title: NSD2 targeting reverses lineage plasticity and drug resistance in advanced prostate cancer [bulk RNA-seq]

city: New York

first: Jia

middle: Jessie

last: Li

state: NY

department: Medicine

institute: Columbia University Irving Medical Center

summary: Treatment with highly potent androgen receptor signaling inhibitors (ARSIs), such as enzalutamide and abiraterone promotes lineage plasticity in metastatic castration-resistant prostate cancer (mCRPC), which results in intra-tumor heterogeneity and emergence of mCRPC subtypes. The histological transformation from adenocarcinoma to aggressive neuroendocrine prostate cancer (CRPC-NE) has been associated with a loss of dependency on lineage-survival signals, leading to targeted drug resistance. Epigenomic reprogramming might be a fundamental driver of lineage plasticity. To determine CRPC-NE vulnerabilities, we have performed image-based screening using a small library of antibodies targeting different histone marks on CRPC-NE organoids derived from genetically engineered mice. We find that the histone mark H3K36me2 and the histone methyltransferase NSD2 (also known as MMSET/WHSC1) play important roles in maintaining the state of CRPC-NE. Knockout of NSD2 or ablation of H3K36me2 using H3.3K36M oncohistone reverted CRPC-NE to CRPC-AR-like phenotype. Simultaneous profiling of the transcriptome and epigenome from single cells collected from control (sgCtrl) and NSD2 knockout (sgNSD2) organoids, or from empty vector (EV) and H3.3K36M transduced organoids confirms the lineage reversal of treatment-resistant cells and maps the re-establishment of canonical AR signaling dependency. Moreover, H3K36me2 or NSD2 depleted mouse and human CRPC-NE organoids responded to enzalutamide treatment in vitro and in vivo, suggesting a restoration of ARSI sensitivity. Most importantly, a small molecule inhibitor of NSD2, which is similar to KTX-1001 (NCT05651932), in combination with enzalutamide leads to growth suppression or apoptosis of mouse and human CRPC-NE organoids, organoids of other CRPC subtypes and xenografts in vitro and in vivo. In conclusion, inhibition of NSD2 reverses lineage plasticity and reverts the state of treatment-resistant cells back into an ARSI-sensitive state. Thus, we suggest that the combination of NSD2 inhibition with AR inhibition may represent a novel therapeutic approach for patients with CRPC-NE and other CRPC subtypes.

overall_design: The mouse NE tumors derived from NPp53 (Nkx3.1CreERT2/+; Ptenflox/flox; TrpP53flox/flox; Rosa26-EYFP) mice were cultured as organoids. To detect intercellular heterogeneity within organoids, I used single-cell RNA sequencing (scRNA-seq) to analyze lineage diversity of five mouse NE tumor organoids on passage 2. I also used single-nucleus ATAC sequencing (snATAC-seq) to profile the chromatin accessibilities of NPPO-1 organoids on passage 8, a multilineage NE tumor organoid line. The non-NE tumor cells from NPPO-1 organoids on passage 3 were isolated by fluorescence-activated cell sorting (FACS) according to a higher side scatter (SSC) value in non-NE compared to NE tumor cells. Some isolated non-NE tumor cells were analyzed by scRNA-seq to detect any contamination by NE tumor cells. Other isolated non-NE tumor cells were transduced with H2B-RFP lentivirus without antibiotic selection. H2B-RFP labeled tumor cells were either cultured alone or co-cultured with NE tumor cells to induce non-NE to NE conversion. To trace cell lineage of H2B-RFP-labeled cells, organoids from either monoculture or co-culture on passage 4 were analyzed by scRNA-seq (gene name: addgene26001). To quantitively compare the histone mark expression between NE and non-NE cells, epigenomic profiling using Cut&Tag was performed on 4 NE and 4 non-NE organoids derived from the same NPp53 background. To accurately quantify gene regulation, bulk RNA-seq was performed alongside epigenomic profiling on the organoids. To study the effects of NSD2 on NE lineage maintenance, we knockout of NSD2 on two mouse NE tumor organoids using CRISPR/Cas9. After antibiotic selection for 14 days, positive-transfected cells were isolated by FACS sorting according to the presence of RFP reporter signal. We used multiome single-nucleus ATAC and RNA sequencing (multiome snATAC + snRNA-seq) to analyze changes of cell lineage and state in sgNSD2 transfected NE tumor organoids in compared with sgControl (sgCtrl) organoids. To provide direct evidence of H3K36me2 histone modification in modulating NE lineage, mouse NE tumor organoids were transduced with lentivirus carrying the gene encoding the H3.3K36M oncohistone that can cause significant reduction in global levels of H3K36me2. After antibiotic selection for 14 days, cells positively transfected with H3.3K36M or empty vector (EV) control were analyzed using multiome snATAC + snRNA-seq to detect changes in cell lineage and state. In parallel, quantitative epigenomic profiling using Cut&Tag was performed on sgCtl and sgNSD2, or EV and H3.3K36M organoids to quantify global epigenomic changes. We have synthesized a small molecular inhibitor of NSD2 (NSD2i), which is similar to KTX-1001 (NCT05651932) in phase 1 clinical trial in patients with relapsed and refractory multiple myeloma. To evaluate if NSD2i recapitulates the effects of genetic interventions in reversal of lineage plasticity and restoring androgen receptor signaling inhibitor (ARSI) sensitivity, 1 µM NSD2i or DMSO was applied every other day on NPPO-1NE organoids cultured in a medium without DHT for 12 days. Organoids treated with either DMSO or 1 µM NSD2i were then collected for multiome snATAC + snRNA-seq analysis or Cut&Tag epigenomic profiling.

middle: J

first: Kristjan

middle: H

last: Gretarsson

first: Michael

middle: M

last: Shen


title: NSD2 targeting reverses lineage plasticity and drug resistance in advanced prostate cancer [Cut&Tag]

status: Public on Dec 22 2025

country: USA

type: Genome binding/occupancy profiling by high throughput sequencing

overall_design: The mouse NE tumors derived from NPp53 (Nkx3.1CreERT2/+; Ptenflox/flox; TrpP53flox/flox; Rosa26-EYFP) mice were cultured as organoids. To detect intercellular heterogeneity within organoids, I used single-cell RNA sequencing (scRNA-seq) to analyze lineage diversity of five mouse NE tumor organoids on passage 2. I also used single-nucleus ATAC sequencing (snATAC-seq) to profile the chromatin accessibilities of NPPO-1 organoids on passage 8, a multilineage NE tumor organoid line. The non-NE tumor cells from NPPO-1 organoids on passage 3 were isolated by fluorescence-activated cell sorting (FACS) according to a higher side scatter (SSC) value in non-NE compared to NE tumor cells. Some isolated non-NE tumor cells were analyzed by scRNA-seq to detect any contamination by NE tumor cells. Other isolated non-NE tumor cells were transduced with H2B-RFP lentivirus without antibiotic selection. H2B-RFP labeled tumor cells were either cultured alone or co-cultured with NE tumor cells to induce non-NE to NE conversion. To trace cell lineage of H2B-RFP-labeled cells, organoids from either monoculture or co-culture on passage 4 were analyzed by scRNA-seq (gene name: addgene26001). To quantitively compare the histone mark expression between NE and non-NE cells, epigenomic profiling using Cut&Tag was performed on 4 NE and 4 non-NE organoids derived from the same NPp53 background. To accurately quantify gene regulation, bulk RNA-seq was performed alongside epigenomic profiling on the organoids. To study the effects of NSD2 on NE lineage maintenance, we knockout of NSD2 on two mouse NE tumor organoids using CRISPR/Cas9. After antibiotic selection for 14 days, positive-transduced cells were isolated by FACS sorting according to the presence of RFP reporter signal. We used multiome single-nucleus ATAC and RNA sequencing (multiome snATAC + snRNA-seq) to analyze changes of cell lineage and state in sgNSD2 transfected NE tumor organoids in compared with sgControl (sgCtrl) organoids. To provide direct evidence of H3K36me2 histone modification in modulating NE lineage, mouse NE tumor organoids were transduced with lentivirus carrying the gene encoding the H3.3K36M oncohistone that can cause significant reduction in global levels of H3K36me2. After antibiotic selection for 14 days, cells positively transduced with H3.3K36M or empty vector (EV) control were analyzed using multiome snATAC + snRNA-seq to detect changes in cell lineage and state. In parallel, quantitative epigenomic profiling using Cut&Tag was performed on sgCtl and sgNSD2, or EV and H3.3K36M organoids to quantify global epigenomic changes. We have synthesized a small molecular inhibitor of NSD2 (NSD2i), which is similar to KTX-1001 (NCT05651932) in phase 1 clinical trial in patients with relapsed and refractory multiple myeloma. To evaluate if NSD2i recapitulates the effects of genetic interventions in reversal of lineage plasticity and restoring androgen receptor signaling inhibitor (ARSI) sensitivity, 1 µM NSD2i or DMSO was applied every other day on NPPO-1NE organoids cultured in a medium without DHT for 12 days. Organoids treated with either DMSO or 1 µM NSD2i were then collected for multiome snATAC + snRNA-seq analysis or Cut&Tag epigenomic profiling.


title: Combined epigenetic therapy with FACT and BET inhibitors remodels chromatin and disrupts oncogenic transcription in Diffuse Midline Glioma [Xenium]

city: Kensington

first: Holly

last: Holliday

address: Lowy Cancer Research Centre

country: Australia

institute: Children's Cancer Institute

summary: Aberrant epigenetic regulation is a hallmark of Diffuse Midline Glioma (DMG), an incurable pediatric brain tumor. The H3K27M driver histone mutation leads to transcriptional dysregulation, indicating that targeting the epigenome and transcription may be key therapeutic strategies against this highly aggressive cancer. One such target is the Facilitates Chromatin Transcription (FACT) histone chaperone. We found FACT to be enriched at developmental gene promoters, coinciding with regions of open chromatin and binding motifs of core DMG regulatory transcription factors. Furthermore, FACT interacted and co-localized with the Bromodomain and Extra-Terminal Domain (BET) protein BRD4 at promoters and enhancers, suggesting functional cooperation between FACT and BRD4 in DMG. In vitro, a combinatorial therapeutic approach using the FACT inhibitor CBL0137, coupled with BET inhibition revealed potent and synergistic cytotoxicity across a range of DMG cultures. These results were recapitulated in vivo, significantly extending survival in three independent orthotopic PDX models of DMG. Mechanistically, we show that CBL0137 treatment decreased chromatin accessibility, synergizing with BET inhibition to cause broad transcriptional collapse, silencing several key oncogenes including MYC, PDGFRA, MDM4 and SOX2, as well as causing alterations to the splicing landscape. Notably, this combination also elicited immune-related effects, including activation of the interferon response and antigen presentation mechanisms in DMG cells and induction of an activated state in macrophages and T cells, as demonstrated in an immunocompetent setting with spatial transcriptomics. Altogether, our data highlights the therapeutic promise of simultaneously targeting FACT and BET proteins in DMG, offering a dual tumor-intrinsic and immune-mediated strategy for combating this devastating pediatric brain tumor.

overall_design: 5-7 weeks old female C57BL6 mice were intracranially injected into the brainstem with IUE 24B7 cells that harbor H3.3K27M mutation generated in (doi: 10.1093/neuonc/noz197 and 10.1093/noajnl/vdac079). The timing of treatment initiation was 18 days post intracranial surgery. The drug treatments were as follows: CBL0137 - dissolved in 5% dextrose, CBL0137 was administered intravenously at a dosage of 40 mg/kg/day. The treatment was given once every four days for a total of three weeks. JQ1 - dissolved in DMSO and subsequently in a solution containing 0.9% saline with 5% PEG400 and 5% TWEEN-80, JQ1 was administered at a dosage of 50 mg/kg/day intraperitoneally. The treatment was given five days a week over three weeks. Brain samples were collected on the final day of treatment and fixed in a 10% formalin neutral buffered solution (Sigma-Aldrich). The fixed samples were then embedded in paraffin wax, and 5 µm sections were cut and mounted onto Xenium slides, with two brains per slide. Xenium Prime In Situ (10x Genomics) was performed according to the manufacturer’s instructions using the 5000 pre-designed gene panel and cell segmentation staining. Briefly, priming oligos were hybridized to the target RNA, followed by site-specific cleavage using RNase treatment to release the RNA strand from the hybridized oligo. The 5000-gene panel was then added to the sections to hybridize with the RNA, followed by ligation to generate circular DNA probes, which were subsequently amplified via rolling circle amplification. After a series of ethanol washes, the tissue was blocked and stained with the cell segmentation stain and DAPI. Slides were then loaded into the Xenium Analyzer for imaging and decoding. Spatial Transcriptomics Services for this study were provided by the Garvan Genomics Platform at the Garvan Institute of Medical Research.


title: IFNγ-induced memory in human macrophages is not sustained by epigenetic changes but the durability of the cytokine itself [ATAC-Seq]

city: Los Angeles

first: Aleksandr

last: Gorin

state: CA

address: 611 Charles E Young Dr E 570C Boyer Hall

institute: UCLA

summary: Macrophages, as key sentinel cells of the innate immune system, can retain memory of prior stimulus exposure. Interferon gamma (IFNγ) plays a central role in maintaining trained immunity in vivo and can induce potent memory in macrophages. Such memory is associated with the formation of de novo enhancers that alter gene expression responses to subsequent stimuli. However, how such enhancers are maintained after cytokine exposure remains unclear. We report that durable IFNγ-induced enhancers can last for days after cytokine washout, yet the underlying persistence mechanism is not cell-intrinsic. IFNγ-treated macrophages continue to exhibit JAK/STAT signaling days after cytokine removal. Blocking IFNγ signaling with a JAK inhibitor or anti-IFNγ neutralizing antibodies after cytokine removal is sufficient to reverse IFNγ-induced enhancers and erase the potentiated state of the treated macrophages. Our findings suggest that epigenetic changes in macrophages do not inherently encode innate immune memory or a “potentiated” macrophage state, but that rather epigenetic changes are themselves dependent on ongoing cytokine signaling. These findings suggest new possibilities for pharmacologic interventions to reverse aberrantly trained immune states associated with pathology.

overall_design: ATACseq was performed to identify chromatin opening induced by interferon gamma in human macrophages and assay the necessity of continued JAK/STAT signaling on chromatin accessibility after interferon withdrawal.

first: Vyas

last: Koduvayur

first: Noa

last: Harriott

first: Quen

last: Cheng

first: Alexander

last: Hoffmann


title: Epigenetic Signatures of Brain Aging by Single Nucleus Methylation Sequencing [NACB-9mo-m-012]

status: Public on Dec 23 2025

city: La Jolla

first: Joseph

middle: R

last: Ecker

address: 10010 North Torrey Pines Road

department: Genomic Analysis Laboratory

institute: HHMI-Salk-Institute

type: Methylation profiling by high throughput sequencing

summary: Illustrating aging effects on the epigenomic features of brain is critical to understand brain functional decline and neurodegeneration. Single nucleus methylation sequencing was applied to brain regions of mouse at the age of 2, 9 and 18 months. We use these data to identify the epigenomic changes with aging in mouse brains.

overall_design:
Apply snmC-seq3 to mouse brain regions to identify aging effects on epigenomics

Processed data for these samples are available at:

https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE313770

first: Q

last: Zeng

first: Wei

last: Tian

first: Anna

last: Bartlett

first: J

last: Nery

first: Rosa

last: Gomez Castanon

first: Julia

middle: K

last: Osteen

first: Jordan

last: Altshul

first: N

middle: D

last: Johnson

first: M

last: Kenworthy

first: C

last: Valadon

first: Huaming

last: Chen

last: Behrens


title: E2F1 methylation by SETD6 disrupts BRD4-E2F1 binding and modulates gene regulation in prostate cancer cells

city: Stuttgart

first: Albert

last: Jeltsch

address: Allmandring 31

department: Institute of Biochemistry and Technical Biochemistry

country: Germany

institute: University Stuttgart

type: Expression profiling by high throughput sequencing

summary: The SETD6 protein lysine methyltransferase has a role in the regulation of various cellular processes including cancer initiation and progression. It methylates several cellular proteins including K117 of the E2F1 transcription factor, but the functional consequences of this methylation event are unknown. In this study, the role of SETD6 mediated E2F1 methylation on the progression of prostate cancer was investigated. We identified distinct sets of genes that are bound and upregulated by methylated and unmethylated E2F1. Genes upregulated by methylated E2F1 lead to the negative regulation of cell migration,  while genes upregulated by unmethylated E2F1 have roles in cell proliferation and apoptosis. BRD4 is known to bind E2F1 acetylated at K117 and K120. Here, we demonstrate that methylation of E2F1 at K117 prevents E2F1-BRD4 binding of in vitro and in cells establishing the molecular mechanism of the differential gene regulation by methylated and unmethylated E2F1 that depends on SETD6 activity.

overall_design: The DU145 human prostate cancer cell line was used in experiment.  SETD6 knock-out cells was generated by CRISPR/Cas9 system. Cells had stable expression of Flag-E2F1 WT and E2F1 K117R.

first: Gizem

middle: T

last: Ulu

first: Margarita

last: Kublanovsky

first: Michal

last: Feldman

first: Sara

last: Weirich

first: Dan

last: Levy


title: Effect of pancreatic cancer derived extracellular vesicles on bone marrow drived macrophages

city: Washington

first: Amrita

last: Cheema

state: DC

address: 3900 Reservoir Road NW

department: Oncology

institute: Georgetown University Medical Centre

type: Non-coding RNA profiling by high throughput sequencing

summary: Our results show that PaCa cell derived EVs were taken up by the macrophage in a time-dependent manner that resulted in an immunosuppressive phenotype characterized by higher expression of CD206 and PD-L1 and higher secretion of immunosuppressive cytokines including TGF-beta, IL-10 and GM-CSF.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: EVs were isolated from the conditioned medium that was used to culture one established pancreatic cancer cell (PANC-1), one PDX cell line (PPCL68) and a normal (non-tumorigenic) established pancreatic epithelial cell line (hTERT-HPNE), using size exclusion chromatography. Macrophages were generated from the bone marrow cells of C57/BL6 mice cultured in vitro using macrophage colony stimulating factor (M-CSF). The macrophages were seeded into 24 well plates and co-cultured with isolated PaCa or normal cell derived EVs at the concentration of 10 ug/mL EVs for 48h.

first: Baldev

last: Singh

first: Pankaj

last: Gaur

first: Pritha

last: Bose

first: Yanjun

last: Zhang

first: Yaoxiang

last: Li

first: Zihao

first: Jeyalakshmi

last: Kandhavelu

first: William

last: Klotzbier

first: Meth

last: Jayatilake

first: Shivani

last: Bansal

first: Mohd

last: Farhan

first: Sunain

last: Deol

first: Partha

middle: P

last: Banerjee

first: Keith

last: Unger

first: Seema

last: Gupta

first: Vivek

last: Verma


title: Single cell and isoform-specific translational profiling of the mouse brain [ribostamp]

first: Brian

last: Yee

address: 9500 Gilman Dr

department: Health

institute: UCSD

summary: The brain displays the richest repertoire of post-transcriptional mechanisms regulating mRNA translation1–12. Among these, alternative isoform expression has been shown to drive cell type specificity and, when disrupted, is strongly linked to neurological disorders13–18. However, genome-wide measurements of mRNA translation with isoform-sensitivity at single-cell resolution have not been achieved. To address this, we deployed Ribo-STAMP (Surveying Ribosomal Targets by APOBEC-Mediated Profiling) coupled with short- and long-read single-cell RNA-sequencing in the brain19. We generated the first isoform-sensitive single cell translatomes of the mouse hippocampus, discovering 797 alternative isoforms across 325 genes, with CA1 exhibiting the most differentially expressed and CA3 the most differentially translated isoforms. We defined high and low translational states in CA1 and CA3 neurons, with ribosomal, metabolic, and synaptic genes enriched in high states. Surprisingly, we found that CA3 neurons exhibited higher basal translation compared to CA1, as confirmed by metabolic labeling of newly synthesized proteins and immunohistochemistry of mRNA translation factors. This easily adoptable platform will expand our understanding of how isoform-specific translation drives brain physiology and disease.

overall_design: To optimize Ribo-STAMP for expression in brain cells, we compared RPL10a, RPL22 and RPS2 NT (deposited at GSE275091) Ribo-STAMP constructs in mouse cortical neurons.

first: Samantha

middle: L

last: Sison

first: Eugene

middle: W

last: Yeo


title: E2F1 methylation by SETD6 disrupts BRD4-E2F1 binding and modulates gene regulation in prostate cancer cells [ChIP-seq]

first: Albert

last: Jeltsch

overall_design: The DU145 human prostate cancer cell line was used in the experiment. SETD6 knock-out cells were generated using the CRISPR/Cas9 system. Flag-E2F1 was stably integrated and constitutively expressed.

first: Gizem

last: Ulu

first: Margarita

last: Kublanovsky

first: Michal

last: Feldman

first: Sara

last: Weirich

first: Dan

last: Levy


title: Single cell and isoform-specific translational profiling of the mouse brain [Bulk RNA-seq]

overall_design: To determine if RPS2-STAMP accurately reports ribosome occupancy on mRNAs in neurons, we benchmarked it against enhanced cross-linking and immunoprecipitation (eCLIP) of ribosomal proteins.

first: Sison

last: Samantha

first: Yeo

last: Eugene


title: Extracellular matrix protein 1 shapes metabolic and spatial dynamics of the kidney fibrotic microenvironment

city: Pittsburgh

first: Shuchang

last: Liu

state: PA

address: 203 Lothrop Street

institute: University of Pittsburgh

summary: The fibrotic kidney microenvironment is a battlefield shaped by cellular crosstalk, ECM remodeling, metabolic shifts, and spatial heterogeneity, all fueling fibrosis. In late-stage kidney disease, myofibroblast-produced ECM dominates organ function by altering bioenergetics and metabolite signaling. However, the role of early-activated matrix proteins remains unclear. This study identifies four expression patterns of core matrisome proteins, with Ecm1 emerging as a key early player in kidney remodeling. Ecm1 knockout in mice leads to spontaneous kidney fibrosis and early death, yet serum Ecm1 levels rise in CKD patients. Targeting Ecm1 via adeno-associated virus-mediated knockdown or fibroblast-specific deletion substantially reduces kidney fibrosis. Mechanistically, proteomics and spatial transcriptomics show that fibroblast-Ecm1 ablation disrupts Hippo-Yap signaling via integrin α2β1 and RhoC in tubules, enhancing OXPHOS in tubular cells and promoting repair, as validated across pharmacological, in vivo, ex vivo, and in vitro models. Notably, diminished Hippo-Yap in fibroblasts silenced their aberrant activation without crippling their own OXPHOS. This selective ECM-mitochondrial crosstalk uncovers a mechano-metabolic pathway where mitochondrial shifts drive defense against kidney fibrosis.

overall_design: Heterozygous Ecm1 mice (Product #: S-KO-01828) were obtained from Cyagen (Santa Clara, CA). Male and female Ecm1 heterozygous mice were mated to generate Ecm1-wild type (WT) and Ecm1-knockout (KO) offspring. Homozygous ECM1-floxed mice (Product #: S-CKO-02130) were also acquired from Cyagen (Santa Clara, CA). Col1α2-CreER mice (Strain #: 029567) and Pdgfrβ-P2A-CreERT2 mice (Strain #: 030201) were obtained from the Jackson Laboratory (Bar Harbor, ME). By crossing Ecm1-floxed mice with Col1α2-CreER or Pdgfrβ-P2A-CreERT2 transgenic mice, conditional knockout mice were generated where Ecm1 gene was specifically deleted in kidney Col1α2+ or Pdgfrβ+ fibroblasts (genotype: ECM1fl/fl, Cre ±) were created. These conditional knockouts were bred with homozygous Ecm1-floxed mice (genotype: ECM1fl/fl), yielding litters with 50% Col1α2-Ecm1-/- or Pdgfrβ-Ecm1-/- mice and 50% control mice (Col1α2-Ecm1+/+ or Pdgfrβ-Ecm1+/+). Kidney ECM1 WT slide and ECM1 KO slide were pooled into one spatial transcritpomics imaging for analysis.

first: Yuan

last: Gui

first: Wenxue

last: Li

first: Jia-Jun

first: Yuanyuan

last: Wang

first: Cameron

last: Jones

first: Riddhi

last: Bansal

first: Samantha Mae

last: Mallari

first: Henry Wells

last: Shaffer

first: Saiya

last: Zhu

first: Edward

last: Malakh

first: Yanbao

last: Yu

first: Haiyan

last: Fu

first: Tracy

last: Tang

first: Silvia

first: Yansheng

first: Dong

last: Zhou


title: Defective RNA processing and ELOA-mediated transcriptional elongation in reversible cellular senescence suggest aging by transcription

city: Chicago

first: Ali

last: Shilatifard

state: IL

address: 320 E Superior St

department: Department of Biochemistry and Molecular Genetics

institute: Northwestern University Feinberg School of Medicine

summary: Elongation factors control post-initiation steps of transcription by RNA polymerase II (RNAPII). We have established distinct mechanistic roles for the essential elongation factors PAF1, NELF, SPT5, and SPT6 via acute depletion of each individually in auxin-inducible degron lines. Here, we leverage these degron lines to explore the regulatory intersection of elongation control and mRNA processing. Integrating long- and short-read RNA-seq data to quantify transcript isoform usage at single-molecule resolution, we identify elongation factor-specific RNA processing regulons including a cellular senescence-enriched regulon shared by NELF and SPT6. We then show that long-term depletion of NELF or SPT6 results in reversible growth arrest following early upregulation of a small group of genes, which include the senescence-associated genes CDKN1A (p21) and CCN2. We perform genetic suppressor screens that implicate the elongation factor Elongin A (ELOA) in NELF or SPT6 depletion-induced growth arrest. ELOA KO suppresses NELF depletion-induced mRNA processing defects and the 3’ extension of RNAPII occupancy past transcription end sites (TES) at genes induced by NELF depletion. ELOA also occupies TES-proximal regions under normal conditions, and acute ELOA depletion results in a loss of RNAPII processivity at the 3’ end of genes, opposing the effects of NELF or SPT6 depletion. Finally, we demonstrate that genetic ELOA loss confers a growth advantage to aging human primary dermal fibroblasts. These findings establish the existence of novel ELOA-dependent mechanisms regulating transcription termination-coupled processes, and links these to the complex phenomena of cellular senescence and aging.

overall_design: Long-read profiling of transcriptome in control and perturbed DLD1 cells to investigate the impact of the depletion of transcriptional elongation factors PAF1, SPT5, SPT6, NELF-C, and ELOA

first: Saeid

last: Parast

first: Simai

first: Deniz

middle: G

last: Olgun

first: Marta

last: Iwanaszko

first: Vijay

last: Ramani


title: IFNγ-induced memory in human macrophages is not sustained by epigenetic changes but the durability of the cytokine itself [CUT&Tag]

first: Aleksandr

last: Gorin

overall_design: H3K4me1 CUT&Tag was performed to enhancer marks induced by interferon gamma in human macrophages and assay the necessity of continued JAK/STAT signaling on the persistence of chromatin marks after interferon withdrawal.

first: Vyas

last: Koduvayur

first: Noa

last: Harriott

first: Quen

last: Cheng

first: Alexander

last: Hoffmann


title: Single cell and isoform-specific translational profiling of the mouse brain [Ribo-STAMP]

first: Brian

last: Yee

overall_design: To evaluate Ribo-STAMP's sensitivity in detecting rapid translational changes on the order of minutes to hours, we treated neurons with Brain-Derived Neurotrophic Factor (BDNF) after Ribo-STAMP expression.

first: Sison

last: Samantha

first: Yeo

last: Eugene


title: NSD2 targeting reverses lineage plasticity and drug resistance in advanced prostate cancer [scRNA-seq]

first: Jia

last: Li

first: Alessandro

last: Vasciaveo

first: Michael

last: Shen


title: Combined epigenetic therapy with FACT and BET inhibitors remodels chromatin and disrupts oncogenic transcription in Diffuse Midline Glioma [ATAC-seq]

first: Holly

last: Holliday

summary: Aberrant epigenetic regulation is a hallmark of Diffuse Midline Glioma (DMG), an incurable pediatric brain tumor. The H3K27M driver histone mutation leads to transcriptional dysregulation, indicating that targeting the epigenome and transcription may be a key therapeutic strategy against this highly aggressive cancer. One such target is the Facilitates Chromatin Transcription (FACT) histone chaperone. We found FACT to be enriched at developmental gene promoters, coinciding with regions of open chromatin and binding motifs of core DMG regulatory transcription factors. Furthermore, FACT interacted and co-localized with the Bromodomain and Extra-Terminal Domain (BET) protein BRD4 at promoters and enhancers, suggesting functional cooperation between FACT and BRD4 in DMG. In vitro, a combinatorial therapeutic approach using the FACT inhibitor CBL0137, coupled with BET inhibition revealed potent and synergistic cytotoxicity across a range of DMG cultures, with H3K27M-mutant cells demonstrating heightened sensitivity. These results were recapitulated in vivo, significantly extending survival in three independent orthotopic PDX models of DMG. Mechanistically, we show that CBL0137 treatment decreased chromatin accessibility, synergizing with BET inhibition to disrupt transcription, silencing several key oncogenes including MYC, PDGFRA and MDM4, alongside alterations to the splicing landscape. Combined, these data highlight the therapeutic promise of simultaneously targeting FACT and BRD4 in DMG, proposing a novel strategy for combating this devastating pediatric brain tumor.

overall_design: Two H3.3 K27M DMG cell lines (HSJD-DIPG007 and SU-DIPGVI) were treated with CBL0137 (1µM for DIPG7 and 2µM for DIPGVI) or JQ1 (1µM) as single agents and in combination for 4h. Vehicle cotrol cells were treated with DMSO at the same concentration. ATAC-seq was then performed to measure chromatin accessibility

first: Nisitha

last: Jayatilleke

first: Chelsea

last: Mayoh

first: Samuel

middle: E

last: Ross


title: Combined epigenetic therapy with FACT and BET inhibitors remodels chromatin and disrupts oncogenic transcription in Diffuse Midline Glioma [RNA-seq]

overall_design: Two H3.3 K27M DMG cell lines (HSJD-DIPG007 and SU-DIPGVI) were treated with CBL0137 (1µM for DIPG7 and 2µM for DIPGVI) or JQ1 (1µM) as single agents and in combination for 4h. Vehicle cotrol cells were treated with DMSO at the same concentration. RNA was then extracted and sequenced. RNA is matched from ATAC-seq experiments.

first: Xinyi

last: Guo


title: Combined epigenetic therapy with FACT and BET inhibitors remodels chromatin and disrupts oncogenic transcription in Diffuse Midline Glioma [CUT&RUN]

overall_design: Cleavage Under Targets and Release Using Nuclease (CUT&RUN) for SPT16, BRD4, H3K27M, H3K27me3 and H3K4me3 in 3 DMG cell lines (HSJD-DIPG007, SU-DIPGVI, VUMC-DIPG10). IgG was used as a negative control.


first: Ali

last: Shilatifard

summary: Negative elongation factor (NELF) is a transcriptional regulator that is primarily known for its role in the promoter-proximal pausing of RNA polymerase II (RNAPII) [1-2]. We have previously demonstrated that acute depletion of the NELF subunit NELF-C causes genome-wide RNAPII pausing at a second site associated with the +1 nucleosome [3]. Here, we show that acute depletion of the NELF subunit NELF-C results in a reversible cellular senescence phenotype that includes impaired growth, robustly detectable β-galactosidase activity, and upregulated expression of senescence-associated genes. We also identify a small set of genes at which NELF-C depletion induces RNAPII release rather than second site pausing. We identify Elongin A (ELOA) as a transcription factor mediating the impacts of NELF depletion, and we demonstrate that the primate-specific ELOA homolog ELOA3 can also play this role. Mechanistically, ELOA mediates the impacts of NELF depletion at least in part by altering the kinetic coupling between the elongation and splicing of nascent transcripts, which results in intron retention and other splicing defects. Despite reports implicating splicing defects in aging-related phenomena [4-6] [7-9], mechanisms have remained elusive. The findings presented here indicate that the crosstalk between splicing factors, ELOA, and NELF may be critical to understanding cellular senescence and aging

overall_design: TT-seq in NELFC-AID and ELOA-KO DLD1 cells

first: Saeid

last: Parast

first: Marta

last: Iwanaszko

first: Yuki

last: Aoi


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

Elongation factors control post-initiation steps of transcription by RNA polymerase II (RNAPII). We have established distinct mechanistic roles for the essential elongation factors PAF1, NELF, SPT5, and SPT6 via acute depletion of each individually in auxin-inducible degron lines. Here, we leverage these degron lines to explore the regulatory intersection of elongation control and mRNA processing. Integrating long- and short-read RNA-seq data to quantify transcript isoform usage at single-molecule resolution, we identify elongation factor-specific RNA processing regulons including a cellular senescence-enriched regulon shared by NELF and SPT6. We then show that long-term depletion of NELF or SPT6 results in reversible growth arrest following early upregulation of a small group of genes, which include the senescence-associated genes CDKN1A (p21) and CCN2. We perform genetic suppressor screens that implicate the elongation factor Elongin A (ELOA) in NELF or SPT6 depletion-induced growth arrest. ELOA KO suppresses NELF depletion-induced mRNA processing defects and the 3’ extension of RNAPII occupancy past transcription end sites (TES) at genes induced by NELF depletion. ELOA also occupies TES-proximal regions under normal conditions, and acute ELOA depletion results in a loss of RNAPII processivity at the 3’ end of genes, opposing the effects of NELF or SPT6 depletion. Finally, we demonstrate that genetic ELOA loss confers a growth advantage to aging human primary dermal fibroblasts. These findings establish the existence of novel ELOA-dependent mechanisms regulating transcription termination-coupled processes, and links these to the complex phenomena of cellular senescence and aging.

overall_design: Refer to individual Series


title: NAC controls nascent chain fate through tunnel sensing and chaperone action [Ribo]

city: Stanford

first: Jae Ho

last: Lee

address: 318 Campus Drive E200

department: Biology

institute: Stanford University

summary: The nascent polypeptide-associated complex (NAC) is a conserved ribosome-bound factor with essential yet incompletely understood roles in protein biogenesis1. Here, we show that NAC is a multifaceted regulator that coordinates translation elongation, cotranslational folding, and organelle targeting through distinct interactions with nascent polypeptides both inside and outside the ribosome exit tunnel. Using NAC-selective ribosome profiling in C. elegans, we identify thousands of sequence-specific NAC binding events across the nascent proteome, revealing broad cotranslational engagement with hydrophobic and helical motifs in cytosolic, nuclear, ER, and mitochondrial proteins. Unexpectedly, we discover an intra-tunnel sensing mode, where NAC engages ribosomes with extremely short nascent polypeptides inside the exit tunnel in a sequence-specific manner. Moreover, initial NAC interactions induce an early elongation slowdown that tunes ribosome flux and prevent ribosome collisions, linking NAC’s chaperone activity to kinetic control of translation. We propose NAC action protects aggregation-prone intermediates by shielding amphipathic helices, thus promoting cytonuclear folding. NAC also supports mitochondrial membrane protein biogenesis and ER targeting by early recognition of signal sequences and transmembrane domain. Our findings establish NAC as an early-acting, multifaceted orchestrator of cotranslational proteostasis, with distinct mechanisms of action on nascent chains depending on their sequence features and subcellular destinations.

overall_design: Selective-ribosome profiling of NAC and NAC(deltaN1-53), and translatome profiling of C. elegans upon NAC_KD

first: Jae

first: Laurenz

last: Rabl

first: Martin

last: Gamerdinger

first: Fabian

last: Morales-Polanco

first: Elke

last: Deuerling

first: Judith

last: Frydman


title: TRIM40 Drives Pathological Cardiac Hypertrophy and Heart Failure through PKN2 Ubiquitination

status: Public on Dec 21 2025

city: Jilin City

first: Mengyang

last: Wang

state: Jilin

address: No. 3999, Huashan Road, Fengman District, Jilin City, Jilin Province

country: China

institute: Beihua university

summary: Pathological cardiac hypertrophy is a key predisposing factor for heart failure (HF). This study investigates the role of the E3 ubiquitin ligase Tripartite Motif-Containing 40 (TRIM40) in cardiac hypertrophy. Using TRIM40 knockout (TRIM40-/-) and cardiac-specific overexpressing mice, pathological hypertrophy was induced by angiotensin II (Ang II) infusion or transverse aortic constriction (TAC). Results demonstrated that TRIM40 expression was upregulated in hypertrophic hearts. TRIM40 deficiency attenuated cardiac hypertrophy and dysfunction, whereas its overexpression exacerbated pathological remodeling. Mechanistically, TRIM40 binds Protein Kinase N2 (PKN2) via its B-box domain, promoting K63-linked ubiquitination at cysteine 29 that enhances PKN2 phosphorylation at Ser815 and activates downstream signaling. Pharmacological inhibition of PKN2 attenuated cardiac remodeling induced by TRIM40 overexpression. These findings indicate that TRIM40 promotes cardiac hypertrophy through K63-linked ubiquitination and activation of PKN2, identifying TRIM40 as a potential therapeutic target for HF.

overall_design: We performed RNA sequencing analysis on the heart tissues of three control group mices and three Ang II-induced mices.

first: Risheng

last: Zhao


overall_design: polyA RNA-seq in NELFC-AID and ELOA-KO DLD1 cells


title: Epigenetic Signatures of Brain Aging by Single Nucleus Methylation Sequencing [DHC-18mo-m-003]

first: Joseph

last: Ecker

summary: Illustrating aging effects on the epigenomic features of brain is critical to understand brain functional decline and neurodegeneration. Single nucleus methylation sequencing was applied to brain regions of both male and female mouse at the age of 2, 9 and 18 months. We use these data to identify the epigenomic changes with aging in mouse brains.

overall_design:
Apply snmC-seq3 to mouse brain regions to identify aging effects on epigenomics

Processed data for these samples are available at:

https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE313770

first: Q

last: Zeng

first: Wei

last: Tian

first: Anna

last: Bartlett

first: J

last: Nery

first: Rosa

last: Gomez Castanon

first: Julia

last: Osteen

first: Jordan

last: Altshul

first: N

last: Johnson

first: M

last: Kenworthy

first: C

last: Valadon

first: Huaming

last: Chen

last: Behrens


title: Epigenetic Signatures of Brain Aging by Single Nucleus Methylation Sequencing [DHC-18mo-m-002]


title: NAC controls nascent chain fate through tunnel sensing and chaperone action[RNA]

first: Jae Ho

last: Lee

overall_design: Transcriptome profiling of whole lysate, membrane-bound, and soluble fraction of C. elelgans upon NAC_KD.

first: Natalia

last: Barbosa

first: Jae

first: Fabian

last: Morales-Polanco

first: Martin

last: Gamerdinger

last: Deuerling


title: Essential role of NONO-HOXA1-Wnt axis in cardiomyocyte differentiation [ChIP-seq]

city: Shanghai

first: Feizhen

last: Wu

state: Shanghai

address: Zhangheng Road 825, Pudong

department: Intelligent Medicine Institute

institute: Fudan Univ, Shanghai, China

summary: NONO is recognized as a critical molecular scaffold involved in both transcriptional and posttranscriptional regulation. Mutations in NONO are frequently linked to congenital heart diseases (CHDs) in humans. However, the mechanisms by which NONO regulates cardiac development remain elusive. Here, we identified NONO as a pivotal dual-function regulator of cardiomyocyte differentiation in human induced pluripotent stem cells (hiPSCs). NONO deficiency in hiPSCs results in a distinct defect in early cardiomyocyte differentiation. Mechanistically, NONO interacts with HOXA1 and regulates the dynamic expression of key genes during early cardiomyocyte differentiation. ChIP-seq analysis reveals that NONO loss reduces HOXA1 occupancy at target genes, compromising its transcriptional regulation. Additionally, NONO and HOXA1 cooperatively activate the Wnt signaling. Taken together, these findings establish the NONO-HOXA1-Wnt axis as a key molecular mechanism in cardiomyocyte differentiation and provid insights into the etiology of CHDs associated with NONO mutations.

overall_design:
We performed ChIP-seq of NONO in Day2-differentiated cells. We performed ChIP-seq of HOXA1 in WT and NONO-KO Day2-differentiated cells overexpressing Dox-induced HOXA1-HA using anti-HA tag antibody, Input of ChIP-seq in Day2-differentiated cells, mRNA-seq of WT,  NONO-KO Day2, 5, 7, 9-differentiated cells and HOXA1-KO Day2-differentiated cells.
We performed reChIP-seq of NONO in Day2-differentiated cells overexpressing Dox-induced HOXA1-HA using anti-HA for primary ChIP and anti-NONO for re-ChIP. We performed ChIP-seq of HOXA1 in WT and NONO-KO Day2-differentiated cells using anti-H3K27AC antibody, Input of ChIP-seq in Day2-differentiated cells. We performed ATAC-seq in Day2-differentiated cells.

last: Sheng

first: Guoying

last: Huang

first: Zhiyu

last: Feng

first: Yuan

last: Gao


title: Nonsense-mediated mRNA decay orchestrates neuronal migration and cortical lamination while modulating reelin and ciliary gene regulatory networks (snRNAseq)

status: Public on Dec 20 2025

city: Riverside

first: Sika

last: Zheng

address: University Ave 900

department: School of Medicine

institute: University of California, Riverside

summary: Nonsense-mediated mRNA decay (NMD) is associated with various neurodevelopmental disorders. Here, we demonstrate that NMD, mediated by UPF2, previously not linked to cortical organization, is indispensable for neuronal migration and cortical lamination. Conditional deletion of Upf2 in radial glial cells delays neuronal migration and disrupts cortical lamination. Trp53 knockout rescues microcephaly from Upf2 deficiency but cannot rescue lamination defects, showing that UPF2’s role in neuronal migration is uncoupled from its regulation of cell cycle and independent of p53. UPF2 deficiency downregulates key neuronal migration genes in the Reelin signaling pathway and microtubule assembly (e.g., Dab1, Lrp8, Tubb2b, Tuba1a), partly through upregulation of the transcriptional repressor Ino80. Additionally, NMD inhibition causes widespread upregulation of ciliary genes. Ectopic expression of Foxj1, a master regulator of ciliary genes and NMD target, impedes neuronal migration, mimicking the Upf2 knockout phenotype. Therefore, NMD is a central post-transcriptional mechanism that coordinates migration and ciliary gene networks crucial for cortical structure development, providing insight into how NMD dysfunction contributes to neurodevelopmental disorders.

overall_design: Comparative gene expression analysis for E17.5 control and Emx1-Upf2cKO cortices at single-nucleus resolution.

first: Lin

last: Lin

first: Naoto

last: Kubota


title: Mapping breakome of multiple cancer cell-lines

city: Jerusalem

first: Rami

last: Aqeilan

address: Ein Kerem

country: Israel

institute: Hebrew University of Jerusalem

summary: Cancer is characterized by uncontrolled proliferation accompanied by the hypertranscription of oncogenes, leading to transcription stress, a key source of DNA double-strand breaks (DSBs) that jeopardize genomic stability. Yet, transcription stress is still underexplored. In this study, we utilized maps of DSBs identified through in-suspension break labeling in situ and sequencing (sBLISS), along with transcription stress markers, revealing that transcription stress regions coincide with the super-enhancer regulatory landscape. Notably, γH2AX mapping indicates its enrichment at transcription stress sites while not all DSB-enriched genes show equal γH2AX marking, but those with DSBs tied to transcription stress are distinctly marked. Intriguingly, genes with high-DSBs marked by γH2AX exhibited significantly higher DSB turnover and repair than those with γH2AX-low genes, manifesting vulnerability to mutagenesis. These findings underscore super-enhancer activity as a determinant of the transcription stress landscape in cancer, posing a threat to the genomic stability of oncogenes in cancer.

overall_design: MCF7 (HTB-22) and T-47D (HTB-133) cells were grown in RPMI supplemented with 10% (vol/vol) fetal bovine serum FBS (GIBCO), glutamine, and penicillin/streptomycin. HEK-293 (CRL-1573), MDA-MB-468 (HTB-132), MDA-MB-231 (HTB-26), and HeLa cells (CCL-2) were grown in DMEM supplemented with 10% (vol/vol) FBS, glutamine, and penicillin/streptomycin. A-4098 cells (HTB-44) were grown in. EMEM supplemented with 10% FBS, glutamine, and penicillin/streptomycin. SH-SY5Y (CRL-2266) were grown in DMEM/F12 supplemented with 10% FBS, glutamine, and penicillin/streptomycin. HMLE cells were grown in Promocell mammary epithelial cell basal media (C-21010) with added supplements (c-93110) whereas MCF10A were grown on DMEM/F12 supplemented with 5% Horse serum, 20 ng/ml EGF, 0.5 mg/ml Hydrocortisone, 100 ng/ml Cholera toxin, 10 mg/ml Insulin and Pen/Strep. MCF-7 cells were synchronized in the G1 cell cycle stage by culturing in serum-free media for 72 hours followed by replacing the media with 5% FBS media for 5 hours. Cells were grown at 37°C under a humidified atmosphere with 5% CO2. Cells were routinely authenticated by STR profiling, and tested for mycoplasma, and cell aliquots from early passages were used. Cells were treated with DMSO (Sigma-Aldrich), dBET6 (Sigma-Aldrich, SML2638), or CDK9i (NVP-2) (MCE, Cat# Y-12214A) 100 nM for 4 hours. Cells were then treated with DMSO or ATMi (20 uM, KU-55933, Sigma) for 5 hours. Transient transfection of siXRCC4 (Dharmacon; SMARTpool siRNA, Catalog ID: L-004494-00-0005) and siSc (Dharmacon; D-00181010) was carried out using Lipofectamine (ThermoFisher). On the day of transfection, cells were seeded to achieve a confluency of 60-70%. The transfection solution was prepared by mixing 1.5 ml of serum and antibiotic-free RPMI with 30 µL of Lipofectamine and incubating for 5 minutes before adding a mixture of 1.5 ml RPMI and 0.6 nmoles of siRNA, followed by a 20-minute incubation at room temperature. This mixture was then added to cells in a 10 cm plate cultured in antibiotic and serum-free media. The media was replaced with fully supplemented media after 5-6 hours, and the cells were incubated in the incubator for 48 hours.

first: Osama

last: Hidmi

first: Sara

last: Oster

first: Diala

last: Shatleh

first: Jonathan

last: Monin


first: Ali

last: Shilatifard

overall_design: ChIP-seq for Pol2, pSer2, PAF1, SPT6, ELOA, ELOA3 in NELFC-AID DLD-1 cells

first: Saeid

last: Parast

first: Marta

last: Iwanaszko


title: Nonsense-mediated mRNA decay orchestrates neuronal migration and cortical lamination while modulating reelin and ciliary gene regulatory networks (RNAseq)

overall_design: Comparative gene expression analysis for E18.5 control and Emx1-Upf2cKO cortices.


title: Longitudinal monitoring of Type 1 Diabetes progression to disease onset

city: Ann Arbor

first: Lonnie

last: Shea

state: MI

address: 1600 Huron Parkway

department: Biomedical Engineering

institute: University of Michigan

summary: Preventing autoimmune type 1 diabetes (T1D) necessitates improved monitoring for disease progression prior to symptom onset. Current diagnostic methods assess circulating autoantibodies, C-peptide levels, or dysglycemia, yet these approaches fail to identify β cell destruction preceding glucose dysregulation. Herein, a subcutaneous microporous scaffold is employed as an immunological niche (IN), which provides a non-vital accessible tissue reflecting many immune changes occurring in the pancreas. RNA sequencing analysis of the IN successfully delineates at-risk from non-risk groups, as well as disease progressors from non-progressors at six weeks of age in the NOD mouse model. Within progressors, we identify disease five to seven weeks prior to disease onset. Collectively, disease occurring in a poorly accessible site can be identified early by sampling a distant non-vital tissue, indicating the systemic nature of the disease and informing the timing of disease modifying therapies to halt or delay the progression of T1D.

overall_design: RNA-seq profiling of microporous scaffolds implanted into NOD and NOR mice. Samples were taken from all mice at 6 weeks of age. Additional samples were taken at 7, 5, 3, and 1 week before symptom onset in NOD mice that progressed to hyperglycemia, as well as at the time of symptom onset. Samples were taken at 10, 14, 18, and 22 weeks of age for NOD mice that did not develop hyperglycemia and NOR controls.

first: Jessica

last: King


title: Fission yeast Whi5 represses MBF-dependent transcription in quiescent cells

city: Salamanca

first: Sergio

last: Moreno

email: smo@usal.es, ibfg_smp@usal.es

state: Salamanca

address: Zacarias Gonzalez 2

department: IBFG

country: Spain

institute: Institute of Functional Biology and Genomics (IBFG) (CSIC)

summary: When cells arrest the cell cycle in G1 to enter the non-dividing quiescent state, the transcriptional machinery that promotes G1/S transition must be inactivated. Central to this regulation is Whi5, in budding yeast, and the Retinoblastoma (Rb) protein, in mammalian cells, acting as repressors of the transcription factors SBF, and E2F, respectively. In fission yeast, the MBF complex is the functional homologue of E2F in metazoans. Fission yeast Whi5/Mug54 is an uncharacterized protein predicted to function as a cell cycle transcriptional repressor at the G1/S transition. In this work, we show that after induction of quiescence by nitrogen starvation, fission yeast Whi5 is localised to the nucleus, and it is required for the repression of MBF-dependent genes. Mass spectrometry and bimolecular fluorescence complementation (BiFC) analyses revealed that Whi5 physically interacts with components of both the MBF and the histone deacetylase Clr6-I complexes. Moreover, Whi5 is required for the interaction of the MBF complex with Clr6-I, suggesting a role for Whi5 in maintaining transcriptional repression of MBF-dependent genes in quiescent cells by the recruitment of HDACs to gene promoters.

overall_design: We performed a comparative analysis of gene expression profiles from RNA-seq data between wild-type cells and the mutant whi5∆ growing in minimal medium (MM) and in minimal medium lacking nitrogen (MM-N) after 10 and 24 hours.

first: Celia

last: Gálvez-Merchán

first: Rafael

last: López-San Segundo

first: M. Belén

last: Suárez

first: Daniel

last: González-Ávarez

first: José

last: Ayte

first: Livia

last: Pérez-Hidalgo


title: Fission yeast Whi5 represses MBF-dependent transcription in quiescent cells II

overall_design: We performed a comparative analysis of gene expression profiles from RNA-seq data between wild-type cells and the mutants whi5∆, nmt41-whi5+, nrm1-3xsAID and nrm1-3xsAID whi5∆ growing in minimal medium (MM) and in minimal medium lacking nitrogen (MM-N) after 24 hours.


title: Chronic NK cell activation results in a dysfunctional, tissue resident-like state mediated by KLF2 deficiency

city: Providence

first: Jacob

middle: Andrew

last: Myers

state: RI

address: 171 Meeting St.

institute: Brown University

summary: Adoptive transfer of NK cells can induce complete remissions in 30-50% of patients with refractory acute myeloid leukemia and lymphoma. While blood chimerism occurs, attaining functional homing to the site of tumor without exhaustion has been elusive. During chronic infections and tumorigenesis, exposure to activating stimuli weakens the effector activity of NK cells. Despite this knowledge, there is little known about the mechanisms that govern this dysregulation and whether these disparate activating stimuli use distinct pathways to downregulate effector immunity. In this study, we reveal that chronic NK cell activating receptor (NKAR) stimulation and chronic IL-15 exposure impart distinct modes of dysregulation, with NKAR stimulation inducing a tissue resident-like state that resembles that of tumor-infiltrating NK cells in cancer patients. Using loss- and gain-of-function studies, we identify the transcription factor KLF2 as a master regulator of the NK cell response to chronic activation and provide evidence that KLF2 overexpression promotes NK cell cytotoxicity, cytokine production, and chemotaxis, and inhibits the development of dysfunctional, tissue resident-like features. Using KLF2 reporter mice, we show that in certain tissues, tissue resident NK cells are predominantly KLF2-negative while circulating NK cells in these tissues are overwhelmingly KLF2 + . Lastly, using mixed bone marrow chimeras, we demonstrate that conditional KLF2 deficiency in NK cells leads to altered homing and the acquisition of tissue resident-like features in vivo. Together, these findings highlight the profound changes NK cells undergo during prolonged activation and advance our understanding of how some NK cell therapies fail during malignant relapse.

overall_design: Details of experimental design can be found within the associated publication. There are datasets from two separate figures in this directory. The following is a summary of design for the data in Figure 2: in brief, NK cells from healthy human donor peripheral blood mononuclear cells (PBMCs) were isolated and stimulated with either NK cell activating receptor (NKAR) agonists (i.e. anti-NKp46 and MICA/B) or IL-15 for 4 or 10 days. 10 day stimulations were performed with either continuous IL-15 or with a 2-day (days 5 and 6) reduction in the concentration of IL-15. The following is a summary of design for the data in Figure 4: in brief, human NK cells were isolated as previously described and expanded using K562 leukemia cells expressing membrane-bound IL-21 and 4-1BB-L. Following expansion, NK cells were transduced with AAV vectors carrying either a control insert (mNG) or KLF2-overexpression vector (KLF2-mNG). Following transduction, cells were stimulated for 4 days on control or NKAR-agonist plates as previously described. Cell samples were also collected post-transduction, but prior to plate-bound activation.

middle: A


title: Deoxynivalenol(DON)  and  related compounds target protein processing to induce immunotoxic effects in human T cells and macrophages [Jurkat]

city: Beijing

first: Hui

last: Yang

address: 7 Panjiayuan Nanli, Room314

department: Department of Toxicology

institute: China National Center for Food Safety Risk Assessment (CFSA)

summary: Deoxynivalenol (DON) is a fungal toxin mainly produced by Fusarium graminearum and Fusarium graminearum, and is present in cereal crops such as wheat, barley, oats, rye, and corn. It is widely polluted worldwide. DON and its related derivatives have a wide range of toxic effects on humans and animals. This study used zebrafish models and different cell models to verify the toxic effects of DON, and explored the mechanism of DON mediated Jurkat apoptosis and inhibition of M2 macrophage differentiation. The results indicate that the toxic effects of these fungal toxins differ significantly among various cell types. For Jurkat T cells, the IC50 values of DON, NIV, 15 Ac DON, and 3-Ac DON were 488.6 nM, 318.2 nM, 544 nM, and 6516 nM, respectively. There are significant differences in the effects on M2 polarization of macrophages, and DON, NIV, and 15 Ac DON can inhibit M2 differentiation and function of macrophages at lower dose levels. Indicating that immune toxicity is an important toxic effect of DON and its related substances. Research on its mechanism has shown that DON can target protein processing processes, interfere with the normal synthesis and folding of proteins in cells, and trigger cellular stress responses. These findings reveal the toxic effects and potential mechanisms of DON on human immune cells, providing a scientific basis for preventing and controlling DON pollution and its harm to human health.

overall_design: Study the immunotoxicity of DON and its related substances using Jurkat T cells and M2 macrophages. Total RNA was extracted using TRIzol for transcriptome sequencing, and three biological replicates were set for Jurkat T cells and M2 macrophages.

first: Jing

first: Xiaohong

last: Wang

first: Lijuan

last: You

first: Qiannan

last: Zhang

first: Xudong

last: Jia


title: Silencing lipid catabolism determines longevity in response to fasting [fasting RNA-Seq]

status: Public on Dec 19 2025

city: Dallas

first: Peter

last: Douglas

state: Texas

address: 6000 Harry Hines Blvd

department: Department of Molecular Biology

institute: University of Texas Southwestern Medical Center at Dallas

summary: Oscillations between lipid anabolism and catabolism allow for long-term preservation of cellular health amid systemic metabolic fluctuations. As a conserved aging determinant, fasting can improve disease outcomes and extend lifespan. Yet, the relative importance of activating lipid catabolism versus its attenuation in fasting-induced longevity remains unclear. The robust adaptability of the soil-dwelling worms, C. elegans, to variable nutrient availability provides an excellent means to better understand how metabolic transitions alter aging trajectories. Here, we show that, rather than activation, the silencing of lipid catabolism upon nutrient replenishment is essential for lifespan extension through fasting. The fasting-responsive nuclear hormone receptor, NHR-49, is pivotal in activating lipid catabolism through β-oxidation. Unlike traditional ligand-regulated nuclear hormone receptors, NHR-49 employs a unique regulatory mechanism that bypasses ligand binding, instead relying on cofactors to mediate its transcriptional attenuation and turnover during times of nutrient stress. Here, we identify casein kinase 1 alpha 1 (KIN-19) as a central regulator of metabolic plasticity and fasting-induced longevity, which attenuates β-oxidation via primed phosphorylation of NHR-49. Overall, cooperative, ligand-independent silencing of this conserved nuclear hormone receptor promotes longevity associated with fasting.

overall_design: RNAseq profiling of N2 C.elegans were collected at Day 1 (fed), Day 2 (24 hours fasted), and Day 3 (24 hours refed).

first: Lexus

last: Tatge


title: Overactivation of Cdc42 GTPase Impairs the Cytotoxic Function of NK Cells from Old Individuals towards Senescent Fibroblasts

city: Ulm

first: Pallab

last: Maity

address: Meyerhofstrasse/James Franck Ring

department: Dermatology

institute: University of Ulm

summary: Senescent fibroblasts accumulate in the connective tissue of all organs and promote organ aging and aging-related diseases. The underlying mechanisms for the accumulation of senescent fibroblasts are poorly understood. Natural killer (NK) cells of innate immunity play a critical role in the removal of tissue resident senescent cells. We here show that NK cells from old adults and old mice fail to efficiently remove senescent fibroblasts due to severely reduced efficiency of aged NK cells to induce perforin mediated membrane disruptions and granzyme B induced targeted death of senescent fibroblasts. We demonstrate elevated activation of the small Cdc42 Rho GTPase in aged NK cells to be responsible for the disruption of the microtubular organisation of aged NK cells. A correct microtubular organisation of NK cells is essential for cellular polarity, degranulation of perforin and granzyme B into the synaptic cleft between NK cells and senescent fibroblasts, and for a proper function of energy metabolism. Attenuation of the elevated activity of Cdc42 in aged human NK cells with CASIN, a small molecule Cdc42 inhibitor, rebalances Cdc42 activity to a young level, restores a proper perforin and granzyme B release and attenuates reduced ATP levels and mitochondrial dysfunction in aged NK cells to result in a youthful cytotoxicity of aged NK cells against senescent cells. Collectively, we identified a previously unreported molecular mechanism underlying functional impairment of NK cells from older adults. In perspective, our data hold promise to develop novel strategies against age-related disorders driven by tissue-resident senescent fibroblasts.

overall_design: Total RNASeq profiling of natural killer (NK) cells from young and aged human donors


title: Immunotoxic effect and mechanisms of Fusarium mycotoxins on human immune cells: A focus on T cells and macrophages [M2]

first: Ying

last: yang

last: wang

last: zhang


title: Identification of a JAK/STAT/miR155HG positive feedback loop in regulating NK cell proliferation and effector functions

first: Yongjie

last: Liu

address: No. 1 West Beichen Road, Chaoyang District

institute: Institute of Genetics and Developmental Biology

summary: The Janus kinase (JAK)-signal transducer and activator of transcription (STAT) signaling pathway control NK cell development, expansion and cytotoxic functions. Whether lncRNA regulates the JAK-STAT signaling in NK cells remains unknown. We found that miR155HG was upregulated in activated NK cells and promoted their proliferation and effector functions. Mechanistically, miR155HG bound to miR-6756 and relieved its repression of JAK3 expression, resulting in increased JAK3 level and enhanced NK cells proliferation and function. Further investigations disclosed that upon cytokine stimulation, STAT3 directly interacts with miR155HG promoter and induces miR155HG transcription. Collectively, we identify a miR155HG-mediated positive feedback loop of the JAK-STAT signaling. In brief, STAT3 interacts with miR155HG promoter and induces miR155HG transcription upon cytokine stimulation, and miR155HG may work as a sponge for miR-6756, preventing the miR-6756-mediated repression of JAK3 expression, thereby promoting the JAK-STAT pathway and enhancing NK cell proliferation and function. These findings may provide a promising therapeutic target to improve NK cell therapy.

overall_design: RNA-seq of NK-92-shCtrl/shmiR155HG cells with three replicates


title: Silencing lipid catabolism determines longevity in response to fasting

overall_design: RNAseq profiling of N2 C.elegans were collected at Day 1 under empty vector control RNAi or kin-19 RNAi


title: NSD2 targeting reverses lineage plasticity and drug resistance in advanced prostate cancer [snATAC-seq and multiome snATAC-seq]

city: New York

first: Jia

last: Li

institute: Columbia University Irving Medical Center

summary: Treatment with highly potent androgen receptor signaling inhibitors (ARSIs), such as enzalutamide and abiraterone promotes lineage plasticity in metastatic castration-resistant prostate cancer (mCRPC), which results in intra-tumor heterogeneity and emergence of mCRPC subtypes. The histological transformation from adenocarcinoma to aggressive neuroendocrine prostate cancer (CRPC-NE) has been associated with a loss of dependency on lineage-survival signals, leading to targeted drug resistance. Epigenomic reprogramming might be a fundamental driver of lineage plasticity. To determine CRPC-NE vulnerabilities, we have performed image-based screening using a small library of antibodies targeting different histone marks on CRPC-NE organoids derived from genetically engineered mice. We find that the histone mark H3K36me2 and the histone methyltransferase NSD2 (also known as MMSET/WHSC1) play important roles in maintaining the state of CRPC-NE. Knockout of NSD2 or ablation of H3K36me2 using H3.3K36M oncohistone reverted CRPC-NE to CRPC-AR-like phenotype. Simultaneous profiling of the transcriptome and epigenome from single cells collected from control (sgCtrl) and NSD2 knockout (sgNSD2) organoids, or from empty vector (EV) and H3.3K36M transduced organoids confirms the lineage reversal of treatment-resistant cells and maps the re-establishment of canonical AR signaling dependency. Moreover, H3K36me2 or NSD2 depleted mouse and human CRPC-NE organoids responded to enzalutamide treatment in vitro and in vivo, suggesting a restoration of ARSI sensitivity. Most importantly, a small molecule inhibitor of NSD2, which is similar to KTX-1001 (NCT05651932), in combination with enzalutamide leads to growth suppression or apoptosis of mouse and human CRPC-NE organoids, organoids of other CRPC subtypes and xenografts in vitro and in vivo. In conclusion, inhibition of NSD2 reverses lineage plasticity and reverts the state of treatment-resistant cells back into an ARSI-sensitive state. Thus, we suggest that the combination of NSD2 inhibition with AR inhibition may represent a novel therapeutic approach for patients with CRPC-NE and other CRPC subtypes.

overall_design: The mouse NE tumors derived from NPp53 (Nkx3.1CreERT2/+; Ptenflox/flox; TrpP53flox/flox; Rosa26-EYFP) mice were cultured as organoids. To detect intercellular heterogeneity within organoids, I used single-cell RNA sequencing (scRNA-seq) to analyze lineage diversity of five mouse NE tumor organoids on passage 2. I also used single-nucleus ATAC sequencing (snATAC-seq) to profile the chromatin accessibilities of NPPO-1 organoids on passage 8, a multilineage NE tumor organoid line. The non-NE tumor cells from NPPO-1 organoids on passage 3 were isolated by fluorescence-activated cell sorting (FACS) according to a higher side scatter (SSC) value in non-NE compared to NE tumor cells. Some isolated non-NE tumor cells were analyzed by scRNA-seq to detect any contamination by NE tumor cells. Other isolated non-NE tumor cells were transduced with H2B-RFP lentivirus without antibiotic selection. H2B-RFP labeled tumor cells were either cultured alone or co-cultured with NE tumor cells to induce non-NE to NE conversion. To trace cell lineage of H2B-RFP-labeled cells, organoids from either monoculture or co-culture on passage 4 were analyzed by scRNA-seq (gene name: addgene26001). To quantitively compare the histone mark expression between NE and non-NE cells, epigenomic profiling using Cut&Tag was performed on 4 NE and 4 non-NE organoids derived from the same NPp53 background. To accurately quantify gene regulation, bulk RNA-seq was performed alongside epigenomic profiling on the organoids. To study the effects of NSD2 on NE lineage maintenance, we knockout of NSD2 on two mouse NE tumor organoids using CRISPR/Cas9. After antibiotic selection for 14 days, positive-transduced cells were isolated by FACS sorting according to the presence of RFP reporter signal. We used multiome single-nucleus ATAC and RNA sequencing (multiome snATAC + snRNA-seq) to analyze changes of cell lineage and state in sgNSD2 transfected NE tumor organoids in compared with sgControl (sgCtrl) organoids. To provide direct evidence of H3K36me2 histone modification in modulating NE lineage, mouse NE tumor organoids were transduced with lentivirus carrying the gene encoding the H3.3K36M oncohistone that can cause significant reduction in global levels of H3K36me2. After antibiotic selection for 14 days, cells positively transduced with H3.3K36M or empty vector (EV) control were analyzed using multiome snATAC + snRNA-seq to detect changes in cell lineage and state. In parallel, quantitative epigenomic profiling using Cut&Tag was performed on sgCtl and sgNSD2, or EV and H3.3K36M organoids to quantify global epigenomic changes. We have synthesized a small molecular inhibitor of NSD2 (NSD2i), which is similar to KTX-1001 (NCT05651932) in phase 1 clinical trial in patients with relapsed and refractory multiple myeloma. To evaluate if NSD2i recapitulates the effects of genetic interventions in reversal of lineage plasticity and restoring androgen receptor signaling inhibitor (ARSI) sensitivity, 1 µM NSD2i or DMSO was applied every other day on NPPO-1NE organoids cultured in a medium without DHT for 12 days. Organoids treated with either DMSO or 1 µM NSD2i were then collected for multiome snATAC + snRNA-seq analysis or Cut&Tag epigenomic profiling.

first: Alessandro

last: Vasciaveo

first: Michael

last: Shen


title: Excess FGFR3 signaling in achondroplasia disrupts turnover of resting zone chondrocytes via CREB signaling

city: Suita

first: Noriyuki

last: Tsumaki

state: Select One

address: 2-2 Yamadaoka

department: Department of Biochemistry and Molecular Biology

country: Japan

institute: Osaka University, Graduate School of Medicine

summary: Achondroplasia, associated with gain-of-function mutations in FGFR3, causes growth plate cartilage dysfunction, resulting in short-limb dwarfism. However, its precise molecular and cellular mechanisms remain unclear. To address this, we aimed to generate knock-in mice (Fgfr3Ach) harboring the achondroplasia mutation (p.Gly380Arg). In addition to previously reported abnormalities, we observed an expansion of the resting zone. EdU labeling and lineage tracing analysis revealed disrupted stem cell properties of resting zone chondrocytes, preventing self-renewal, limiting cell divisions and migration toward the primary ossification center. Single-cell RNA-seq and immunohistochemical analysis identified a cell cluster that corresponded to the expanded resting zone. Pathway analysis and functional experiments revealed that CREB disrupts stem cell properties in resting zone chondrocytes and contributes to dwarfism. Administration of CREB inhibitor 666-15 restored growth plate pathology and bone length. These findings demonstrate that how excess FGFR3 signaling disrupts resting zone chondrocyte properties and suggest potential therapeutic targets for achondroplasia.

overall_design: We harvested growth plate cartilage from wild-type mice and knock-in mice bearing the p.Gly374Arg mutation in Fgfr3 (Fgfr3Ach) at 19- and 22-days old. We subjected these samples to scRNA-seq analysis.

first: Nanao

last: Horike

first: Saeko

last: Koyamatsu

first: Noriko

last: Tanaka


title: NSD2 targeting reverses lineage plasticity and drug resistance in advanced prostate cancer [multiome snRNA-seq]


title: High-resolution in vivo binding data show that TBP initiates transcription at TATA promoters with and without TFIID

city: Kansas City

first: Melanie

last: Weilert

state: MO

institute: Stowers Institute for Medical Research

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

overall_design: Refer to individual Series


title: Skeletal Muscle Methylome-Transcriptome Disruptions During the Onset and Progression of Colorectal Cancer-Induced Cachexia

city: Fayetteville

first: Ana

middle: Regina

last: Cabrera

state: AR

address: 155 Stadium Dr

institute: University of Arkansas

summary: Cancer cachexia (CC) is experienced by 80% of cancer patients, representing up to 40% of cancer-related deaths. Evidence suggests biological sex dimorphism associated with CC. The initial triggering factors of muscle wasting, biological sex differences, and the epigenetic regulation during the onset and progression of cancer cachexia are not fully understood. The purpose of this study was to define the time course of C26-induced CC in males and females using transcriptomics, compare the transcriptomic changes with a more sever APC+/Min genetic model and identify the impact of epigenetic regulations (methylation) associated to CC.

overall_design: At week-old Balb/c male and female mice (n=6-8/group) receivd 1x10^6 C26 cells injected bilateral, and tumors were developed for 10-, 20-, and 25-days, at endpoints, Tibialis Anterior (TA) muscle were collected.  The control mice received PBS injections  and TAs were colllected 25-days after PBS injections (controls). TA was also collected from 20-weeks-old APC+/Min mice (C57 black mice background) and their wild types (controls). DNA was extracted for RRBS.

first: Nicholas P.

last: Greene

first: Kevin A.

last: Murach

first: Ana Regina


title: High-resolution in vivo binding data show that TBP initiates transcription at TATA promoters with and without TFIID [ChIP-nexus]

summary: This study investigates how the general transcription factor TFIID binds to different core promoter types in vivo. We performed high-resolution ChIP-nexus for all 14 subunits of the Drosophila TFIID complex (TBP and TAF1 to TAF13), as well as for NC2α, NC2β, and Mot1, under both DMSO (control) and Triptolide treatments to capture various transcriptional states.The goal of the study was to define TFIID's in vivo architecture and compare its function across promoters. We show that TBP binding is promoter-specific, enabling de novo classification of TATA, DPR, and housekeeping promoters. Crucially, we provide in vivo evidence for TAF-independent transcription initiation specifically at TATA promoters, which correlates with NC2 binding and offers a mechanistic explanation for their distinct transcriptional bursting properties.

overall_design: ChIP-nexus of General Transcription Factors in Drosophila melanogaster Kc167 cells. Cells are treated with either DMSO (control) or 10 µM triptolide for 1 hour at room temperature. Non treated samples have not been treated for 1h with DMSO.

first: Sergio

last: G-M Alcantara

first: Simon

last: Bourdareau

first: Julia

last: Zeitlinger


title: Regenerating Skeletal Muscle of Ovariectomized Mice

city: Arlington

first: Barbra

last: St. Pierre Schneider

address: 701 S. Nedderman Drive

institute: The University of Texas at Arlington

summary: In this study, we found more stable Siglec1 expression and stronger CD169+ macrophage staining in the muscle of 17-β-estradiol-treated mice than in that of the placebo-treated mice. However, Siglec1 was correlated with more interferon-related genes in the muscle of the placebo-treated mice. These findings suggest that Siglec1/IFN1 activity may be part of the inflammatory response during muscle regeneration. Future studies are warranted to investigate this role.

overall_design: We studied female mice implanted with placebo or 0.18 mg 17‑β‑estradiol pellets, gave a closed crush injury to the right hindlimb after 7 days of treatment, harvested right lateral gastrocnemius at ~32, 96, and 192 h post‑crush for Affymetrix Mouse Gene 1.0 ST microarray profiling, and compared gene expression between estradiol- and placebo-treated groups at each timepoint.

first: Barbara

first: Juli

last: Petereit


title: FGFR signaling and neddylation facilitate SARS-CoV-2 infection by modulating interferon induction and viral entry respectively

city: Edmonton

first: Alberto

last: Felix-Lopez

state: Alberta

department: Medical Microbiology and Immunology

country: Canada

institute: University of Alberta

summary: SARS-CoV-2 is the causative agent of COVID-19, and although vaccines have reduced disease severity, emerging variants remain a significant public health issue. Broadly effective therapeutics are still needed, particularly those targeting host pathways essential for coronavirus infection. Here, we used a CRISPR knockout screen to identify druggable host factors required for SARS-CoV-2 infection. The screen revealed NAE1 and FGFR1 as key contributors to infection. FDA-approved or in clinical trials inhibitors of these pathways reduced replication of both ancestral and contemporary viral variants. Mechanistic studies showed that FGFR1 promotes viral replication through downstream MEK/ERK signaling, while neddylation appears to support viral entry or infectivity rather than replication itself. In a murine model of severe COVID-19, inhibitors of NAE1 and FGFR1 significantly decreased viral load and lung pathology. These findings support the development of host-targeted antiviral strategies.

overall_design: CRISPR KO screen to identify host factors required for SARS-CoV-2 infection

first: Joaquin

last: Lopez-Orozco

first: Mohamed

last: Elaish

first: Nawell

last: Fayad

first: Zaikun

last: Xu

first: Tekeleselassie

last: Woldemariam

first: Bardes B.

last: Hassan

first: Rashmi

last: Panigrahi

first: Juveriya

last: Qamar Khan

first: Megha

last: Rohamare

first: Irv

last: Mayers

first: Mark

last: Glover

first: Joyce

last: Wilson

first: Darryl

last: Falzarano

first: Anil

last: Kumar

first: Tom C

last: Hobman


title: Harnessing lipid-driven immuno-metabolic pathways in omental metastases to enhance immunotherapy in ovarian cancer

city: Gosselies

first: Meggy

last: Suarez-Carmona

address: Rue des Professeurs Jeener et Brachet, 12

country: Belgium

institute: ULB

summary: Despite the potential of immune checkpoint blockade (ICB) in epithelial ovarian carcinoma (EOC), its clinical efficacy remains limited. Our study investigates the immuno-metabolic landscape of omental metastases to identify therapeutic targets. Using patient-derived explants (PDEs), we demonstrate that lipid-rich omental environments preserve effector T cell function, while lipid processing in tumor-associated macrophages (TAMs) induces oxidative stress and immune suppression. Targeting lipid metabolism via CCR5 inhibition (e.g., maraviroc) or anti-CD36 treatment reprograms TAMs, restores T cell activity, and enhances anti-tumor immunity. These findings reveal a metabolic-inflammatory axis that can be modulated to improve ICB outcomes in EOC.

overall_design: Following MDA-MB-231 tumor growth in MISTRG humanized mice, we perfored scRNA-sequencing analysis on human tumor-infiltrating immune cells to analyse the effects of Maraviroc on the antitumor immune response. In total, cells from 7 treated mice and 7 untreated mice were sequenced after hashing.

first: Mareike

last: Hampel

first: Xin-Wen

last: Zhang

first: Alexandra

last: Pöchmann

first: Silke

last: Grauling-Halama

first: Nektarios

last: Valous

first: Pornpimol

last: Charoentong

first: Dyke

last: Ferber

first: Jannis

last: Wissfeld

first: Alicia

last: Höflich

first: Stanislas

last: Goriely

first: Aurélie

last: Detavernier

first: Abdulkader

last: Azouz

first: Anthony

last: Rongvaux

first: Sven

last: Zukunft

first: Ingrid

last: Fleming

first: Jürgen

last: Okun

first: Vickie

last: Baracos

first: Mathias

last: Heikenwälder

first: Laurence

last: Zitvogel

first: Xinyi

first: Chenqi

first: Michael

last: Volkmar

first: Daniel

last: Schraivogel

first: Lars

last: Steinmetz

first: Junzo

last: Hamanishi

first: Masaki

last: Mandai

last: Gaida

first: Theresa

last: Mokry

first: Johanna

last: Nattenmüller

first: Oliver

last: Sedlaczek

first: Nanna

last: Monjé

first: Roxanna

last: Schwab

first: Annette

last: Hasenburg

first: Regina

last: Johanna Boger

first: Frederik

last: Marmé

first: Athanasios

last: Mavratzas

first: Sarah

last: Schott

first: Niels

last: Halama


title: High-resolution in vivo binding data show that TBP initiates transcription at TATA promoters with and without TFIID [RNA-seq]

first: Melanie

last: Weilert

summary: This study investigates how the general transcription factor TFIID binds to different core promoter types in vivo. We performed high-resolution ChIP-nexus for all 14 subunits of the Drosophila TFIID complex (TBP and TAF1 to TAF13), as well as for NC2α, NC2β, and Mot1, under both DMSO (control) and Triptolide treatments to capture various transcriptional states.The goal of the study was to define TFIID's in vivo architecture and compare its function across promoters. We show that TBP binding is promoter-specific, enabling de novo classification of TATA, DPR, and housekeeping promoters. We provide in vivo evidence for TAF-independent transcription initiation specifically at TATA promoters, which correlates with NC2 binding and offers a mechanistic explanation for their distinct transcriptional bursting properties.

overall_design: Total mRNA from Drosophila melanogaster Kc167 cells.

first: Sergio

last: G-M Alcantara

first: Simon

last: Bourdareau

first: Julia

last: Zeitlinger


title: Transcriptomics DNAJC15 and DNAJC19 knockdown in HeLa cells

city: Cologne

first: Lara

last: Kroczek

address: Joseph-Stelzmann-Straße 9B

institute: Max-Planck Institute for Biology of Ageing

summary: To better understand how OMA1 affects mitochondrial proteostasis and stress responses, we performed a proteomic survey of OMA1-deficient cells for proteolytic substrates. We demonstrate that OMA1 cleaves the mitochondrial chaperone DNAJC15 facilitating its degradation by the mitochondrial m-AAA protease. The loss of DNAJC15 alters protein import by TIM23 protein translocases in the IM and limits the accumulation of OXPHOS-related mitochondrial matrix and IM proteins. Non-imported mitochondrial preproteins accumulate at the endoplasmic reticulum (ER) and trigger an ATF6-related unfolded protein response. These results demonstrate that OMA1 allows to adapt mitochondrial protein biogenesis to stress and reveal an intricate network of cellular stress responses to proteostasis disturbances.

overall_design: RNA-seq profile of scrambled control HeLa cell versus DNAJC15 and DNAJC19 knock down.

first: Kroczek

last: Lara

first: Langer

last: Thomas


title: Prenatal and postnatal effects of gestational immune activation on synaptic and neurodevelopmental pathways via epigenetic mechanisms (ChIP-seq)

city: Blacksburg

first: Chang

last: Lu

state: VA

address: 235 Goodwin Hall, 635 Prices Fork Road, Virginia Tech

department: Chemical Engineering

institute: Virginia Tech

summary: Previous epidemiological research suggests that maternal immune activation (MIA) during early pregnancy is a significant risk factor for neurodevelopmental and psychiatric disorders in offspring. Epigenetic factors and chromatin-related phenomena remain largely plastic during the course of prenatal and early postnatal development, allowing a dynamic impact of environmental effects on access to genes and regulatory elements. In this study, we tested the effects of maternal infection with the mouse-adapted influenza A/WSN/33 (H1N1) virus as a MIA model. Additionally, to assess the prenatal and postnatal effects of MIA, we cross-fostered half of the neonatal mice immediately after birth.This study includes RNA-seq and ChIP-seq profiling (H3K27ac, H3K4me3) from the prefrontal cortex of MIA-exposed and control mice. RNA-seq and ChIP-seq data are submitted under separate Series and linked via sample annotations.

overall_design: This study examined transcriptomic and epigenomic changes in offspring following maternal immune activation (MIA) via prenatal influenza infection. Timed pregnant CD1 mice were intranasally infected with 5 × 10³ pfu of influenza A/WSN/33 (H1N1) or mock-treated on embryonic day 9.5. At birth, ~50% of pups were cross-fostered to surrogate dams (MIA or control) to separate prenatal and postnatal influences. Offspring were weaned at P21, housed by sex, and assigned to one of seven groups based on birth and rearing conditions: Group A (Mock-Mock): Offspring born to control mother and raised by their birth mother, together with stepsiblings born to MIA-exposed mother. Group B (MIA-Mock): Offspring born to MIA-exposed mother but raised by control surrogate mother, together with stepsiblings born to this control mother. Group C (Mock-MIA): Offspring born to control mother but raised by MIA-exposed surrogate mother, together with stepsiblings born to this MIA mother. Group D (MIA-MIA): Offspring born to MIA-exposed mother and raised by their birth mother, together with stepsiblings born to control mother. Group E (Mock-Mock_1): Offspring born to control mother and raised by their birth mother, together with stepsiblings born to a different control mother. Group F (Mock-Mock_2): Offspring born to control mother but raised by control surrogate mother, together with stepsiblings born to the control surrogate mother. Group G (Mock-Mock_3): Offspring born to control mother and raised by their birth mother with no stepsiblings. Mice were sacrificed in adulthood. Frontal cortex (bregma 1.90–1.40) was collected for MOWChIP-seq profiling of H3K4me3 and H3K27ac. Neuronal nuclei were isolated via FACS (NeuN⁺), with ~10,000 nuclei used per ChIP replicate. Libraries were prepared using Accel-NGS 2S Plus and sequenced on an Illumina HiSeq 4000 (single-end, 50 bp), yielding ~15 million reads per sample.

first: Bohan

last: Zhu

first: Gaoshan

last: Li


title: A microreactor system for point-of-care viral genome sequencing

summary: Viral genome sequencing is critical for understanding viral biology, identifying strain mutations, and managing public health during pandemics. Although sequencing technologies have progressed over the years in terms of throughput and cost, a portable platform that integrates sample processing, library preparation, and sequencing remains critical for point-of-care viral sequencing that combats viral outbreaks in the field. In here, we demonstrate a semi-automated, field-deployable microreactor system to produce nanopore sequencing libraries from viral samples. The system integrates isothermal amplification, purification, and enzymatic processing within a closed-channel format, minimizing contamination risk and operational complexity. Using Senecavirus A (SVA) as a surrogate for high-consequence pathogens, we demonstrate the system’s ability to prepare sequencing libraries targeting three dispersed genomic regions comprising ~16% of the viral genome and identify single nucleotide variations with high confidence. Together with a portable nanopore sequencer, this microreactor offers a robust solution for decentralized genomic surveillance and rapid diagnostics in resource-limited or outbreak-prone environments.

overall_design: RT-LAMP sequencing of vesicular lession fluid extracted from SVA-positive pigs using an Oxford Nanopore Minion device with Flongle attachment. SVA positive pigs (as determined by RT-PCR) had total RNA extracted from collected vesicular lession samples. RNA  samples underwent reverse transcription loop mediated isothermal amplification (RT-LAMP), performed on a custom designed microfluidic device. This device was then used to purify the generated RT-LAMP amplicons, before performing library preparation for nanopore sequencing. Sequenced samples were analysed to evaluate diagnostic and variant monitoring capabilities of the designed platform.

first: Thomas

last: Hadlock

first: Jacob

last: Neice


title: Transient ER stress cell-autonomously promotes beta cell cycling

city: Brussels

first: Daliya

last: Kancheva

address: Laarbeeklaan 103, Building E

institute: Vrije Universiteit Brussel - VIB

summary: Regenerating endogenous pancreatic ß-cells is a potentially curative yet currently elusive strategy for diabetes therapy. Mimicking the microenvironment of the developing pancreas and leveraging vascular signals that support pancreatic endocrinogenesis may promote ß-cell regeneration. We aimed to investigate whether recovery from experimental hypovascularization of the endocrine pancreas, achieved by modulating the transgenic production of a VEGF-A blocker in ß-cells, could trigger mouse ß-cell proliferation. Serendipitously, we found that transgene overexpression in ß-cells induces endoplasmic reticulum (ER) stress and that subsequent relief from this stress stimulates ß-cell proliferation independent of vessel recovery. Transient GFP overexpression in vivo and transient chemical induction of ER stress in vitro replicated this ß-cell cycling response. Our findings highlight the potential side effects of ER stress due to transgene overexpression in ß-cells and assert that ER stress relief serves as a potent regenerative stimulus.

overall_design: Pancreatic islets were isolated from RIP-rtTA;TetO-sFLT1 mice and subsequently dissociated. For each condition (No DOX, +DOX, and WD4D), two biological replicates were included, resulting in a total of six samples that were analyzed using scRNAseq.

first: Stephanie

last: Bourgeois

first: Annelore

last: Van Mulders

first: Yves

last: Heremans

first: Gunter

last: Leuckx

first: Lien

last: Willems

first: Sophie

last: Coenen

first: Laure

last: Degroote

first: Julie

last: Pierreux

first: Isabelle

last: Scheyltjens

first: Kiavash

last: Movahedi

first: Miriam

last: Cnop

first: Xiaoyan

last: Yi

first: Chiara

last: Vinci

first: Yue

last: Tong

first: Eelco

last: de Koning

first: Françoise

last: Carlotti

first: Heimberg

last: Harry

first: Nico

last: De Leu

first: Staels

last: Willem


title: LOXL2 identifies inflammation-associated myofibroblasts predicting kidney allograft dysfunction and fibrosis

city: Münster

first: Carolin

last: Walter

address: Domagkstraße 9

department: Medical Faculty of the WWU Münster

summary: Chronic allograft nephropathy (CAN) is a major obstacle in kidney transplantation, often leading to long-term graft failure. Early identification of patients at high risk for CAN could be instrumental to improve outcomes. Here, we investigate Lysyl oxidase ligand 2 (LOXL2) as a predictive biomarker for CAN. Using single cell RNA sequencing of transplant biopsies, immunohistochemistry and immunofluorescence microscopy, we find that LOXL2 labels a distinct intertubular myofibroblast subtype with a smooth muscle actin (SMA)-, CXCL12- and PDGFRb-negative and CD68-positive phenotype. These cells are present in non-fibrotic and fibrotic regions utilizing collagen 3 as scaffold, and are associated with a high expression of extracellular matrix (ECM) proteins and related pathways. Native kidneys also harbor LOXL2+ myofibroblasts, albeit at much lower levels, and their presence correlates with the incidence of chronic kidney disease. Following transplant surgery, LOXL2+ cells can rapidly emerge within days, particularly during episodes of rejection, where they associate with leukocyte aggregates. Notably, elevated cell numbers can be normalized as shown in follow up biopsies. A retrospective analysis of 118 biopsies revealed a significant association between LOXL2+ cell abundance and transplant fibrosis, inflammation, and kidney function. However, no correlation was found with transplant age, rejection type and other Banff parameters. Non-rejecting allografts displayed high variability in the number of LOXL2+ cells, with high abundance serving as a long-term predictor of fibrosis and reduced allograft function. Collectively, our findings suggest that LOXL2+ intertubular cells are a subset of inflammation-associated SMA- myofibroblasts and may be useful as a risk biomarker for early fibrogenesis in non-rejecting allografts.

overall_design: Renal graft biopsies of KTx patients were analysed in this study. The indication for graft biopsy was unexplained graft injury as assessed by creatinine or proteinuria. Per patient, 1 biopsy was analysed. In total, 18 patients were included in this study.

first: Konrad

last: Buscher

first: Rebecca

last: Rixen

first: Paula

last: Schütz


title: Spatially distinct cellular and molecular landscapes define prognosis in triple negative breast cancer [2]

city: Pasadena

first: David

middle: Eugene

last: Frankhouser

address: 1500 E Duarte Rd

department: Population Sciences

institute: City of Hope

summary: Triple-negative breast cancer is a prevalent breast cancer subtype with the lowest 5-year survival. Several factors contribute to its treatment response, but the inherent molecular and cellular tumor heterogeneity are increasingly acknowledged as crucial determinants. Here we report on the spatial and molecular heterogeneity underlying a retrospective, treatment-naïve group of women with differential prognoses (17 with >15 years survival- good prognosis (GPx) and 15 with <3 years survival-poor prognosis (PPx)) profiled using GeoMX® DSP. Analyses revealed that the state of epithelia and its microenvironment (TME) are transcriptionally distinct between the two groups. Invasive epithelia in GPx shows a significant increase in immune transcripts, with the TME exhibiting increased immune cell presence (via IF). PPx epithelia, in contrast, are more metabolically and translationally active, with the TME being more mesenchymal/fibrotic. Notably, pre-cancerous epithelia in PPx display a prescience of aggressiveness, as evidenced by increased EMT-signaling. We identify distinct epithelial gene signatures for PPx and GPx, that can, with high accuracy, classify samples at the time of diagnosis and are likely to inform therapy.

sample_organism:
Homo sapiens
synthetic construct

platform_organism:
Homo sapiens
synthetic construct

overall_design: GeoMx human whole transcriptome atlas was performed on ROIs that were selected from 32 FFPE sections from good prognosis (GPx) and poor prognosis (PPx) triple negative breast cancer samples.

first: Kavitha

last: Mukund

first: Darya

last: Veraksa

first: Lixin

last: Yang

first: Jerneja

last: Tomsic

first: Raju

last: Pillai

first: Srijan

last: Atti

first: Zahra

last: Mesrizadeh

first: Daniel

last: Schmolze

first: Xio-Cheng

last: Wu

first: Mary-Anne

last: LeBlanc

first: Lucio

last: Miele

first: Augusto

last: Ochoa

first: Victoria

last: Seewaldt

first: Shankar

last: Subramaniam


title: Enhanced non-enzymatic H2S generation extends lifespan and healthspan in male mice

city: Seville

first: Alejandro

last: Martin-Montalvo

address: Av/ Américo Vespucio 24

institute: CABIMER

summary: Hydrogen sulfide is a gasotransmitter with biological functions, including roles in antioxidant defenses, mitochondrial bioenergetics and cellular signaling via cysteine persulfidation. Several longevity-promoting interventions enhance endogenous hydrogen sulfide generation. However, whether enhanced hydrogen sulfide generation extends healthspan and lifespan in mammals remains unknown. Here, we investigated the in vivo effects of the non-enzymatic hydrogen sulfide generation promoted by natural diallyl sulforated compounds. Diallyl sulforated compounds extended lifespan and improved the main aspects of healthspan, including glucoregulation, locomotor function and neurocognition in wild type male mice across their lifespan. At histological and molecular levels, we observed reductions in hepatic lipid-droplet size, attenuation of transcriptional and proteomic signatures associated with mTOR and immune-related pathways, and increased cysteine persulfidation in proteins. In humans, greater protein persulfidation in individuals with polypathological conditions was associated with increased muscle strength and lower triglyceride levels, supporting its physiological relevance. Our findings uncover the potential of enhanced hydrogen sulfide generation to promote healthy aging.

overall_design: Mice were fed with a standard healthy diet, a high fat diet, a high sucrose diet or a high fat high sucrose diet. Mice were treated with DAS, DAD or DAT for 3 hours or 18 weeks. Liver tissue was harvested and processed for RNA-seq.

first: María Ángeles

last: Cáliz-Molina

first: Raúl

last: López-Fernández Sobrino

first: Inmaculada

last: Pino-Pérez

first: Concepción

last: Panadero-Morón

first: Maria del Carmen

last: Vilches-Pérez

first: María

last: Camacho-Cabrera

first: Almudena

last: García-Ruíz

first: Leopoldo

last: Pérez-Rosendo

first: Isabel

last: Espadas

last: Sola-García

first: Mario

last: Soriano-Navarro

first: Franz

last: Martin

first: Mónica

last: Venegas-Calerón

first: Joaquin José

last: Salas

first: Enrique

last: Marínez-Force

first: Luis

middle: C

last: Romero

first: Ángeles

last: Aroca

first: Román

last: González-Prieto

first: Máximo

last: Bernabeu-Wittel

first: Vivian

last: Capilla-González

first: Christopher

last: Hine

last: Martín-Montalvo