C57BL/6 embryos and 2 pooled C57BL/6 embryos respectively. SMART-Seq2 sequencing of E15.5 heart tdTomato positive sorted cells labelled with tamoxifen induction at E9.5 (1mg) and E11.5 (1mg) of Wt1CreERT2; Srsf3fl/fl from 4 pooled C57BL/6 embryos.

first: Nicola

last: Smart

first: Irina-Elena

last: Lupu


title: Alterations in energy production in a Drosophila model for the X-linked dystonia-parkinsonism-related Taf1 deficiency

city: Luebeck

first: Melissa

last: Vos

address: Ratzeburger Allee 160, building 67

department: Institute of Neurogenetics

country: Germany

institute: University of Luebeck, UKSH

summary: Background: X-linked dystonia-parkinsonism (XDP), an adult-onset neurodegenerative disorder, is caused by an SVA insertion in the TAF1 gene, containing a hexanucleotide, the length of which is correlated to the severity of the disease. The SVA insertion moderately disrupts gene expression; however, the underlying disease mechanism remains enigmatic. Methods: Here, we characterized a fly model for Taf1 deficiency and performed a pilot RNA sequencing analysis. Subsequently, we validated these findings in Taf1-deficient flies and in XDP patient-derived fibroblasts.  Results: We identified an upregulation of genes involved in lipid-dependent energy production as a compensatory mechanism to maintain proper ATP levels. However, studies in XDP patient-derived fibroblasts with minor TAF1 reduction did not confirm these findings. Conclusion: β-oxidation is elevated in flies with severe TAF1 reduction but not detected in XDP-patient fibroblasts, suggesting that this compensatory mechanism may only manifest above a critical TAF1 dosage threshold, absent in patient basal conditions. This finding thus suggests that dosage-dependent metabolic responses occur following TAF1 loss.

overall_design: A heterozygous Taf1 mutant line and its control was used to isolate RNA. RNAseq was performed to get a preliminary output alterations in KEGG pathways

first: Philip

last: Seibler

first: Christine

last: Klein


title: KCNH2 regulates developmental gene networks and cell fate in human pluripotent stem cells

city: Ann Arbor

first: David

middle: Kelly

last: Jones

state: Mi

address: 1150 W Medical Center Drive

institute: University of Michigan

summary: KCNH2 encodes the voltage-gated potassium channel hERG1, which conducts the rapid delayed rectifier current (IKr) that is essential for cardiac repolarization. Loss-of-function KCNH2 variants or pharmacological inhibition of hERG1 cause congenital long QT syndrome type 2 and acquired long QT syndrome, respectively. KCNH2 variants are also associated with intrauterine fetal death and sudden infant death syndrome, yet the contribution of KCNH2 in early human development remains unclear. To explore this, we generated a KCNH2-/- human embryonic stem cell (hESC) line, which exhibited reduced proliferation and increased cell death at high confluency compared to isogenic controls. Transcriptomic analysis revealed widespread dysregulation of developmental genes, indicating impaired differentiation and altered cardiovascular gene networks. Pathways involved in calcium homeostasis, cell motility, and chemotaxis were also disrupted. These findings identify KCNH2 as a bioelectric modulator of early human developmental gene networks, extending its function beyond cardiac electrophysiology and informing congenital disease mechanisms.

overall_design: RNA-seq profiling of isogenic WT and KCNH2-/- human embryonic stem cells was performed to identify KCNH2-dependent transcriptional programs. Cells were analyized under steady-state pluripotent conditions. Loss of hERG1 expression and maintenance of pluripotency were confirmed by immunocytochemistry. WT and KCNH2-/- hESCs were compared for proliferation, survival, and apoptotic activity using cell counting, phase-contrast imaging, and live-cell imaging with caspase-3/7 reporters. Differential expression and gene ontology analyses revealed disruption of developmental, cell cycle, and survival pathways, with selected targets validated by qPCR.

first: Eric

middle: N

last: Jimenez-Vazquez

first: Abhilasha

last: Jain

first: Arfa

last: Nazneen


title: Micro-RNA networks in T-cell prolymphocytic leukemia reflect T-cell activation and shape DNA damage response and survival pathways

city: Leipzig

first: Dennis

last: Jungherz

address: Johannisallee 32a

department: Hematology

institute: University of Leipzig Medical Center

summary: T-cell prolymphocytic leukemia (T-PLL) is a poor-prognostic mature T-cell malignancy. It typically presents with exponentially rising lymphocyte counts, splenomegaly, and bone marrow infiltration. Effective treatment options are scarce and a better understanding of TPLL's pathogenesis is desirable. Activation of the TCL1 proto-oncogene and loss-of-function perturbations of the tumor suppressor ATM are TPLL's genomic hallmarks. The leukemic cell reveals a phenotype of active T-cell receptor (TCR) signaling and aberrant DNA damage responses. Regulatory networks based on the profile of microRNA (miR) have not been described for T-PLL. In a combined approach of small-RNA and transcriptome sequencing in 46 clinically and moleculary well-characterized T-PLL, we identified a global T-PLL-specific miR expression profile that involves 34 significantly deregulated miR species. This pattern strikingly resembled miR-ome signatures of TCR-activated T cells. By integrating these T-PLL miR profiles with transcriptome data, we uncovered regulatory networks associated with cell survival signaling and DNA damage response pathways. Despite a miR-ome that discerned leukemic from normal T cells, there were also robust subsets of T-PLL defined by a small set of specific miR. Most prominently, miR-141 and the miR- 200c-cluster separated cases into two major subgroups. Furthermore, increased expression of miR-223-3p as well as reduced expression of miR-21 and the miR-29 cluster were associated with more activated Tcell phenotypes and more aggressive disease presentations. Based on the implicated pathobiological role of these miR deregulations, targeting strategies around their effectors appear worth pursuing. We also established a combinatorial miR-based overall survival score for T-PLL (miROS-T-PLL), that might improve current clinical stratifications.

overall_design: polyA-RNA data of healthy donors and T-PLL patients

first: Till

last: Braun

first: Linus

last: Wahnschaffe

first: Markus

last: Glaß

first: Janine

last: Altmüller

first: Stefan

last: Hüttelmaier

first: Alexandra

last: Schrader

first: Marco

last: Herling


title: Early events in transcription and genome organization after STAG2 reconstitution in human cancer cells [Hi-C]

city: CLEVELAND

first: Fulai

last: Jin

state: Ohio

address: 10900 Euclid Avenue

department: Genetics and Genome Sciences

institute: Case Western Reserve University

summary: Truncating mutations of the tumor suppressor gene STAG2, encoding a component of the chromatin-bound cohesin complex, are present in many human cancer types. Here we report for the first time the early effects of reconstituting physiological  levels of wild-type STAG2 in STAG2-mutant human cancer cells. Acute STAG2 reconstitution only modestly affected global gene expression - <1% of genes were altered by two-fold or greater. Only EFEMP1, encoding a secreted extracellular matrix glycoprotein, was induced by STAG2 in all experimental systems tested. There were similarly modest effects on chromatin loops - <1% of all chromatin loops were altered in intensity. and no loops were entirely STAG2-dependent. Loops strengthened by STAG2 reconstitution tended to be small, consistent with prior observations that STAG2-cohesin has less loop extrusion processivity than STAG1-cohesin. The STAG2-regulated “immediate early genes” were not accompanied by STAG2-regulated enhancer-promoter chromatin loops and no chromatin features were identified by integrative bioinformatics with newly generated ChIP-seq and ATAC-seq data that could explain their regulation by STAG2. Together these data indicate that the number of genes directly regulated by STAG2-cohesin is small, suggest that current models are likely insufficient to explain the mechanism of direct transcriptional regulation by STAG2-cohesin, and identify EFEMP1 as a potential effector of STAG2 tumor suppression.

overall_design: Human glioblastoma multiforme cell lines H4 and MOG-G-UVW (both with endogenous truncating mutations of the STAG2 tumor suppressor gene) were infected with adenoviral vector alone or a derivative expressing wild-type STAG2. 36 hours after infection the cells were harvested and prepared for Hi-C.

first: Tianyi

last: Yang

first: Jung-Sik

last: Kim

first: Clara

last: Mellows

first: Wanying

last: Xu

first: Alvin

last: Ya

first: Lisa

last: Sadzewicz

first: Luke

last: Tallon

first: Jann

last: Sarkaria

first: Todd

last: Waldman


title: Early events in transcription and genome organization after STAG2 reconstitution in human cancer cells [RNA-Seq]

overall_design: Human glioblastoma multiforme cell lines H4 and MOG-G-UVW (both with endogenous truncating mutations of the STAG2 tumor suppressor gene) were infected with adenoviral vector alone or a derivative expressing wild-type STAG2. 36 hours after infection the cells were harvested and prepared for RNA-seq.


title: CosMX spatial transcriptomics of mouse subcutaneous EO771 tumors to study synergistic effects of NEDD8 depletion and anti-PD-1 therapy

city: Uppsala

first: Yumeng

last: Mao

address: Dag Hammarskjölds Väg 20

country: Sweden

institute: Uppsala University

summary: We have established the phenotype that depletion of NEDD8 in tumor cell has synergestic effect with anti-PD1 therapy in mouse EO771 subcutaneous tumor model. In this study we performed CosMX spatial transcriptomics of mouse tumor samples to study the tumor intrinsic and immune environment changes that could explain the outcome.

overall_design: 4 mouse (1 per group) were injected subqutaneously with WT or NEDD8 KO EO771 breast cancer cell line, followed by i.p anti-PD1 treatment or IgG2a control treatment for every three days after tumor establishment. Tumors were collected on day 19 after cancer cells injection and three days after the last anti-PD1 treatment for scRNAseq.

first: Papakyriacou

last: Irineos

first: Lu

last: Yonglin

first: Bedos

last: Marta Rubies

first: Jarvius

last: Malin

first: Lapins

last: Maris

first: Puigvert

last: Jordi Carreras

first: Mao

last: YuMao


title: [18F]FDG PET/CT multiomics identifies Hedgehog-driven HPV-negative head and neck squamous cell carcinoma

city: Brno

first: Core Facility

last: Bioinformatics

address: Kamenice 5

institute: CEITEC - Masaryk University

summary: Head and neck squamous cell carcinoma (HNSCC) is a highly heterogeneous malignancy with limited predictive markers to guide personalized treatment, particularly in human papillomavirus (HPV)-negative cases, which exhibit poor outcomes. Identifying reliable biomarkers for prognosis and therapeutic response remains a critical challenge. In a retrospective cohort of 51 patients with primary HPV-negative HNSCC, we investigated the prognostic significance of the Hedgehog (HH) signaling pathway and its association with imaging biomarkers. Genomic and transcriptomic analysis revealed that HH pathway activation correlated with distinct [18F]FDG PET/CT radiomic features, notably the PET-derived “histogram:ih.max” - a surrogate for peak [18F]FDG uptake and was associated with inferior survival outcomes. Functionally, pharmacologic inhibition of HH signaling demonstrated anticancer efficacy across multiple models, including HNSCC cell lines, patient-derived tumoroids, and in vivo xenograft models. Importantly, HH inhibition altered imaging characteristics in HNSCC xenografts, leading to a measurable reduction in [18F]FDG uptake. This imaging phenotype closely mirrored our clinical findings, suggesting that [18F]FDG PET/CT radiomics may serve as a non-invasive biomarker to identify and monitor HH-driven HNSCC tumors. The integration of multi-level molecular profiling and functional imaging supports a potential precision oncology strategy, in which HH inhibition may offer a viable therapeutic approach for HPV-negative HNSCC. Our study underscores the value of [18F]FDG PET/CT multiomics in linking tumor biology with imaging features, paving the way for improved patient stratification and treatment monitoring. These findings provide a compelling rationale for further investigation into HH-targeted therapies in this aggressive subset of HNSCC and their clinical implications.

overall_design: Total RNA was isolated according to manufacture's recommenations with the RNeasy Mini Kit or the RNeasy FFPE Kit from control or JK184-treated HNSCC cell lines (CAL27 or SCC25) at three timepoints (6, 12 and 24 h) or HNSCC FFPE patient samples, repsectively. poly(A) mRNA from cell lines was sequenced with the Illumina NextSeq 2000 sequencer. poly(A) mRNA from patient samples was sequenced with AVITI 2x75 Sequencing Kit Cloudbreak FS High Output using the QuantSeq 3′ mRNA-Seq V2 Library Prep Kit FWD with UDI 12 nt in combination with UMI Second Strand Synthesis Module for QuantSeq FWD preparation.

first: Stefan

last: Stoiber

first: Daniel

last: Pölöske

first: Clemens

last: Spielvogel

first: Elisabeth

last: Gurnhofer

first: Michaela

last: Schlederer

first: David

last: Haberl

first: Cécile

last: Philippe

first: Dominik

last: Elmer

first: Richard

last: Moriggl

first: Heidi

middle: A

last: Neubauer

first: Nicolas

last: Blavet

first: Vojtěch

last: Bystrý

first: Karolína

last: Trachtová

first: Hanne

last: Verswyvel

first: Hannah

last: Zaryouh

first: Abraham

last: Lin

first: Christophe

last: Deben

first: Gregor

last: Heiduschka

first: Maik

last: Dahlhoff

first: Fritz

last: Aberger

last: Schramek

first: Marcus

last: Hacker

first: Alexander

last: Haug

first: Lukas

last: Kenner


title: MIB1-mediated H2AK119 mono-ubiquitination chromatin landscapes independent of canonical PRC1

city: Seoul

first: Hyebin

last: Park

address: 84 Heukseok-ro, Dongjak-gu

department: Life science

country: South Korea

institute: Chung-Ang University

summary: Polycomb repressive complex 1 (PRC1) is a central regulator of transcriptional repression through RING1B-mediated mono-ubiquitination of histone H2A at lysine 119 (H2AK119ub). Recent studies have suggested that the E3 ubiquitin ligase MIB1, previously characterized in Notch signaling, may also contribute to H2AK119ub deposition through a PRC1-independent mechanism. To investigate the chromatin-associated roles of MIB1, we generated ChIP-seq datasets for MIB1, RING1B, and H2AK119ub in the human colorectal cancer cell line HCT116. These datasets enable comparative analysis of genome-wide binding patterns, identification of shared and distinct target regions, and assessment of PRC1-dependent and PRC1-independent modes of H2AK119ub regulation.

overall_design: ChIP-seq was performed in HCT116 colorectal cancer cells. H2AK119ub was profiled using an H2AK119ub-specific antibody, with MIB1 and RING1B analyzed by anti-FLAG ChIP following overexpression of FLAG-tagged MIB1 or FLAG-tagged RING1B.

first: Sang Beom

last: Seo


title: RNA-seq of BJ fibroblasts transfected with control or FH siRNA

city: Paris

first: Mikel

last: Rezola Artero

state: Ile de France

address: 15 Rue de l'École de Médecine

department: Centre de Recherche des Cordeliers

country: France

institute: Inserm

summary: Bulk RNA sequencing was performed on BJ human fibroblasts transfected with control siRNA or FH-targeting siRNA to investigate transcriptomic changes associated with FH silencing. Four biological replicates per condition were analyzed.

overall_design: The RNAseq sequencing was performed by the GenomIC Cochin Institute INSERM U1016. BJ cells were transfected (siC vs. siFH) and cultured for 5 days before RNA extraction (4 samples per condition). Following RNA quality evaluation, libraries were prepared using the Stranded mRNA Prep protocol from Illumina, starting from 250 ng of total RNA. Paired end (2 × 51 bp) sequencing was performed on an Illumina NovaSeq platform.

first: Margot

last: Revel

first: Alessandra

last: Zarantonello


title: PHIP suppresses NuRD to enable the growth of SWI/SNF-mutant cancers

city: Memphis

first: John

last: Navarro

state: Tennessee

address: 262 Danny Thomas Place

institute: St. Jude Children's Research Hospital

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

overall_design: Refer to individual Series


title: Transcriptome sequencing study of WT iPSCs and R693Q iPSCs

city: Hangzhou

first: Jiayang

last: Wan

address: No. 79, Qingchun Road, Shangcheng District

institute: The First Affiliated Hospital, Zhejiang University School of Medicine

summary: Objective: Genome-wide gene expression profiles of patient-derived R693Q iPSCs and wild-type iPSCs were compared by performing a comprehensive RNA-Seq analysis.The goals of the study was to compare the differential gene expression and possible enriched signaling pathways between the two groups by RNA-seq analysis, including quantitative analysis, differential analysis and enrichment analysis.

overall_design: The samples in this study included the experimental group: R693Q iPSCs and the control group: WT iPSCs. Total RNA was extracted from three biologically independent replicates of each cell line (a total of six samples with three biological replicates per group), followed by library preparation and sequencing on Illumina platforms.


title: Duox-driven ROS release by glia promote regeneration in the adult Drosophila brain

city: Sant Joan d'Alacant

first: Marta

last: Neto

state: Alicante

address: Av. Ramon y Cajal, s/n

country: Spain

institute: Instituto de Neurociencias - CSIC/UMH

summary: Tissue damage activates immediate responses to restrict further harm and initiate repair. How injury sensing is coupled to regeneration is still not well understood. Here, we studied regenerative responses in the adult Drosophila brain, where proliferation is normally strongly restricted.  We show that localized brain damage triggers oxidative stress and diverse brain protective programs. We find that ROS generation by the NADPH Oxidase Duox on glial cells is responsible for injury-induced oxidative stress. Both genetic and chemical suppression of ROS in injured brains lead to impaired regeneration. In particular, selective knock-down of calcium-sensitive duox in glia, which show elevated calcium after injury, strongly reduces injury-induced proliferation. We further provide evidence that diffusing ROS can sustain the activity of pro-regenerative signaling, which is required to stimulate cell divisions. Although oxidative stress is generally considered as harmful in the brain, we uncover here an unanticipated beneficial role of transient ROS release by glia to promote brain repair.

overall_design: RNAseq profiling of glial cells from optic lobes of adult fly brains in a intact condition (non-injured, NI) and 48 hours after an injured condition (injured, INJ).

first: Carolina

last: Alves

first: Anabel

last: Simões

first: Carmo

last: Soares

first: Beatriz

last: Ferreira

first: Andreia

last: Augusto

first: Christa

last: Rhiner


title: single cell RNA sequencing (scRNA-seq) of mouse first molar and adjacent alveolar bone tissues with and without orthodontic loading

city: Chongqing

first: miao

last: tan

address: 426 Songshi North Road, Yubei District, Chongqing, China

institute: College of Stomatology,Chongqing Medical University

summary: Single-cell RNA sequencing (scRNA-seq) was performed on mouse first molar and adjacent alveolar bone tissues (within 1 mm, buccal and palatal sides). Two groups were analyzed: non-loaded control and orthodontic tooth movement (OTM) after 1 day. The study aims to reveal early cellular and transcriptional changes induced by orthodontic force.

overall_design: Healthy C57BL/6J mice (6–8 weeks old, 50 g) were used in this study. A nickel–titanium coil spring was attached to the maxillary right first molar and maxillary incisors using a 0.010-inch stainless steel ligature to provide mesial orthodontic force (50 g constant force) on the right first molar. The contralateral (left) first molar, with springs attached but without force application, served as the control. On day 1 after force application, the upper right maxillary first molar was extracted following removal of the orthodontic appliance. Alveolar bone tissues within 1 mm of the extraction socket (buccal and palatal sides), with gingiva carefully removed, were immediately collected and rinsed in ice-cold PBS containing 1% FBS. Tissues from each condition were pooled, minced into 1–2 mm fragments, and digested with 2 mL GEXSCOPE® Bone Tissue Dissociation Solution at 37 °C for 60 min. The digested suspension was filtered and centrifuged, followed by removal of red blood cells using 2 mL GEXSCOPE® RBC lysis buffer at 25 °C for 10 min. Single-cell suspensions were then obtained and processed for scRNA-seq using the 10x Genomics platform.Samples were pooled from 10 mice for each condition prior to sequencing.

first: Miao

last: Tan

first: Minyu

last: He


title: Integrated  single cell multiomic profiling and functional validation reveal distinct cellular routes to human plasma cell differentiation.

city: Tucson

first: Deepta

last: Bhattacharya

state: AZ

address: 1656 East Mabel St

department: Immunobiology

institute: University of Arizona

summary: Upon activation, B cells undergo profound transcriptional, epigenetic, and metabolic reprogramming to form antibody secreting plasma cells bearing little resemblance to their progenitors. Here, we used single cell RNA and ATAC sequencing of primary and in vitro differentiating human B cells to identify multiple distinct plasma cell subsets and differentiation routes depending on the starting cell type. In primary tonsils, we observed two distinct plasma cell subsets distinguished by expression of CD44 variant 9 (CD44v9), CD38, CD31, and CD10. A transient and rare CD30+ intermediate was identified both in primary tonsils and in vitro. CD40, TLR9, and cytokine stimulation of naïve and memory B cells formed CD30+ cells, which then formed plasma cells that were exclusively CD44v9+. CD30+ intermediates were not generated from primary germinal center B cells as they differentiated into plasma cells, which mostly lacked expression of CD44v9. Analysis of single cell multiomic data and pharmacological inhibition experiments demonstrated that the CD30+ intermediate was promoted by the transcription factor MEF2C. BAFF and APRIL promoted further maturation of these cells to CD44v9+ plasma cells. These data suggest that human germinal center-independent and -dependent ontogenies are biased towards distinct differentiation routes and terminal plasma cells.

overall_design: Multiome-seq: Single nuclei were isolated from tonsil, bone marrow, and in vitro differentiating samples. Joint scRNA- and scATAC libraries were prepared using the 10x Genomics platform.

first: Colin

last: Fields


title: ACSS2-Mediated Lysine Crotonylation Attenuates Senescence and Enhances the Therapeutic Efficacy of Adipose-derived Stem Cells in Inflammatory Bowel Disease [RNA-Seq]

city: Guangzhou

first: Ming

last: Yuan

state: Guangdong

address: 26 Yuancun Erheng Rd

institute: The sixth affiliated hostpital, sun yat-sen university

summary: Background Inflammatory bowel disease (IBD) remains a significant clinical challenge with limited curative options. Adipose-derived mesenchymal stem cells (ADSCs) hold therapeutic promise, but their anti-inflammatory efficacy is often compromised by cellular senescence. This study investigates the role of lysine crotonylation (Kcr) in ADSCs senescence and explores its therapeutic potential. Methods We analyzed Pan-Kcr levels in senescent ADSCs and evaluated the effects of sodium crotonate (NaCr), a crotonyl-CoA precursor, on senescence, proliferation, and anti-inflammatory function. A murine colitis model was used to assess therapeutic efficacy. Molecular mechanisms focusing on ACSS2-mediated Kcr regulation and H3K9 crotonylation (H3K9cr) at the ACSS2 promoter. Results Senescent ADSCs exhibited a marked decline in Pan-Kcr levels. NaCr treatment ameliorated senescence, enhanced proliferation, and improved anti-inflammatory capacity. ACSS2, a key regulator of Kcr, was downregulated in senescent ADSCs. Moreover, the anti-senescence effect of NaCr depended on ACSS2-mediated crotonylation. NaCr promoted H3K9cr modification at the ACSS2 promoter, forming a positive feedback loop that elevated Kcr levels. Mechanistically, ACSS2-mediated Kcr suppressed the NF-κB pathway to delay ADSCs senescence. Conclusion Our findings reveal an epigenetic pathway (ACSS2-Kcr-H3K9cr) regulating ADSCs senescence and propose Kcr modulation as a novel strategy to enhance ADSCs-based therapy for IBD.

overall_design: RNA-seq profiling of ADSCs at  passage 3 and passage 7

first: Senmao

last: Li

first: Shaopeng

last: Chen

first: Minghui

last: Zhu

first: Runfeng

last: Yu

first: Junfeng

last: Huang

first: Guanzhan

last: Liang

first: Chi

last: Zhang

first: Xiaowen

first: Ping

last: Lan

first: Xianrui

last: Wu


title: ACSS2-Mediated Lysine Crotonylation Attenuates Senescence and Enhances the Therapeutic Efficacy of Adipose-derived Stem Cells in Inflammatory Bowel Disease [ChIP-Seq]

overall_design: ChIP-seq of H3K9cr in ADSCs at  passage 3 and passage 7


title: Integrative transcriptomic analysis uncovers the miRNA-centric regulation of Japanese encephalitis virus infection in porcine trophoblast cells [RNA-Seq]

city: Yangzhou

first: Juan

last: Yang

address: 大学南路88号

department: College of Veterinary Medicine

institute: Yangzhou University

summary: To characterize porcine placental miRNAs, we performed small RNA-seq experiments using porcine placental tissues and the porcine placental trophoblast cell line pTr. To investigate the intracellular responses of pTr cells following JEV infection, RNA-seq and small RNA-seq experiments were conducted on pTr cells with or without JEV infection.

overall_design: We collected placental samples from first - parity naturally delivered Duroc gilts, pTr cell samples under normal culture conditions, and pTr cell samples at 24 and 48 hours post - infection with JEV as well as non - infected samples. Two replicates per sample were used for RNA-seq and small RNA-seq experiments.

first: Jiayao

last: Jiang

first: Jiajia

last: Gong

first: Shuai

first: Ming-an

last: Sun


title: Integrative transcriptomic analysis uncovers the miRNA-centric regulation of Japanese encephalitis virus infection in porcine trophoblast cells [miRNA-Seq]

type: Non-coding RNA profiling by high throughput sequencing


title: Independent validation of circulating microRNAs as biomarkers in a case-control study of adolescents with type 1 diabetes for more than 8 years

city: Vienna

first: Matthias

last: Hackl

state: Austria

address: Leberstrasse 20

country: Austria

institute: TAmiRNA GmbH

summary: MicroRNAs (miRNAs) are epigenetic regulators of gene activity. Analysis of circulating miRNAs enables minimal-invasive studies of disease mechanisms and identification of novel disease biomarkers. The aim of this case-control validation study was to investigate previously identified circulating miRNAs in Swedish adolescents with long-duration (8.0–16.5 years) type 1 diabetes (T1D), and healthy matched controls to confirm their utility as biomarker candidates to diagnose and monitor the progression of T1D. Quantitative PCR analysis of 23 previously reported miRNAs was performed in 24 T1D and 24 control individuals. Body composition was assessed by dual-energy X-ray absorptiometry and peripheral quantitative computed tomography. Prospectively collected clinical data were retrieved from the Swedish diabetes quality registry. The selected miRNAs showed higher variability in both male and female T1D groups compared to controls. Statistical analysis confirmed differences for 12 miRNAs in comparison with controls, including miR-223-3p and miR-135a-5p, which previously were reported to be associated with T1D. MiR-34a-5p and miR-210-3p were positively associated with T1D duration and HbA1c (average from the last year), respectively. In conclusion, 12 previously reported miRNAs showed consistent differential expression between individuals with T1D and controls. Among these were miR-223-3p and miR-135a-5p, which are associated with cardiovascular/inflammatory disease and cancer, respectively.

overall_design: Quantitative PCR (qPCR) expression profiling of plasma microRNA. Analysis of 23 previously reported miRNAs was performed in 24 T1D and 24 control individuals.

first: Diana

last: Swolin-Eide

first: Auste

last: Lycka

first: Gun

last: Forsander

first: Daniel

last: Novak

first: Johannes

last: Grillari

first: Andreas

middle: B

last: Diendorfer

first: Per

last: Magnusson


title: The SMARCA5–DMRT1 Pioneer Complex Establishes Epigenetic Priming to Direct Male Germline Development [RNA-Seq]

city: Davis

first: Satoshi

middle: H

last: Namekawa

address: One Shields Ave

institute: University of California, Davis

summary: Establishing cell type–specific chromatin landscapes is essential for cellular identity, yet how these landscapes are generated and maintained remains poorly understood. Here, we demonstrate that the chromatin remodeler SMARCA5 facilitates epigenetic priming required for retinoic acid–induced differentiation in the male germline. Germ cell–specific deletion of Smarca5 results in a complete loss of differentiating spermatogonia, phenocopying vitamin A–deficient mice lacking retinoic acid signaling. During the perinatal transition from prospermatogonia to undifferentiated spermatogonia, SMARCA5 is recruited to DMRT1-binding sites located at distal putative enhancers and promoters of germline genes. The SMARCA5–DMRT1 pioneer complex establishes chromatin accessibility at these loci, generating poised enhancers and promoters that serve as retinoic acid receptor (RAR)–binding sites. Thus, SMARCA5 licenses transcriptional responses to retinoic acid that enable spermatogenic differentiation. Our findings uncover a critical epigenetic priming mechanism that links pioneer factor activity to external signal responsiveness in the germline.

overall_design: RNA-seq analysis using Smarca5-ctrl and Smarca5 cKO P8 Aundiff and Smarca5-ctrl P8 Adiff; ATAC-seq analysis using Smarca5-ctrl P0 prospermatogonia, Smarca5-ctrl and Smarca5 cKO P8 Aundiff and Smarca5-ctrl P8 Adiff; CUT&Tag analysis on SMARCA5 using Smarca5-ctrl P0 prospermatogonia, Smarca5-ctrl P8 Aundiff, and on DMRT1 using Smarca5-ctrl and Smarca5 cKO P8 Aundiff.

first: Yuka

last: Kitamura

first: Yasuhisa

last: Munakata


title: Cell-autonomous control coupled with tissue context regulates the cessation of migration at the site of organ development

city: Berlin

first: Johanna

middle: Berenike

last: Kroll

address: Hannoversche Str.28

department: Berlin Institute for medical Systems Biology

institute: Max Delbrück Center for Molecular Medicine

summary: Organ development relies on interactions among different cell types that form three-dimensional structures to carry out specific tasks. This process often involves active migration of progenitor cells toward specific positions within the embryo, where the cells then become immotile and form stable connections among each other and with neighboring cell types. In this work, we study the process of motility loss using zebrafish primordial germ cells as an in vivo model. We show that changes in embryonic tissues as well as cell-autonomous events regulate germ cells’ behavior as they arrive at their target region. Importantly, we find that reduction in germ cell motility is correlated with the decay of RNA encoding for Dead end 1 (Dnd1), a conserved vertebrate RNA-binding protein that is essential for PGC migration. Indeed, decreasing or increasing the level of Dnd1 results in a premature or delayed stop to motility, respectively. These findings represent a novel RNA decay-based mechanism for timing the duration of cell migration in vivo.

overall_design: To generate spatially-resolved RNA-seq data for 24 hpf (hours post fertilization) old embryos, we cryosectioned samples, extracted RNA from the individual sections, and amplified and barcoded mRNA using the CEL-seq protocol (Hashimshony et al., Cell Reports, 2012) with a few modifications. Libraries were sequenced on Illumina NextSeq500.

first: Zahra

last: Labbaf

first: Moritz

last: Ophaus

first: Michal

last: Reichman-Fried

first: Jan Philipp

last: Junker

first: Erez

last: Raz


title: Selective targeting of type II tRNAs underlies SLFN14-mediated translational repression and its dysregulation by thrombocytopenia-linked mutations

city: Ithaca

first: Saori

last: Uematsu

state: New York

address: 526 Campus Road, Cornell University

institute: Cornell University

summary: Schlafens (SLFNs) are interferon-inducible proteins with emerging roles in RNA metabolism and antiviral defense. SLFN14 is a ribosome-associated endoribonuclease whose pathogenic variants cause an autosomal dominant form of inherited thrombocytopenia (IT), yet the molecular basis of this disorder has remained unclear. Here, we show that SLFN14 represses global protein synthesis and viral replication primarily through selective cleavage of type II tRNAs. Using cell lines engineered to express SLFN14 variants, we demonstrate that thrombocytopenia-linked mutations alter RNA substrate specificity, enhancing degradation of type II tRNAs while diminishing rRNA cleavage. This shift in substrate preference increases ribosome stalling at codons decoded by type II tRNAs, leading to global translational arrest, stress signalling, and ultimately cell death. The heightened degradation of type II tRNAs by SLFN14 mutants provides a mechanistic explanation for the translational defects and impaired megakaryocyte maturation observed in SLFN14-associated IT. Our findings suggest that type II tRNA targeting is a conserved regulatory strategy within the SLFN family and identify enhanced tRNA cleavage and the resulting impaired translation as the central pathogenic processes driven by thrombocytopenia-linked SLFN14 mutations.

overall_design: Ribo-seq from HEK293T cells expressing SLFN14 variants treated with Dox at different timepoints (0 and 24 h).


title: Investigating the fruitless-directed molecular and circuit architecture of courtship behavior in Drosophila melanogaster, Drosophila simulans and their hybrids

city: Tallahassee

first: Michelle

last: Arbeitman

address: 1115 W. Call Street

department: Department of Biomedical Sciences

institute: Florida State University

summary: Understanding the mechanisms driving behavioral evolution enhances our knowledge of speciation and how behavioral potentials are genetically encoded. Male courtship behaviors, which evolve rapidly, are critical for pre-mating isolation. To investigate the genetic basis of species-specific courtship, we generated hybrid males by crossing two Drosophila melanogaster strains with D. simulans Lhr males. This design allowed us to assess how genetic background and female species identity influence male courtship behavior. In both single-pair and mate-choice assays, hybrid males displayed behavioral plasticity, adjusting their courtship strategies based on the female species. The maternal D. melanogaster strain significantly shaped hybrid behavioral repertoires. To identify the neural correlates, we examined fruitless (fru)-expressing neurons in hybrids. Their projection patterns resembled those in D. melanogaster, indicating conserved circuit architecture. Using CUT&Tag, we identified FruM target genes in both species, revealing conserved core and species-specific FruM targets. Focusing on chemosensory receptors with D. melanogaster-specific FruM binding, we conducted a genetic screen in D. melanogaster, silencing neurons co-expressing fru P1 and specific receptor genes. Courtship preference assays identified three additional olfactory receptor neuron subtypes that modulate species-specific behavior. Using trans-Tango, we mapped second-order projection neurons of these subtypes, revealing their targets in higher-order brain centers. This study uncovers new molecular and neural mechanisms underlying the specification and evolution of courtship behavior, highlighting how genetic and sensory inputs shape species-specific behavioral outcomes.

sample_organism:
Drosophila melanogaster
Drosophila simulans

overall_design: CUT&Tag was performed to identify FruM target genes in male D. melanogaster and D.simulans brains. Males used in CUT&Tag were 15-24 hour old adults. The rat polyclonal anti-FruM antibody was used at a 1:50 dilution. The secondary rabbit anti-rat IgG was used at 1:100 dilution. For control libraries, the brains were only incubated in secondary antibody.

first: Colleen

middle: M

last: Palmateer


title: Repeat expansions in C9orf72 rewire the 3D chromatin landscape in ALS

city: Chapel Hill

first: Hyejung

last: Won

state: North Carolina

address: 116 Manning Dr

department: Genetics

institute: University of North Carolina at Chapel Hill

summary: Amyotrophic lateral sclerosis (ALS) is frequently driven by GGGGCC short tandem repeat (STR) expansions in C9orf72, yet the mechanisms by which these expansions lead to neurodegeneration remain incompletely understood. Here, we propose a novel mechanism involving higher-order chromatin architecture where C9orf72-STR expansions induce widespread, neuron-specific gains in chromatin loops that are closely linked to transcriptomic dysregulation in ALS. These ectopic loops colocalize with the genomic binding sites of C9orf72-STR RNAs and the architectural protein CTCF, supporting a model in which RNA-DNA interactions promote aberrant loop formation.

overall_design: Characterization of chromatin organizations in C9orf72 mediated ALS.

first: Tatiana

last: Ulloa Avila

first: Jingying

last: Wang


title: The SMARCA5–DMRT1 Pioneer Complex Establishes Epigenetic Priming to Direct Male Germline Development [ATAC-seq]


title: The SMARCA5–DMRT1 Pioneer Complex Establishes Epigenetic Priming to Direct Male Germline Development


title: Spatial Transcriptomics Reveals Local Subtype-Specific Identity and Signaling within Multifocal Small Intestinal Neuroendocrine Tumors

city: San Francisco

first: Ayushi

last: Agrawal

address: 1650 Owens St

institute: Gladstone Institutes

summary: Small intestinal neuroendocrine tumors (SI-NETs) frequently present as multifocal lesions, but the spatial and molecular mechanisms underlying their development and heterogeneity remain unclear. This study aimed to characterize the phenotypic subtypes of tumor cells across anatomical sites in multifocal SI-NETs and identify local microenvironmental factors influencing tumor development.

overall_design:
Spatial transcriptomics was performed on 72 tissue microarray cores derived from four patients with multifocal SI-NETs, covering tumor and non-tumor tissues from various anatomical layers and metastatic sites. Unsupervised clustering, over-representation analysis (ORA), and ligand-receptor (L-R) pair analysis were used to define tumor subtypes and associated signaling networks.

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

first: Akitada

last: Yogo

first: Naoki

last: Akanuma

first: Grace

last: Kim

first: Eric

last: Nakakura


title: Novel transcriptomic microglia taxonomy across mouse and human pathologies

city: Freiburg

first: Chintan

last: Chhatbar

address: Breisacher strasse 64

department: Department of Neuropathology

institute: Uniklinikum Freiburg

summary: Microglia are the key immune cells involved in virtually all diseases of the central nervous system (CNS). Numerous previous single-cell sequencing approaches to the study of microglia have revealed extensive diversity in this cell type in the human and mouse brain in different conditions. Importantly, a unifying, comprehensive, disease- and species spanning transcriptomic immune cell-focused atlas of the perturbed CNS, including true single-cell spatial transcriptomic information on neighborhood relationships, is lacking. Here, we generated a novel large-scale single-cell RNA-sequencing-derived taxonomy through the analysis of > 1 million CNS cells enriched for myeloid cells across > 30 different pathologies and conditions. Disease-associated human microglia, together with CNS-associated macrophages and monocytes, were differentiated iteratively into 27 superclusters and 192 clusters, showing substantial overlap with their counterparts in the corresponding murine pathologies. This top-down approach, in combination with targeted and genome-wide subcellular spatial transcriptomics, enabled us to examine and compare the spatial interactome of the identified superclusters and clusters within and across pathologies and species. Using cell-specific mutants, we further characterized the underlying transcriptional programs of activation-associated microglial clusters, identifying distinct dependencies on type I and II interferons and colony stimulating factor 1 receptor pathways. Our data provide new insights into the spatial dynamics of the endogenous CNS immune system during development, health and disease in human and mouse.

overall_design: CNS nuclei were isolated from frozen specimen, stained with anti-NeuN and anti-Olig2, FACS purified (DAPI+NeuN-Olig2-), and analyzed with single nucleus RNA-seq.

first: Marco

last: Prinz


title: SETD2 methyltransferase activity aids gene definition by promoting correct transcription initiation and efficient pre-mRNA 3’ cleavage [U2OS_T4PmNETseq]

city: Fukuoka

first: Takayuki

last: Nojima

state: Japan

address: Maidashi

department: Medical Sciences

country: Japan

institute: Kyushu University

summary: SETD2 is a key histone methyltransferase responsible for depositing histone H3 lysine 36 trimethylation (H3K36me3). Loss of its enzymatic activity has been implicated in various cancers, including renal cell carcinoma (RCC). In RCC, SETD2 mutations have been linked to defects in transcription termination, though the underlying mechanism remains unclear. Here, using nascent transcriptomics in SETD2 knockouts and RCC patient-derived cells, we show that SETD2 loss leads to delayed transcription termination, but only in a subset of genes. POINT-seq analysis reveals widespread transcriptional readthrough beyond canonical termination sites upon SETD2 loss/mutation, affecting 15–24% of genes. We demonstrate that defective termination following SETD2 loss/mutation is driven by a combination of increased cryptic transcription initiation and impaired 3′ pre-mRNA cleavage. In contrast, genes that maintain proper termination show significant downregulation. Additionally, we find that SETD2 ensures proper termination independently of alternative polyadenylation, underscoring the distinction between transcription termination and 3′ end processing. Our findings highlight that methyltransferase activity of SETD2 is a crucial regulator of transcriptional integrity, preventing cryptic initiation and promoting efficient 3′ end processing

overall_design: T4p-mNETseq profiling U2OS parental cells and SETD2 KO cells.

first: Magda

last: Kopczyńska

first: Chihiro

last: Nakayama

first: Agata

last: Stępień

first: Shoko

last: Ito

first: Koshi

last: Imami

middle: R

last: Gdula

first: Kinga

last: Kamieniarz-Gdula


title: Elucidating the fibrotic niche and cellular dynamics in crohn’s disease-associated fibrostenosis

first: Fang

last: Xie

address: Southern Medical University, No. 1838, North Guangzhou Avenue

department: Nanfang Hospital

institute: Southern Medical University

summary: Fibrostenosis is a common and debilitating complication of Crohn’s disease (CD), characterized by intestinal fibrosis, excessive extracellular matrix deposition, and luminal narrowing, for which effective antifibrotic therapies are lacking. To better understand the cellular landscape driving fibrosis, we performed high-resolution spatial transcriptomic analysis on ileal surgical samples from fibrostenotic CD patients, comparing strictured and non-strictured regions. This approach enabled us to delineate a distinct fibrotic niche enriched with stromal and immune cells, providing spatial context to the cellular interactions that underpin fibrotic remodeling in CD.

overall_design: Ileal surgical tissues from CD patients, including strictured and non-strictured regions, were used for spatial transcriptomic analysis.

first: Da

last: Zhang

first: Xinru

last: Zou

first: Mengjia

last: He

first: Anqi

last: Yu

first: Jintao

last: Shi

first: Ting

last: Ouyang

first: Xiaoxia

last: Lin

first: Hui

last: Yang

first: Lan

last: Bai

first: Shibo

last: Sun

first: Kai

first: Jiaolong

first: Fangting

last: Wu

first: Wenting

first: Jingyan

last: Li


title: SETD2 methyltransferase activity aids gene definition by promoting correct transcription initiation and efficient pre-mRNA 3’ cleavage [U2OS_POINTseq]

first: Takayuki

last: Nojima

overall_design: POINTseq profiling U2OS parental cells and SETD2 KO cells.

first: Magda

last: Kopczyńska

first: Chihiro

last: Nakayama

first: Agata

last: Stępień

first: Shoko

last: Ito

first: Koshi

first: Michal

first: Kinga


title: SETD2 methyltransferase activity aids gene definition by promoting correct transcription initiation and efficient pre-mRNA 3’ cleavage [U2OS_3mRNAseq]

overall_design: 3mRNAseq profiling U2OS parental cells and SETD2 KO cells.

last: Imami

last: Gdula

last: Kamieniarz-Gdula


title: SETD2 methyltransferase activity aids gene definition by promoting correct transcription initiation and efficient pre-mRNA 3’ cleavage [RCC_3mRNAseq]

overall_design: 3mRNAseq performed for ACHN and A498 cells.


title: Evaluating MiRomics and Integrative Proteomics for Identification of Plasma miRNAs as Potential Pharmacodynamic Biomarkers of IFNβ-1a Biologics

city: Silver Spring

first: Lakshmi Manasa

middle: Sakuntala

last: Chekka

state: Maryland

address: 10903 New Hampshire Avenue

department: DARS, CDER

institute: U.S. FDA

summary: We used miRNA-sequencing to identify candidate plasma pharmacodynamic (PD) biomarkers of interferon beta-1a (IFNβ-1a) biologics and to explore miRNA-mRNA targeted protein relationships and networks to support identified miRNA candidates. Plasma samples from 36 healthy subjects from a placebo-controlled randomized single dose clinical study with IFNβ-1a and pegIFNβ-1a were used. Mature miRNAs were measured using an in-house miRNA-seq workflow at baseline, at 9 timepoints over 6 days in IFNβ-1a group (n=11[30µg]), and at 11 timepoints over 13 days in pegIFNβ-1a group (n=11[125µg]) and placebo-specific groups (n=6 each). A miRNA was only considered expressed if it had a read count ≥10 in more than 50% of the samples across all treatment groups. Linear mixed-effects models (lmer) regressing miRNA changes with treatment, time and their interaction were used to identify differentially expressed miRNAs, which were further prioritized based on magnitude of response and biological relevance using in silico predicted miRNA-protein targets and regulatory network analyses. Ten and 13 miRNAs were impacted by IFNβ-1a and pegIFNβ-1a, respectively (lmer FDR-corrected p- value <0.1), compared to placebo, of which miR.223.3p and miR.21.5p were prioritized as candidate PD biomarkers for both products. Systems level analysis of integrated proteomics data highlighted miRNA protein targets for both miR.223.3p and miR.21.5p that were linked to previously reported top proteomic response proteins and predicted regulatory networks including IFN beta. Using miRNA-sequencing and linking candidates to predicted target proteins and regulatory networks, results suggest miR.223.3p and miR.21.5p as potential novel PD biomarkers of IFNβ-1a biologics for further investigation.

overall_design: To identify potential PD miRNA biomarkers of IFNβ-1a biologics, we collected plasma samples from healthy participants randomized into three treatment groups (IFNβ-1a, pegIFNβ-1a, and placebo). These samples were obtained at time zero (baseline), at  9 timepoints over 6 days following administration of IFNβ-1a, and at 11 timepoints over 13 days following administration of pegIFNβ-1a and placebo. We then performed miRNA sequencing (next generation sequencing) on these samples. Uisng the normalized sequencing data, we conducted miRNA differential expression analysis to identify differentially expressed miRNAs by treatment from baseline.

first: Mai

last: Mehanna

first: Paula

middle: L

last: Hyland


title: Extensive enhancer crosstalk governs the induction of PPARG2 expression during adipogenic lineage commitment [Chip-Seq]

city: Odense M

first: Susanne

last: Mandrup

address: Campusvej 55

department: Biochemistry and Molecular Biology

country: Denmark

institute: University of Southern Denmark

summary: Transcriptional master regulators drive cell fate transitions. Peroxisome proliferator- activated receptor γ (PPARγ) is the master regulator of adipogenesis, and its expression must therefore be tightly regulated and efficiently induced in response to adipogenic cues. Here we show that the PPARG locus is primed for activation in the mesenchymal stem cell state by a highly connected enhancer (HICE) community. By systematically deleting 9 individual enhancers, spanning upstream, promoter-proximal, and downstream super-enhancer constituents, we demonstrate elaborate enhancer crosstalk in cis involving stabilization of C/EBPβ recruitment prior to chromatin remodeling. We show that the super-enhancer constituent E+102 plays a dual role in cis crosstalk and feedback activation and is obligate for activation of PPARG expresison. Non-coding genetic variants associated with cardiometabolic traits and predicted to regulate PPARG expression map E+102 and other essential enhancers in the community, thereby supporting the importance of these enhancers in human physiology and disease.

overall_design: Genome-wide profilling of transcription factors, co-factors and histone marks using ChIP-seq during adipocyte differentation of human mesenchymal stem cell, and human mesenchymal stem cell, and human mesenchymal stem cells with inducible expression of Cas9, and selected enhancer individual deletions.

first: Anna

last: Cetnarowska

first: Mette

last: Hyldahl

first: Marcus

last: Nygård

first: Bo

last: Vagner Hansen

first: Laura

last: Holm

first: Kaja

last: Madsen

first: Maria

last: Stahl Madsen

first: Vallari

last: Shukla

first: Esra

last: Durmaz Mitchell

first: Alexander

last: Rauch

first: Jesper

last: Grud Skat Madsen

first: Melina

last: Claussnitzer


title: LIVER EXERCISE FACTOR REVERSES AGING- AND ALZHEIMER’S-RELATED MEMORY LOSS BY TARGETING BRAIN VASCULATURE

first: Yasuhiro

last: Fuseya

address: 513 Parnassus Ave Bldg S1364, Medical Science

department: Anatomy

institute: University of California, San Franc

summary: Blood factors transfer the benefits of exercise to the aged brain, independent of physical activity. Here we show that liver-derived exercise factor glycosylphosphatidylinositol (GPI)-specific phospholipase D1 (GPLD1), a GPI-degrading enzyme, reverses aging- and Alzheimer’s-related memory loss by targeting brain vasculature. GPLD1 has potential to cleave over 100 putative GPI-anchored proteins, necessitating identification of downstream targets that mediate cognitive rejuvenation for translational application. We identified GPI-anchored tissue-nonspecific alkaline phosphatase (TNAP) on brain vasculature as a GPLD1 substrate. Mimicking the age-related increase in cerebrovascular TNAP impaired blood-brain transport and cognition in young mice and mitigated Gpld1-induced cognitive benefits in aged mice. Inhibiting TNAP recapitulated benefits of GPLD1 in old age, restoring youthful hippocampal transcriptional signatures and rescuing cognition. In an Alzheimer’s disease model, increasing GPLD1, or inhibiting TNAP, ameliorated Ab pathology and improved cognitive deficits. We thus identify brain vasculature as a mediator of the cognitive benefits of a liver-to-brain exercise axis.

overall_design: Hippocampal brain endothelial cells (BECs) isolated from young and aged control mice, aged mice with liver expression of GPLD1 and aged mice treated with a TNAP inhibitor (TNAPi) using fluorescence activated cell sorting (FACS). Pool of 8 mice/group.

first: Fuseya

last: Yasuhiro

first: Bieri

last: Gregor

first: Pratt

last: Karishma


title: The SMARCA5–DMRT1 Pioneer Complex Establishes Epigenetic Priming to Direct Male Germline Development [CUT&Tag]

first: Satoshi

last: Namekawa

first: Yuka

last: Kitamura

first: Yasuhisa

last: Munakata


title: Transcriptional and cellular maturation of the chick spinal cord in the context of distinct neuromuscular circuits

city: Basel

first: Patrick

last: Tschopp

state: Basel-Stadt

address: Vesalgasse 1

department: Umweltwissenschaften

institute: Universität Basel

summary: The chicken spinal cord is a classic model system to study the early specification of neuronal cell types along its anterior-posterior axis. Here, we follow the ensuing maturation dynamics at limb levels with single cell resolution and contrast neuronal populations innervating appendages of distinct form and function. We use gene co-expression modules to identify rare cell populations with specific biological functions, and show that appendages with different motor outputs – wings and legs – rely on largely similar spinal cord cell type repertoires. Challenging the system with experimental alterations to the peripheral limb musculature reveals limited transcriptional changes, but spatially restricted plasticity in spinal cord motor neuron numbers. Collectively, our results provide a resource to investigate the molecular and cellular basis of neuronal maturation in the avian spinal cord and highlight the plastic nature of embryonic cells to adapt to changes in the limb periphery at both developmental and evolutionary timescales.

overall_design: 10X scRNA-seq of Day 5 (brachial), Day 7 (brachial), and Day 10 (brachial, lumbar, and brachial with induced polydactyly) chick spinal cord.

first: Fabio

last: Sacher

first: Bianka

last: Berki

first: Antoine

last: Fages

first: Maëva

last: Luxey


title: SETD2 methyltransferase activity aids gene definition by promoting correct transcription initiation and efficient pre-mRNA 3’ cleavage [U2OS_POINT5]

first: Takayuki

last: Nojima

overall_design: POINT5 profiling U2OS parental cells and SETD2 KO cells.

first: Magda

last: Kopczyńska

first: Chihiro

last: Nakayama

first: Agata

last: Stępień

first: Shoko

last: Ito

first: Koshi

last: Imami

first: Michal

last: Gdula

first: Kinga

last: Kamieniarz-Gdula


title: Deletion 9p drives B-cell leukemia by the concomitant loss of Pax5 and Cd72

city: Salamanca

first: Isidro

last: Sánchez García

address: Campus Miguel de Unamuno

department: Centro de inv. del Cáncer (CIC).

institute: Inst. de Inv. Biomédica de Salamanca (IBSAL). H. U. de Salamanca. Inst. de Biología Molecular y Celular del Cáncer (IBMCC).

summary:
We used microarrays to analyze gene expression changes in preleukemic cells from Cd72+/-;Pax5+/-,  Pax5+/- and WT mice.
The oncogenic role of 9p deletion in B-ALL has remained elusive since its discovery more than 40 years ago. We combined genomic analysis and functional models to show that the co-deletion of two contiguous genes, Pax5 and Cd72, confers gain in malignancy.

overall_design: The total RNA was first isolated using TRIzol (Life Technologies), and then it was subjected to purification with the RNeasy Mini Kit (Qiagen) using also the On-Column DNase treatment option. Quality and quantification of RNA samples were determined by electrophoresis.  Determination of the expression of the different genes in the RNA samples was performed using Affymetrix Mouse Gene 1.0 ST arrays. All bioinformatic analyses of the array data were performed using Applied Biosystems Transcriptome Analysis Console (TAC) software (v4.0.3).  This software was utilized to identify differentially expressed genes by comparing the expression levels between different conditions. To determine statistical significance, a threshold of p-value < 0.05 and |FC| > 2 was applied.

first: Belén

last: Ruiz-Corzo

first: Ana

last: Casado-García

first: Ninad

last: Oak

first: Paula

last: Somoza-Cotillas

first: Andrea

last: LoÌpez-AÌlvarez de Neyra

first: Jorge

last: Martínez-Cano

first: Elena

middle: G

last: Sánchez

first: Oscar

last: Blanco

first: Diego

last: Alonso-López

first: Javier

last: De Las Rivas

first: Susana

last: Riesco

first: Pablo

last: Prieto-Matos

first: Francisco

last: Criado

first: María

last: Cenador

first: Manuel

last: Ramírez-Orellana

first: César

last: Cobaleda

first: Carolina

last: Vicente-Dueñas

first: Kim

middle: E

last: Nichols

last: Sánchez-García


title: Alzheimer’s Disease-Related Mutations in APP Influence Colorectal Tumor Formation in a Sex-Dependent Manner

city: Grand Forks

first: Takako

last: Ishida-Takaku

state: ND

address: 1301 North Columbia Road

institute: University of North Dakota

summary: Cancer and Alzheimer’s disease (AD) are major age-associated diseases. Numerous studies have indicated common mechanisms and biological interactions between these conditions. Furthermore, epidemiological data demonstrate negative correlations between several cancers and AD. Despite this, few studies have explored their mutual influence on pathological outcomes. In this study, we utilized a human APP mutant knock-in AD mouse model to investigate how familial mutations influence colorectal cancer development. Using a colitis-associated colorectal cancer model, APP mutations promoted colon tumor formation in male mice but inhibited it in female mice. Correspondingly, inflammatory changes were reduced in the colons of female mice. Transcriptome analysis revealed differential expression, particularly in the enrichment of neuronal marker, steroid hormone, and immune cell signaling pathways. Additionally, distinct macrophage subtypes and neuronal profiles were observed in the colons of male and female APP mutant mice. These findings provide the first elucidation of the sex-dependent effects of APP mutations on colon cancer formation.

overall_design: To understand why AOM/DSS treatment resulted in differential phenotypic outcomes in an APP mutant background at the molecular level, we conducted RNA-seq using colon cancer tissues, adjacent normal colon tissues, or normal colon tissues when tumors were not formed

first: Mona

last: Sohrabi

first: Sathiya

middle: P

last: Chandrasekaran

first: Heidi

last: Pecoraro

first: Colin

last: Combs