title: Transcriptomic profiling of MCF7 and ZR-75-1 parental and RB1 knockout cells treated with palbociclib

city: Melbourne

first: Shom

last: Goel

state: Victoria

address: 305 Grattan St

country: Australia

institute: Peter MacCallum Cancer Centre

summary: The retinoblastoma protein (Rb) is a tumour suppressor best known for repressing E2F transcription factors and halting cell cycle progression. In hormone receptor-positive (HR+) breast cancer, CDK4/6 inhibitors activate Rb by preventing its phosphorylation, forming a key component of current endocrine therapy regimens. How pharmacologically activated Rb remodels chromatin and influences transcription beyond cell cycle arrest remains poorly understood. Here we show that CDK4/6 inhibition induces redistribution of hypo-phosphorylated Rb to promoters and enhancers. While Rb predictably binds to cell cycle gene promoters to repress transcription, at other sites it unexpectedly promotes expression of oestrogen-responsive genes by integrating into oestrogen receptor (ER)-rich transcriptional hubs. CDK4/6 inhibition enhances ER target gene expression in breast cancer cells, patient-derived xenografts, and clinical HR+ breast cancer samples in an Rb-dependent manner. This reprogramming is mediated in part by KDM5A, whose interaction with Rb contributes to gene regulation at these loci. Critically, components of this Rb-driven ER transcriptional program are pro-proliferative. In endocrine-sensitive tumours, this can be neutralised with anti-oestrogen therapy, explaining therapeutic synergy. In endocrine-resistant settings such as ESR1-mutant breast cancer, the program persists, limiting therapeutic efficacy. These findings reframe Rb as a dual-function transcriptional regulator that, while enforcing cell cycle arrest, can also activate programs that counteract its tumour suppressor function.

overall_design: 3'RNA-seq profiling of MCF7 and ZR-75-1 parental cells and isogenic RB1 knockout cells treated with DMSO control or palbociclib 500 nM. Three biological replicates per group.

first: Antonio

last: Ahn

first: April

last: Watt

first: Catherine

last: Blyth


title: Degenerate long-range interactions drive phase transition of heterochromatin [ChIP-Seq]

status: Public on Jun 30 2026

city: Toulouse

first: Olivier

last: Cuvier

address: 118 route de Narbonne

department: LBME

country: France

institute: CNRS

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Despite intensive efforts, the molecular mechanisms that organize genomes into A and B compartments are still not understood. This is largely due to experimental challenges in targeted manipulation of the molecular components involved. We made use of C. elegans as elementary model system to unveil fundamental concepts of genome compartmentalization. Simultaneous deletion of the H3 lysine 9 methylation-binding proteins HPL-2, a heterochromatin protein 1- type factor, and LIN-61, a malignant brain tudor-domain factor, expression of a dominant negative HPL-2 mutant, or abolishment of H3 lysine 9 methylation resulted in near-complete loss of A/B compartmentalization. Compartmentless genomes follow an equilibrium model of organization with transitions of compartment identity having only moderate effects on local contact domains or on gene expression. We identify degenerate long-range interactions that scalewith the high combined genomic density of HPL-2, LIN-61 and H3 lysine 9 methylation to drive heterochromatin segregation. Using simulations of a minimal polymer system, we show that such interactions are sufficient for the establishment of B compartments and for the phase transition that drives the spatial segregation of heterochromatin from euchromatin. Our findings define genomic, molecular, and mechanistic determinants that are essential for comprehending the role of genome compartmentalization and 3D organization in health and disease.

overall_design: Crosslinking of C. elegans embryos and nuclei preparation followed a protocol described previously (Crane et al., 2015). Gravid adult worms were collected and bleached to obtain late-stage embryo pellets of at least 1 mL or 4 mL for ChIP-seq or Hi-C experiments, respectively. Embryos were washed twice in 1xPBS, flash-frozen in liquid nitrogen, and stored at -80°C. Thawed frozen embryos were crosslinked in 50 mL of 2% formaldehyde (v/v) in PBS for 30 minutes at room temperature on a nutator. After crosslinking, embryos underwent sequential washes in 100 mM Tris-HCl, pH 7.5, 1xPBS, and lysis buffer (10 mM Tris-HCl, pH 8.0; 10 mM NaCl; 0.2% Igepal CA-630; 5 mM DTT; 0.5 mM EGTA; 1x protease inhibitor (cOmplete™)). Embryos were dounced on ice with pestle A and B, followed by centrifugation at 100 xg at 4°C. The supernatant containing nuclei was collected and the extraction process was repeated up to six times, confirming the presence of DAPI-stained nuclei in the supernatants under fluorescence microscopy. Pure nuclei-containing supernatants were pooled and centrifuged at 2000 xg for 5 minutes at 4°C to collect material for subsequent experiments. ChIP was performed with minor modifications (Ercan et al., 2007). After crosslinking, C. elegans embryos were washed with 100 mM Tris-HCl, pH 7.5, followed by two washes with FA buffer (50 mM HEPES-KOH, pH 7.5; 1 mM EDTA; 1% Triton X-100; 0.1% sodium deoxycholate; 150 mM NaCl; 1x protease inhibitor (cOmplete™)). Nuclei were sheared as for Hi-C and centrifuged at 13,000 xg for 15 minutes at 4°C to collect extracts. Immunoprecipitations were set up with extracts equivalent to 3 mg and 3–5 μg of antibodies in 500 μL FA buffer. Input controls were set aside before antibody addition. After 16 hours of incubation at 65°C with rocking, complexes were recovered using Protein A/G magnetic beads and washed with FA buffer supplemented to 1 M NaCl and then to 500 mM NaCl. Beads were eluted with 1% SDS in TE with 250 mM NaCl at 65°C for 30 minutes. Samples and inputs were treated with Proteinase K and decrosslinked overnight at 65°C. DNA was purified using a PCR clean-up kit, quantified with Qubit, and libraries were prepared with NEBNext® Ultra™ II DNA Library Prep Kit for Illumina®. Libraries were sequenced on Novaseq 6000 PE150 S4 flowcells. Antibodies used for ChIP experiments included anti-HPL-2, anti-LIN-61, anti-LEM-2, anti-H3K9me2, anti-H3K9me3, anti-H3K4me1, anti-H3K4me3, anti-H3K27me3, and anti-H3K27ac. All ChIP-seq experiments were conducted as biological independent triplicates.

first: Yogesh

last: Ostwal

first: Fernando

last: Muzzopappa

first: Alexandre

last: Heurteau

first: Raoul

last: Raffel

first: Alaguraj

last: Veluchamy

first: Karthik

last: Eswara

first: Shakila

last: Ali

first: Reema

last: Kumari

first: Ahlem

last: Boudellioua

first: David

last: Depierre

first: Satya

last: Vadlamani

first: Nicolas

last: Chanard

first: Stephane

last: Schaak

first: Fabian

last: Erdel

first: Wolfgang

last: Fischle


title: Transcriptomic profiling of ZR-75-1 wild-type and RB1 knockout cells treated with abemaciclib

status: Public on Jun 29 2026

first: Shom

last: Goel

type: Expression profiling by high throughput sequencing

summary: The retinoblastoma protein (Rb) is a tumour suppressor best known for repressing E2F transcription factors and halting cell cycle progression. In hormone receptor-positive breast cancer, CDK4/6 inhibitors activate Rb by preventing its phosphorylation, forming a key component of current endocrine therapy regimens. How pharmacologically activated Rb remodels chromatin and influences transcriptional networks beyond cell cycle arrest remains poorly understood. Here, we validated the specificity of the Rb antibody for CUT&RUN, and measured changes in Rb chromatin binding after treatment with abemaciclib. We show that CDK4/6 inhibition induces widespread redistribution of Rb to both promoters and enhancers in cancer cells. While Rb predictably binds to cell cycle gene promoters where it represses transcription, at other sites it unexpectedly promotes expression of oestrogen-responsive genes by integrating into oestrogen receptor (ER)-rich transcriptional hubs. This reprogramming is mediated in part by KDM5A and KDM5B, whose functional interaction with Rb contributes to transcriptional regulation at these loci. In patient-derived models and clinical samples of endocrine-sensitive breast cancer, CDK4/6 inhibition enhances ER target gene expression, sensitising tumours to anti-oestrogen therapy. However, in endocrine-resistant contexts – such as ESR1-mutant breast cancer – this Rb-driven oestrogenic programme serves to limit therapeutic efficacy. These findings reframe Rb as a dual-function transcriptional regulator – one that enforces cell cycle arrest while driving transcriptional programs with important therapeutic implications. This expanded model of Rb function provides a new mechanistic basis for the synergy between CDK4/6 inhibitors and endocrine therapy, and offers insight into resistance mechanisms in hormone receptor-positive breast cancer.

overall_design: Isogenic ZR-75-1 clones were derived from CRISPR using a non-targeting sgRNA or an sgRNA against RB1 in bulk ZR-75-1 cells. RNA-seq of ZR-75-1 cells with isogenic RB1 knockout or control genotype following treatment with DMSO or 500 nM abemaciclib for 5 days. Three biological replicates were analysed per condition.

first: Antonio

last: Ahn

first: Keefe

last: Chan


title: PHOTOTROPIN-mediated blue light signaling orients the asymmetry of Marchantia polymorpha spores

city: Rehovot

first: Zohar

last: Meir

state: Shfela

address: Herzl 234

department: Mathematics & CS

country: Israel

institute: Weizmann Institute of Science

summary: Multicellular organisms produced by sexual reproduction develop from single cells and the asymmetry of these cells can define the orientation of the earliest developmental axes. The haploid multicellular stage of the plant, Marchantia polymorpha, develops from a single cell – the spore – that divides asymmetrically, producing an apical germ cell that generates the plant body and a smaller basal cell that differentiates as an anchoring germ rhizoid cell. We show that the orientation of this asymmetric cell division is controlled by an external, environmental cue – blue light – that is perceived by the photoreceptor PHOTOTROPIN and signals in an NCH1-dependent manner. This defines core elements of the mechanism by which a directional environmental signal orients cell division, which in turn orients the first axis of symmetry.

overall_design:
To assess gene expression during their early development, Marchantia polymorpha spores were harvested in two biological replicates at each one of three timepoints: 0, 15, and 30 hours after plating on standard medium plates, half-strength B5 Gamborg with 0.5 g/L MES hydrate, 1% sucrose, and 1% plant agar, adjusted to pH 5.5. Plates were covered with cellophane. For each sample, all spores from six wild type sporangia (derived from a cross of Tak-1 and Tak-2) were collected, and surface-sterilized with 1% NaDCC, followed by washing three times with sterile water. Spore collected at the first time point (0 h) were not plated, but sterilized upon their collection and prior to RNA extraction, in order to match culture conditions with other samples. The spore suspensions of 15h and 30h time points were plated and maintained under continuous white light (PFD 50-60 µmol m⁻² s⁻¹) for the indicated durations. For collection, the cellophane with adhered spores was peeled from the medium, flash-frozen in liquid nitrogen, and ground to a fine powder using a mixer mill (RETSCH MM 400) at 30 Hz for 4 minutes. Total RNA was extracted using the RNeasy Plant Mini Kit (Qiagen) according to the manufacturer’s instructions, followed by Turbo DNase treatment to remove residual genomic DNA.. Differential expression analysis was conducted using DESeq2. Raw sequencing data, as well as Salmon alignment and DESeq2 outputs are uploaded here.

Please note that the 'Transcriptiome of 0 hour Marchantia spores, 2nd biological replicate, lane 5' sample raw data has been lost and thus only processed data is provided.

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Submitter states that missing raw files (GSM9841686) are due to file loss.
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first: Radka

last: Slovak

first: Johannes

last: Rötzer

first: Liam

last: Dolan


title: Transcriptomic profiling of MCF7 ESR1 Y537S mutant cells treated with abemaciclib and/or fulvestrant

overall_design: 3'RNA-seq of MCF7 ESR1 Y537S mutant cells treated with DMSO control, abemaciclib, fulvestrant, or a combination of abemaciclib and fulvestrant. Two biological replicates per group.

first: April

last: Watt

first: Rhiannon

last: Coulson


title: RNA-seq analysis of CAMA-1 cells treated with abemaciclib and/or fulvestrant

overall_design: 3'RNA-seq profiling of CAMA-1 cells treated with DMSO control, abemaciclib, fulvestrant, or a combination of abemaciclib and fulvestrant. Three biological replicates per group.


title: Rb DynaTag profiling of estrogen receptor-positive patient-derived xenograft tumours following treatment with abemaciclib or vehicle control

overall_design: PDX KCC-P-3837 estrogen receptor-positive patient-derived xenograft tumours were grown in NSG mice and treated in vivo with either abemaciclib or vehicle control for 4 days. Tumours were harvested following treatment and immediately subjected to Rb DynaTag chromatin profiling. Two Rb biological replicates per treatment group and four matched IgG controls were sequenced.

first: Michael

last: Taylor

first: Catherine

last: Dietrich

first: Christabella

last: Mahendra


title: Dissecting the radio-protection of hemp seed oil on splenic lymphocytes at single-cell level

city: Beijing

first: Zhijie

last: Bai

state: 北京Beijing

address: Taiping Road, Haidian District

country: China

institute: beijing institute of radiation

summary: In summary, we illustrated that HSO can increase the survival of mice and protects splenic T cells under acute irradiation. The exclusive protection of T cells induced by HSO involved multiple pathways including alleviate oxidative stress, suppressing apoptotic pathways, upregulating anti-inflammatory functions and regulating cell cycle or proliferation. These findings offer vital supports for application of HSO as a prophylactic medicine not only for surviving acute irradiation but also for improving the living quality after exposure.

overall_design: The cells of spleen from Control (mice experiencing no irradiation), IR (mice irradiated at 8.5 Gy and administered with colvent control) and IR+HSO (mice irradiated at 8.5 Gy and administered with HSO) groups were subjected to 10x Genomics scRNA-seq.


title: The risk SNP rs11067228 promotes prostate cancer drug resistance and neuroendocrine differentiation through 3D chromatin interactions (ChIP-seq data set)

city: Tianjin Shi

first: Yuan

last: Jiang

state: Tianjin Shi

address: No. 94 Weijin Road, Nankai District, Tianjin

institute: Nankai University

summary: We performed ChIP-seq in 22Rv1, C4-2B, LNCaP and RWPE1 cells to determine the binding of H3K27ac.

overall_design: Chromatin immunoprecipitation followed by next-generation sequencing (ChIP-seq) in 22Rv1, C4-2B, LNCaP and RWPE1 cells for the active histone mark H3K27ac.


title: Noninvasive Salivary MicroRNA Signatures as Potential Prognostic Biomarkers for Bronchopulmonary Dysplasia in Extremely Preterm Infants: A Pilot Study

city: Almaty

first: Iskander

last: Isgandarov

address: Timiryazev 45

country: Kazakhstan

institute: Institute of plant biology and biotechnology

summary:
Bronchopulmonary dysplasia (BPD) is a major complication of preterm birth, with lasting effects on respiratory outcomes in extremely low birth weight (ELBW) and very low birth weight (VLBW) infants. This pilot study aimed to identify non-invasive salivary microRNA biomarkers for early BPD risk stratification in preterm neonates.
Saliva samples were collected from 20 preterm infants (BPD+: n=10; BPD-: n=10) at days 10-14 of life. Total RNA including the miRNA fraction was isolated from saliva using the MagMAX mirVana Total RNA Isolation Kit. RNA was profiled using the Affymetrix GeneChip miRNA 4.1 Array (GeneTitan MC instrument, 96-well plate format). RMA normalization and limma differential expression identified three candidate salivary miRNA biomarkers (hsa-let-7b-5p, hsa-miR-4454, hsa-let-7c-5p), all significantly upregulated in BPD infants.

overall_design: Case-control study. Saliva samples were collected from preterm newborns (gestational age ≤36 weeks) at days 10-14 of life. Cases (n=10): infants diagnosed with BPD. Controls (n=10): preterm infants without BPD. RNA was hybridized to Affymetrix GeneChip miRNA 4.1 Array. RMA normalization applied (background=FALSE). Differential expression assessed with limma (v3.66.0), adjusted for gestational age and birth weight.

first: Arailym

last: Abilbayeva

first: Iskandar


title: Spatial transcriptomics reveals differences in bronchus-associated lymphoid tissue composition in stable and rejecting human lung allografts

city: St. Louis

first: Daniel

last: Kreisel

state: Missouri

address: 660 Euclid Ave

country: USA

institute: Washington University School of Medicine

summary: Background: Lung allograft rejection is determined by pathologist grading of transbronchial biopsies (TBBx) according to criteria established by the International Society of Heart and Lung Transplantation (ISHLT).  Lymphocytic bronchiolitis (LB), described as mononuclear cells in bronchiolar submucosa, exhibits significant histologic overlap with bronchus-associated lymphoid tissue (BALT) leading to diagnostic uncertainty. Additionally, the role of BALT in tolerance and rejection is debated. We sought to characterize lymphoid aggregates in ISHLT  acute cellular rejection (ACR) grades A0 or A3 biopsies to better understand the spectrum of BALT and rejection lesions. Methods: TBBx were reviewed for the presence of BALT. Representative A0 and A3 biopsies were selected for spatial transcriptomics and multiplex immunofluorescence microscopy. Results: Spatial transcriptomics of allograft biopsies enabled unbiased identification of BALT and rejection lesions via neighborhood analysis. CXCL9/10+ T and PDL1/L2+ myeloid cell clusters were enriched in A0 BALT, while an NKG7+CD8+ T cell cluster was enriched in A3 BALT.  Higher expression of CXCL9 and CXCL10 in the BALT correlated with A0 grade.  Computational methods identified occult areas of lymphoid aggregates in rejecting lung parenchyma with similarity to rejection lesions.  A3 BALT and rejection lesions showed substantial similarities.  Immunofluorescence confirmed the key transcriptomics findings. Conclusions: This study suggests a relationship between the immune microenvironment in BALT and graft rejection.  Additionally, lymphoid infiltration in allografts graded A3 may be more widespread than apparent on Hematoxylin and Eosin (H&E) stain.

overall_design: TBBx from 2 patients graded A0 and 2 patients graded A3 were analyzed by Xenium

first: Alexander

last: Wein

first: Changxu

last: Fan

first: Vladimir

last: Sukhov

first: Junedh

last: Amrute

first: Jon

last: Ritter

first: Amit

last: Bery

first: Maxim

last: Artyomov

last: Krupnick

first: Ruben

last: Nava

first: Andrew

last: Gelman

first: Kory

last: Lavine


title: Effect of full-length laminin 332 treatment on the transcriptome of mouse keratinocytes that express or lack integrin α3β1

city: Albany

first: Whitney

last: Longmate

state: New York

address: 43 New Scotland Avenue

department: Surgery-General Surgery

institute: Albany Medical College

summary: Basement membrane protein Laminin-332 (LN332) is upregulated after injury and promotes keratinocyte migration. It functions as a key ligand for integrin α3β1, which is highly expressed by basal keratinocytes and further induced after wounding. Integrin α3β1 has been previously demonstrated to be a major regulator of the keratinocyte transcriptome. Here, we investigated whether topical application of recombinant full-length LN332 wound impact the keratinocyte transcriptome, and whether integrin α3β1 expression would be required for LN332's effect. It is important to note that keratinocytes make and secrete LN332, therefore the genes that respond to LN332 treatment are responding to treatment with additional (exogenous) recombinant LN332 and likely represent a small fraction of the genes that are regulated by the endogenous LN332-α3β1 axis.

overall_design: Immortalized mouse keratinocytes (MK cells) that lack α3 integrin (MKα3-, null), and MK cells that express α3 integrin (MKα3+; MK-/- cells reconstituted with human α3 integrin, rescued) were cultured on collagen-coated culture dishes in supplemented low-calcium Minimum Essential Medium (MEM; Thermo Fisher Scientific) and incubated at 33°C with 8% CO2. For LN332 treatments, plated MKα3- (null) and MKα3+ (rescued) cells were incubated in MK culture medium supplemented with 0.02 mg/mL LN332 (BioLamina) or equal volume of vehicle control (PBS/10% Glycine/0.02 NaN3) for 16-24 hours.  Cells were pelleted for RNA extraction, library preparation, and RNA-Seq.

first: Salima

last: Lalani


title: Human HEMK1 methylates all mitochondrial release factors and is required for mitochondrial function under stress conditions

city: Stuttgart

first: Albert

last: Jeltsch

address: Allmandring 31

department: Institute of Biochemistry and Technical Biochemistry

country: Germany

institute: University Stuttgart

summary: HEMK1 is the mitochondrial homolog of HEMK2, the enzyme that methylates the conserved glutamine in the GGQ loop of eukaryotic translation termination factor eRF1. Previous work reported altered GGQ-loop glutamine methylation of mitochondrial release factors (MRF) after HEMK1 deletion, but direct biochemical evidence for HEMK1 catalytic activity and the functional consequences of MRF methylation for mitochondrial translation and physiology remained unclear. We generated HEMK1 knockout HEK293 cells and analyzed their transcriptome and proteome but detected only limited effects on the global cellular transcriptome. In contrast, we observed that many mitochondrially encoded transcripts were consistently reduced. The reduction in mitochondrial transcripts may be a secondary consequence of impaired mitochondrial translation leading to mRNA destabilization.

overall_design: RNA-seq profiling of HEMK1 KO HEK293 cells and their WT counterparts

first: Jana

last: Kehl

first: Thyagarajan

middle: T

last: Chandrasekaran


title: Single-cell RNA sequencing reveals endothelial STING-dependent immune remodeling in murine metabolic dysfunction-associated steatohepatitis

city: Xi'an

first: Lin

last: Wang

state: Shaanxi

address: West Changle Road 169

institute: Xijing Hospital, Air Force Medical University

summary: Liver sinusoidal endothelial cells (LSECs) regulate nutrient flux and immune surveillance within the hepatic niche. Here, we investigate how endothelial-intrinsic cGAS-STING signaling integrates metabolic stress signals to reshape adaptive immune responses in metabolic dysfunction-associated steatohepatitis (MASH) at single-cell resolution. We performed single-cell RNA sequencing on liver tissues from endothelial cell-specific Sting knockout mice (Stingfl/fl Cdh5-Cre+) and littermate controls (Stingfl/fl Cdh5-Cre-) subjected to a western diet combined with CCl4 (WD-CCl4) for 12 weeks. Unbiased cell-type annotation revealed broad immune remodeling following LSEC-STING deletion. Among CD4+ T cells, LSEC-STING deletion resulted in a marked reduction of pathological Th17 cells, accompanied by expansion of regulatory T cells (Tregs), leading to a significantly increased Treg/Th17 ratio. LSEC-STING deletion also reduced exhausted CD8+ T cells while increasing effector CD8+ T cells. Mechanistically, LSEC-intrinsic STING signaling represses BMP4 transcription through NF-kB-mediated competition with AP-1, disrupting a tolerance-supporting angiocrine program. These findings identify endothelial STING as a vascular immunometabolic checkpoint that links chronic metabolic stress to adaptive immune remodeling in MASH.

overall_design: Endothelial cell-specific Sting knockout mice (Stingfl/fl Cdh5-Cre+) and littermate controls (Stingfl/fl Cdh5-Cre-) were subjected to WD-CCl4 for 12 weeks to induce MASH. Liver tissues were dissociated into single-cell suspensions and subjected to 10X Genomics Chromium 3' single-cell RNA sequencing. Three biological replicates were included per group.

first: Zhi-Bin

last: Lin

first: Peng

last: Zou

first: Xian-Yi

last: Ma

first: Jun-Bo

last: Song

first: Hong

last: Zhang

first: Wei

last: Du

first: Dan

last: Wei

first: Ping

first: Xin

last: Hong

first: Jing-Jing

last: Liu

first: Zhi-Qiang

last: Fang

first: Hao

last: Xu

first: Fei

last: He

first: Juan-Li

last: Duan

first: Ke-Feng

last: Dou


title: Bulk RNA sequencing reveals endothelial STING-dependent transcriptional programs in murine metabolic dysfunction-associated steatohepatitis

summary: Liver sinusoidal endothelial cells (LSECs) regulate nutrient flux and immune surveillance within the hepatic niche. Here, we investigate how endothelial-intrinsic cGAS-STING signaling integrates metabolic stress signals to reshape the hepatic niche in metabolic dysfunction-associated steatohepatitis (MASH). We performed bulk RNA sequencing on liver tissues from endothelial cell-specific Sting knockout mice (Stingfl/fl Cdh5-Cre+) and littermate controls (Stingfl/fl Cdh5-Cre-) subjected to a choline-deficient, L-amino acid-defined, high-fat diet (CDAHFD) for 10 weeks. Gene set enrichment analysis revealed that multiple cancer-associated and fibrogenic signaling pathways were enriched in control livers but markedly suppressed in StingΔEC livers. Conversely, pathways related to T cell regulation and metabolism were preferentially enriched in StingΔEC livers, indicating that endothelial-intrinsic STING signaling drives MASH progression by reshaping T cell-mediated immune responses.

overall_design: Endothelial cell-specific Sting knockout mice (Stingfl/fl Cdh5-Cre+) and littermate controls (Stingfl/fl Cdh5-Cre-) were generated and subjected to CDAHFD for 10 weeks to induce MASH. Total RNA was extracted from liver tissues and subjected to paired-end bulk RNA sequencing. Three biological replicates were included per group.


title: Comprehensive spatial profiling of the spleen in human myelofibrosis [snRNAseq]

city: Pessac

first: Bérénice

last: Dugué

address: 1 Av. Magellan

institute: INSERM

summary: Splenomegaly is a defining feature of myelofibrosis, yet the contribution of splenic mesenchymal stroma to disease progression remains unclear. We combined spatial and single-nucleus transcriptomics of patient spleens to map extramedullary hematopoiesis niches. Activated red pulp reticular cells localize near hematopoietic stem and progenitor cells, and early disease shows marginal zone disruption with lymphoid depletion preceding stromal remodeling. Macrophage- and megakaryocyte-derived signals activate complement and induce TNFα, TGF-β, extracellular matrix, and Thbs1 programs in reticular cells.

overall_design: Visium Spatial Transcriptomics and Single-Nuclei profiling of human spleens of patients with myelofibrosis

first: Pia

last: Wanner

first: Mayra

last: Ruiz Tejada Segura

first: Mohamed

last: Saad

first: Lucas

last: Greven

first: Katrin

last: Gotz

first: Carmen

last: Schalla

first: Stijn

last: Fuchs

first: Gerjanne

last: Vroeg in de wei

first: Hélène

last: Gleitz

first: Anna Katharina

last: Galyga

first: Jessica

last: Ellen Pritchard

first: Tobias

last: Schlangen

first: Niklas

last: Lutterbach

first: Salim

last: Atakhanov

first: Lina

last: Schmidt

first: Marie

last: Parrens

first: Armelle

last: Peyrat

first: Damien

last: Luque Paz

first: Marie-Christine

last: Copin

first: Alexandre

last: Guy

first: Victor-Emmanuel

last: Brett

first: Olivier

last: Mansier

first: Susanne

last: Schmitz

first: Paul

last: Wanek

first: Bella

last: Banjanin

first: Stephani

first: Martina

last: Crysandt

first: Marian

last: Clahsen-van Groningen

first: Konnie

last: Hebeda

first: Sascha

last: Dietrich

first: Harald

last: Voehringer

first: Matthias

last: Meyer-Bender

first: Kristin

last: Séré

first: Chloé

last: James

first: Adam

last: Benabid

first: Ivan

last: Costa

first: Charles

last: Dussiau

first: Rebekka

last: Schneider


title: Multiple mechanisms lead to differential loss-of-function effects of pathogenic SARS2 variants: implications for genotype-phenotype analyses II

city: Ann Arbor

first: Christina

last: Del Greco

state: MI

address: 1241 Catherine Street

institute: University of Michigan

summary: Seryl-tRNA synthetase 2 (SARS2) encodes the enzyme responsible for charging tRNA with serine in the mitochondria. SARS2 has been associated with a spectrum of recessive diseases including HUPRA syndrome and progressive spastic paresis (PSP). Previous studies showed that pathogenic SARS2 variants cause decreased tRNA charging; however, the mechanism by which specific variants lead to distinct recessive phenotypes has not been defined. To address this lack of knowledge, we studied an allelic series of pathogenic SARS2 variants for differential effects on mitochondrial function. We generated a SARS2 knockout Hap1 cell line with a second, doxycycline-inducible wild-type copy of SARS2 to support cell growth. We first assessed the effects of ablated SARS2 expression on the transcriptome and on mitochondrial function. We found that, in the absence of SARS2 expression, transcriptional changes indicate activation of cellular stress responses including the integrated stress response. Furthermore, loss of SARS2 resulted in severely depleted oxygen consumption in Seahorse assays. We next individually transduced an allelic series of 11 pathogenic SARS2 variants into this cell model to assess the effects of each variant on mitochondrial function. These efforts revealed compelling variant-dependent effects on oxygen consumption that will be useful for phenotype-genotype correlations. Interestingly, the most commonly detected pathogenic SARS2 variant (R402H) had no effect on mitochondrial function in our model system. We performed computational and functional studies, which showed that this missense variant has a partial effect on exon inclusion, which predicts a hypomorphic allele consistent with certain patients being reported as homozygous for this variant. Overall, this study expands our understanding of SARS2 biology, reveals the differential effects that pathogenic variants have on SARS2 function, and provides the foundation for defining the clinical heterogeneity of patient phenotypes.

overall_design: Modified Hap1 cells containing a deletion in the endogenous SARS2 gene, a doxycycline-inducible FLAG-tagged copy, and a constitutively expressed wild-type or mutant copy of SARS2 were grown without doxycycline for 6 days such that only the constitutively expressed copy of SARS2 was expressed.  Cells were harvested and lysed for submission to the Plasmidsaurus RNA-sequencing service to obtain differential gene expression for cells containing wild-type SARS2 vs those containing a variant copy

first: Stephanie

middle: M

last: Laureano-Figueroa

first: Anthony

last: Antonellis


title: Histidine to glutamine substitutants, a regulated process that impacts cell survival and suggests histidine shortage in cancer

city: Amsterdam

first: Pierre-Rene

last: Körner

address: Plesmanlaan 121

department: Oncogenomics

country: Netherlands

institute: Netherlands Cancer Institute (NKI)

summary: Cancer-associated genetic, epigenetic, and microenvironmental factors impose shortages of tryptophan and arginine, which induce specific codon reassignments (substitutants) in their proteomes. Whether cancers are deprived of other amino acids is unknown. Histidine is an essential amino acid reported to be low in certain tumor types. Therefore, we investigated the potential for a histidine shortage in cancer. Using cultured cancer cells, we pinpointed histidine to glutamine (H>Q) substitutants as the most pronounced proteomic event following histidine deprivation. We then scanned cancer proteomes for H>Q proteins and observed a marked enrichment in pancreatic, uterine, and kidney cancers. Mechanistically, we propose that H>Q is a result of tRNA misalignment, and used tRNA glutamine (tRNA(Gln)) modifications at the wobble position to demonstrate it. We further show that URM1-mediated U34 thiolation boosts H>Q production and influences the survival of cancer cells following histidine deprivation, suggesting a potential role for H>Q. Additional characterization of H>Q substitutants revealed preferred sequences and the maintenance of H>Q host protein expression despite the absence of histidine, indicating cellular regulation and functional consequences. Thus, histidine shortage induces H>Q substitutants, a regulated mistranslation event that pinpoints histidine limitation in cancer and impacts cell survival.

overall_design: RNA sequencing of HT29 cells in control condition, histidine depletion condition, and tryptophan depletion condition

first: Domenica

last: Lovecchio

first: Pierre-René

first: Lorenzo

last: Valcanover

first: Onno

last: Bleijerveld

first: Bart R.J.

last: Blokhuis

first: Abhijeet

last: Pataskar

first: Jasmine

last: Montenegro Navarro

first: Demi

last: Wernaart

first: Liesbeth

last: Hoekman

first: Daniil

last: Anastasopoulos

first: Kim

last: Vrijland

first: Xiaodong

last: Feng

first: Julien

last: Champagne

first: Karin E.

last: de Visser

first: Olaf

last: van Tellingen

first: Frank

last: A. Redegeld

first: Reuven

last: Agami


title: RNA-seq gene expression profile of  SUM159PT breast cancer cell line upon manipulation of two TDMD triggers, NREP and SERPINE1

city: Milano

first: Francesco

last: Nicassio

address: Via Adamello 16

department: Center for genomic science

country: Italy

institute: Istituto Italiano di Tecnologia

summary: We investigate the effect of the perturbation of two transcripts involved in target-dependent microRNA degradation (TDMD): NREP is a trigger for miR-29b; SERPINE1 is a trigger for miR-30. We perturbed the genes and monitored the activity of miR-29 or miR-30  analysing the  expression changes of their predicted target genes by RNA sequencing.

overall_design: NREP was perturbed by CRISPR interference, exploiting a SUM159 dcas9-KRAB cell line infected with different sgRNA targeting NREP. SERPINE1 was perturbed by CRISPR/Cas9 genome editing, producing microdeletions in the 3'UTR of the transcript encopassing miR-30 binding site (MRE_KO clones).

first: Matteo

last: Marzi


title: Molecular signatures of T cells targeting multiple myeloma

city: Cambridge

first: Mirco

middle: Julian

last: Friedrich

state: MA

address: 72 Ames St

institute: Broad Institute

summary: Immunotherapeutic approaches including cell therapy are believed to be the next generation of paradigm-changing treatment options in hematological malignancies. Cell therapy using CAR-T cells or expanded autologous T cells (TILs) has shown promising results in non-solid and solid malignancies. However, acquired resistance to immunotherapy or rapid disease progression represent major clinical challenges in hematological cancers. Transgenic T cell receptors (TCRs) redirect patient-autologous lymphocytes to tumor antigens and can induce regression of refractory solid tumors. However, a bone fide population of myeloma reactive TCRs has yet to be identified. Here, we have established a multiplexed optical barcoding assay that allows for ex vivo profiling of single bone marrow-resident T cells in order to identify sequences and transcriptional signatures of patient-individual tumor-reactive lymphocytes. By integrating antigen specificity and the transcriptional profile of single myeloma-associated T cells using the CDR3 sequence as clone-specific barcode, we found that tumor-reactivity results in a specific gene expression state of bone marrow-resident CD8+ T cells. Non-tumor-reactive clones were either CD4+ or enriched for viral specificities, while myeloma reactive TCRs demonstrated an increased clonality within the overall lymphocyte repertoire. We furthermore describe a subset of CD29 (ITGB1)-expressing effector-memory CD8+ T cells that is associated with response to induction immunochemotherapy. Our results demonstrate antitumor reactivity in a subset of bone marrow-resident T cells against malignant plasma cells and provide the rationale for future personalized TCR-transgenic cell therapy approaches in newly diagnosed multiple myeloma patients. More broadly, the generated resources from this project might contribute to identifying and monitoring tumor reactive T cell responses targeting hematological neoplasias.

overall_design:
Bone marrow resident CD3 cells were isolated by magnetic bead separation and analyzed using 10x Genomics 5' RNA/TCR sequencing.

Submitter states that missing raw data are being made available for controlled access in EGA.
EGA submission identifier will be supplied as soon as available.

middle: J

first: Simon

last: Steiger

first: Niklas

last: Kehl

first: Tim

last: Wagner


title: Genome-wide maps in breast cancer MDA-MB-231 cells with or without siRNA interference of YB1.

city: Nanjing

first: Huilong

last: Li

address: China Pharmaceutical University

institute: China Pharmaceutical University

summary: Analysis of MDA-MB-231 cells following YB1 siRNA interference or not. YB1 regulates various RNA metabolic process in MDA-MB-231 cells.Results provide insight into the role of YB1-involved mechanisms underlying YB1-mediated effects on breast cancer stemness.

overall_design: mRNA profiles of MDA-MB-231 cells with or without YB1 siRNA interference were generated by deep sequencing, in 3 repeats, using Illumina NovaSeq 6000.

first: Lufeng

last: Zheng


title: Human spinal cord organoids revealed cell intercalation as a conserved morphogenic mechanism for lumem formation

city: Barcelona

first: Elisa

last: Martí

address: C/ Baldiri Reixac, 4-8, Torre R, 3era Planta

country: Spain

institute: Institut de Biologia Molecular de Barcelona

summary: Neural tube defects (NTDs) are severe anatomical malformations of the nervous system that occur during embryonic development when the neural tube (NT) fails to form correctly. To gain insights into the biological basis of NTDs, we generated human NT organoids with defined anterior and posterior identities. We achieved this by guiding human pluripotent stem cells towards the neural lineage, resulting in the self-assembly of polarized cells around a single lumen, exhibiting subcellular architecture and dynamics similar to those of the embryonic NT. Through transcriptome profiling, we identified that the posterior organoids possessed a spinal cord identity. To better replicate the in vivo morphogenesis of the secondary NT, we developed a modified culture condition, which revealed that lumen resolution in the spinal cord organoids involves cell intercalation and is dependent on the activity of Yes-associated protein (YAP). We believe that these findings will contribute to our understanding of caudal skin-covered NTDs, which are among the most common birth defects observed in humans.

overall_design: Anterior and porterior hNTorg were generated from hESC using drug guidance. Subsequently, the organoids were processed for RNA extraction and samples were sent to bulk RNA sequencing.

first: Jose

last: Blanco-Ameijeiras

first: Mar

last: García-Valero

first: Yara

last: El Majzoub

first: Elena

last: Rebollo

first: Javier

last: Macho-Rendón

first: Jorge

last: Corbacho

first: Juan

middle: R

last: Martinez-Morales

last: Martí-Gorostiza


title: SENP7 deSUMOylase-governed transcriptional program coordinates sarcomere assembly and is targeted in muscle atrophy

city: hannover

first: arnab

last: nayak

address: carl-neuberg-str 1

institute: MHH

summary: Disorganization of the basic contractile unit of muscle cells i.e., the sarcomeres, leads to suboptimal force generation and is a hallmark of muscle atrophy. Here, we demonstrate that the nuclear role of SENP7 deSUMOylase is pivotal for sarcomere organization. SENP7 expression is temporally upregulated in mature muscle cells and directly regulates transcription of the myosin heavy chain (MyHC-IId) gene. We identified SENP7-dependent deSUMOylation of Flightless-1 (Fli-I) as a signal for Fli-I association with Scaffold attachment factor b1 (Safb1). SENP7 deficiency led to higher Fli-I SUMOylation and lower chromatin residency of Safb1, thus generating transcriptionally incompetent chromatin conformation on MyHC-IId. Consequently, lower expression of MyHC-IId caused sarcomere disorganization and disrupted muscle cell contraction. Remarkably, cachexia signaling impeded the SENP7-governed transcriptional program, leading to muscle atrophy, with profound loss of motor protein MyHC-IId. We propose a SENP7-driven distinct transcription program as paramount for muscle cell function, which was found targeted in cachexia

overall_design: 6 samples in total. 3 biological replicates for each conditions (control/Gl2-siRNA and SENP7 siRNA)

first: Mamta

last: Amrute-Nayak

first: Luis

middle: V

last: Gand

first: Bushra

last: Khan

last: Holler

first: Ekaterini

last: Kefalakes

first: Maike

last: Kosanke

first: Theresia

last: Kraft

first: Arnab

last: Nayak


title: small RNA-seq gene expression profile of breast cancer cell lines upon ZSWIM8 perturbation

first: Francesco

last: Nicassio

type: Non-coding RNA profiling by high throughput sequencing

summary: To investigate the ZSWIM8 sensitive (i.e. under the control of target-dependent microRNA degradation) microRNAs in breast cancer, we established a set of dCAS9-KRAB cell lines in which the target gene has been knocked down by CRISPRi and we profiled microRNAs by small RNA sequencing.

overall_design: Each cell line was engineered with a a doxycycline--inducible dCas9-KRAB construct. Then target-dpendent microRNA degradation mechanism was perturbed by CRISPR interference delivering sgRNAs against the critical effector ZSWIM8.  After activation of CRISPRi, cells were cultivated for 7 passages, total RNA was isolated and miRNA expression analyzed by smallRNA-seq.

first: Matteo

last: Marzi


title: Blockade of avb8 integrin synergizes with checkpoint inhibition to enhance anti-tumor immunity through Cxcl9-dependent augmentation of CD8+ T cell cytotoxicity

city: San Francisco

first: Dean

last: Sheppard

state: CA

address: 555 Mission Bay Boulevard South

institute: University of California, San Francisco

summary: Inhibition of the TGF-b activating integrin, avb8, can dramatically enhance the effectiveness of checkpoint inhibition in multiple checkpoint resistant tumor models, but the mechanisms underlying this dramatic synergy have not been previously explored. In this study, we used single cell RNA sequencing to investigate the mechanisms of synergy between inhibition of avb8 and PD1 in the EMT-6 model, a checkpoint resistant model of triple negative breast cancer. Combination therapy dramatically enhanced interferon gamma (Ifng) expression by CD8+ and CD4+ T cells, activated interferon signaling throughout the tumor microenvironment, specifically increased Cxcl9 secretion by tumor macrophages and enhanced Gzmb expression and tumor cell killing by CD8+ T cells. The protective effects of combination therapy were inhibited by Cxcl9 blockade. These findings suggest that Ifng-mediated induction of Cxcl9 is an important driver of synergy between avb8 and checkpoint inhibition and raise the possibility that Cxcl9 might be useful as an early PD biomarker of effective combination therapy.

overall_design: EMT-6 tumor cell suspensions from mice on day 12 of treatment with either isotype control, anti-PD1, anti-avb8, or combination. Three tumors in each condition were collected.

first: Jasmine

middle: W

last: Larrick

first: Hsiao-Yen

last: Ma

first: Caroline

last: Whitty

first: Andrew

middle: D

last: Luster

first: Xin

last: Ren


title: Mediator kinase inhibition exposes a transcriptional vulnerability and primes AML for venetoclax sensitivity

city: Montreal

first: Guy

last: Sauvageau

state: QC

address: 2950, Chemin de Polytechnique

country: Canada

institute: Institute for Research in Immunology and Cancer

summary: Acute myeloid leukemia (AML) is a heterogeneous and aggressive hematologic malignancy associated with poor survival despite recent therapeutic advances. To identify novel anti-leukemic agents, we performed a high-throughput viability screen of 10,000 small molecules across 56 genetically diverse primary AML samples. Compounds with similar activity profiles were grouped into Compound Correlation Clusters (CCCs), leading to the identification of CCC163, which includes AML874, as a potent and selective hit. Integrated pharmacological and CRISPR-Cas9 approaches identified the Mediator complex as the molecular target of AML874. We demonstrate that AML874 acts as a selective CDK8/CDK19 inhibitor, disrupting Mediator-dependent transcription. FLT3-mutated AML showed increased sensitivity to AML874, suggesting a subtype-specific vulnerability. Notably, CDK8 inhibition synergized with venetoclax in AML cell lines and patient-derived xenograft models, resulting in enhanced anti-leukemic activity in vivo. Together, these findings identify AML874 as a promising therapeutic candidate and highlight CDK8/CDK19 inhibition as a potential strategy in AML.

overall_design: Transcriptomic profiling of OCI-AML1 cells treated either with DMSO, AML874 or SEL120-34A at different time points (8h or 24h)

first: Aitchedji

last: Camille

first: Céline

last: Moison

first: Jean-François

last: Spinella

first: Patrick

last: Gendron


title: Gene expression profile at single cell level of breast cell lines HMLE and BT-549

summary: We explored cell heterogeneity by single cell RNA sequencing in normal (HMLE) and cancer (BT-549) mammary cell lines. HMLE is a non-transformed epithelial cell line that possesses only a very small subpopulation of CD44highCD24low cells with mesenchymal and stem-like properties. Given the rarity of the CD44highCD24low subpopulation (1%), we performed single-cell analysis either on the bulk population or on FACS-sorted CD44highCD24low enriched samples.

overall_design: Cell lines were analyzed by scRNA seq either in bulk (BT-549) or upon Fluorescence Activated Cell Sorting using a strategy to enrich for breast stem cells, based on the combination of the two surface proteins CD44 and CD24 (HMLE).


title: Transcriptomic and lipidomic analysis of the cryptococcal lung granuloma reveals macrophage boundary programs and sphingolipid remodeling [Xenium]

city: Stony Brook

first: Maurizio

last: Del Poeta

state: NY

address: 150 Life Science Building

department: Molecular Genetics and Microbiology

institute: Stony Brook University

summary: Cryptococcosis is a life-threatening fungal disease caused by Cryptococcus neoformans. The immunocompetent response results in the formation of a lung granuloma, which contains the fungal cells. We have previously developed a mouse model of the cryptococcal granuloma that is histological similar to the human lung granuloma. In this study, we characterize the lung granuloma in mice using transcriptomics and lipidomics, elucidating the immune cell types present and the transcriptional landscape of macrophages at the granuloma border. Macrophage subtypes are identified, which form a characteristic boundary of the lung granuloma and were observed to have differentially expressed genes involved in the control of the infection. Lipidomic studies revealed that several sphingolipid-associated genes and their products involved in the regulation of phagocytosis were found to be upregulated and in vivo validation studies confirmed their key role in granuloma formation. Lastly, as the granuloma structure develops, the fungal replication inside the granuloma decreases concomitantly to the decrease of the macrophage activation. These studies identified important immune mechanisms of host protection at the granuloma site. Our results provide mechanistic insight into cryptococcal granuloma maturation and refine current models of host–pathogen interaction, which may open avenues for future immunomodulatory strategies

overall_design: Mice are infected with a mutant strain of Cryptococcus neoformans (Δgcs1), which allows for the granulomatous response. Cells were collected from the granuloma at three time points (30, 60, and 90 days-post-infection), as well as from WT-infected and uninfected controls, and were then analyzed via spatial transciptomics

first: Veronica

last: Soares Brauer

first: Kathryn

last: Takemura

first: Barbara

last: Rosati

first: Marinaldo

last: Pacífico Cavalcanti-Neto

first: Deveney

last: Dasilva

first: Neelu

last: Singh

first: Tyler

last: Normile

first: Gabriella

last: Vaccaro

first: Guiseppe

last: Caso

first: Harpreet

first: Bo

last: Chen

first: Iwao

last: Ojima

first: David

last: McKinnon


title: Mediator kinase inhibition exposes a transcriptional vulnerability and primes AML for venetoclax sensitivity [CRISPR Screen]

first: Guy

last: Sauvageau

overall_design: CRISPR-CAS9 screen in OCI-AML1 cells, gDNA obtain at day 17 (d17) in 10% FBS. Analyses were done using DMSO of the corresponding day-FBS (d17-10)  DMSO day0 (d17-10.2)

first: Simon

last: Fortier

first: Tara

last: MacRae

first: Céline


title: Temporal Uncoupling of Radial Glial Progenitor Cell Lineage Progression in Self-organizing Organoid System – MADM-CloneSeq

city: Klosterneuburg

first: Florian

last: Pauler

address: Am Campus 1

country: Austria

institute: Institute of Science and Technology Austria (IST Austria)

summary: Radial glial progenitors (RGPs) produce all excitatory neurons in the developing cerebral cortex. Mosaic Analysis with Double Markers (MADM)-based lineage tracing in vivo revealed a quantitative framework of RGP lineage progression. Here we established MADM technology in embryonic stem cells to probe RGP lineage progression in self-organizing cortical organoid system. We found that RGPs exhibit high level of equipotency in organoid rather than temporally stereotyped lineage progression as observed in vivo. RGPs in organoids showed increased lineage restriction, diminishing cell-type diversity in clones of cortical projection neurons, despite uniform RGP scRNAseq transcriptional signatures and unitary lineage trajectory. Hence, critical non-cell-autonomous cues absent in self-organizing systems and/or genuine stem cell niche are essential for faithful RGP lineage progression and the generation of cortical cell-type diversity.

overall_design: MADM-CloneSeq of MADM-11GT/TG;Emx1-CreER+/- cerebral organoids induced at D13 and collected at D20

first: Melissa

last: Stouffer

first: Osvaldo

middle: A

last: Miranda

first: Fabrizia

last: Pipicelli

last: Hippenmeyer


title: Temporal Uncoupling of Radial Glial Progenitor Cell Lineage Progression in Self-organizing Organoid System

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

overall_design: Refer to individual Series


title: Stroke drives glioma progression through remodeling of the tumor microenvironment [Spatial Transcriptomics]

city: Houston

first: Qi

last: Te

address: 1250 Moursund st

institute: Baylor College of Medicine

summary: Epidemiological studies suggest increased brain tumor risk in patients with prior stroke. Comparative transcriptomic analyses reveal enrichment of stroke-associated gene signatures in high-grade gliomas; nevertheless, the mechanisms that unify these conditions remains unclear. Here we show that stroke promotes tumor infiltration towards the site of stroke injury in human and mouse glioma models, accompanied by reduced overall survival. Stroke induces remodeling of the tumor microenvironment (TME), including the emergence of a unique population of tumor-associated astrocytes (TAAs) that exhibit reduced Ca2+ activity at the infiltrative edge, and enrichment of tumor-associated microglia and macrophages (TAMs). Chemogenetic stimulation of Ca2+ signaling in TAAs or depletion of TAMs reverses stroke-induced glioma progression. We further identify the sodium–bicarbonate cotransporter Slc4a4 as a key regulator of Ca2+ activity in TAAs and associated TAM recruitment. Collectively, our findings demonstrate that stroke accelerates glioma progression through remodeling of astrocytic and immune compartments of the TME.

overall_design: Mice bearing de novo glioma subject to sham or stroke (PTS) surgery, and brain tissue is collected for spatial transcriptomic analysis.

last: Ye

first: Hari Krishna

last: Yalamanchili

first: Hyun Kyoung

last: Lee


title: Stroke drives glioma progression through remodeling of the tumor microenvironment [scRNA-Seq]

overall_design: Mice bearing de novo glioma subject to sham or stroke (PTS) surgery, and brain tissue is collected for scRNA seq  analysis.

first: Akdes Serin

last: Harmanci

first: Kyuson

last: Yun


title: Targeting Lactic Acid Unleashes IFN’s Potent Broad-Spectrum Antiviral Efficacy and Overcomes Clinical Limitations.

city: wuhan

first: Mi

last: Tian

state: Hubei

address: Bayi Street 229 in Wuchang district

department: College of life science

institute: Wuhan University

summary: Type I interferons exhibit potent broad-spectrum antiviral activity in preclinical studies but suffer from critical clinical limitations: narrow intervention windows for acute infections, suboptimal efficacy, and notable adverse reactions. The discrepancy between their robust preclinical and limited clinical performance remains mechanistically unclear. Herein, integrating clinical samples and multi-level infection models, we demonstrate that IFNs exert antiviral effects only when administered pre-infection. Once infection is established, IFNs are ineffective yet induce prominent adverse effects, with host-derived lactic acid (LAC) as the key mediator: it promotes viral immune evasion, impairs IFN therapeutic efficacy, triggers inflammatory storms, and elicits adverse reactions. Mechanistically, LAC suppresses IFN activity via membrane receptor-mediated SIRT1 upregulation and synergizes with IFNs to hyperactivate NF-κB, initiating cytokine storms and forming an "antiviral failure-inflammatory amplification" feedback loop. Based on this mechanism, we developed a combinatorial therapy of IFNs plus an FDA-approved lactate dehydrogenase inhibitor. This regimen reverses LAC-mediated IFN suppression, mitigates inflammation, and achieves dual "antiviral + anti-inflammatory" benefits. Notably, it retains robust efficacy even in late-stage infections, overcoming IFN monotherapy drawbacks and addressing the core bottleneck restricting IFN clinical application. Our study identifies LAC as a pivotal target for broad-spectrum antiviral development and provides a potential strategy to combat emerging viral pandemics.

overall_design: RNA sequence analysis of PM cells infected with VSV virus for 24 hours and the untreated group.

first: Mingfu

first: Zhixin

last: Liu

first: Ke

last: Lan

first: Siwei

last: Chen

first: Ju

last: Huang

first: Xu

last: Li


title: CDK8 integrates jasmonate signaling with sulfur availability to optimize plant chemical defense

city: East Lansing

first: Gregg

last: Howe

state: Michigan

address: 612 Wilson Street

institute: Michigan State University

summary: Plant immunity draws heavily on carbon-, nitrogen-, and sulfur-based precursors for the deployment of specialized defense compounds. The induced biosynthesis of these compounds is triggered by the jasmonate signaling pathway and is often associated with growth penalties. How plants coordinate growth-defense balance with nutrient availability remains poorly understood. Here, we identify CYCLIN-DEPENDENT KINASE 8 (CDK8) as a transcriptional regulator that inhibits growth and seed yield at high levels of jasmonate defense. Transcript and metabolite profiling demonstrated that CDK8 amplifies jasmonate-triggered immunity, including the deployment of sulfur-containing defense compounds. Under sulfur-replete conditions, CDK8 promoted robust accumulation of sulfur-rich defensin peptides and sulfur-containing metabolites such as camalexin. Strikingly, sulfur limitation suppressed the expression sulfur-expensive defensive peptides while elevating production of camalexin, a sulfur-economical output. Our findings indicate that CDK8 optimizes defense strategies to nutrient availability, highlighting how plants maintain immunity during nutritional stress through metabolic prioritization of resource-efficient defenses.

overall_design: RNA-seq profiling of Columbia-0 and cdk8-1 shoots after 10 days of sulfur-sufficient or -deficient conditions.

first: Qiang

last: Guo

first: Huijia

last: Gong

first: Bailey

last: Kleven

first: Jian

last: Yao

first: Jie

last: Wang

first: Kevin

middle: L

last: Childs

first: Hideki

last: Takahashi


title: Gene expression profiling of HSC-3 oral squamous cell carcinoma cells co-cultured with necrotic cells

city: Niigata

first: Manabu

last: Yamazaki

state: Niigata

address: Gakkocho-dori 2-5274, Chuo-ku

department: Division of Oral Pathology

country: Japan

institute: Niigata University Graduate School of Medical and Dental Sciences

summary: Oral squamous cell carcinoma (OSCC) cells are exposed to necrotic cell–derived factors within the tumor microenvironment.  This dataset describes gene expression profiling of the human OSCC cell line HSC-3 following indirect co-culture with necrotic cells for 16 hours using cell culture inserts with 1 µm pores (NC/ID condition), in comparison with monoculture controls. Microarray analysis was performed to characterize transcriptional responses of HSC-3 cells to soluble factors released from necrotic cells.

overall_design: HSC-3 was cultured either alone (monoculture control) or indirectly co-cultured with necrotic HSC-3 cells using cell culture inserts with 1 µm pores (NC/ID condition).  Necrotic HSC-3 cells were prepared by freeze–thaw treatment using liquid nitrogen followed by thawing in a water bath, and were applied to the insert at twice the number of viable HSC-3 cells (2:1 ratio).  Indirect co-culture was performed for 16 hours, allowing the exchange of soluble factors without direct cell–cell contact.  After co-culture, HSC-3 cells were harvested for total RNA extraction and subjected to microarray-based gene expression analysis.  Gene expression profiles were compared between NC/ID and monoculture conditions.


title: Single-cell transcriptomic profiling of frontal fibrosing alopecia reveals preserved cellular architecture with stress-driven epithelial reprogramming and non-cytotoxic inflammation

city: New York

first: Juan

last: Ruano

address: 1230 York ave

institute: Rockefeller University

summary: Background: Frontal fibrosing alopecia (FFA) is a primary cicatricial alopecia characterised by progressive follicular fibrosis and irreversible hair loss. Despite increasing clinical recognition, the cellular mechanisms sustaining disease persistence remain incompletely defined. Objectives: To define cell-type–specific transcriptional states and inter-compartmental signalling networks in lesional scalp from patients with FFA using single-cell RNA sequencing. Methods: Single-cell RNA sequencing was performed on lesional scalp biopsies from patients with FFA (n = 4) and matched control samples (n = 4). Cells were annotated into major lineages and refined subpopulations, and functional programmes and ligand–receptor-mediated communication were analysed. Results: Analysis of 38,984 high-quality cells revealed preservation of overall cellular composition and lineage architecture in FFA compared with controls. Despite this stability, marked disease-associated transcriptional reprogramming was observed across epithelial, immune and stromal compartments. Keratinocytes retained compartment-specific identity but exhibited prominent activation of stress-response and inflammatory pathways, particularly within upper follicle and bulge/outer root sheath populations. Dermal fibroblasts aligned along a continuous inflammation–fibrosis axis, with relative expansion of inflammatory and ACTA2⁺ states and strengthened coupling between inflammatory and fibrotic programmes. Immune profiling demonstrated skewing of CD4⁺ T cells towards Th1-, Th2- and Th22-associated signatures, together with macrophage reprogramming towards pro-fibrotic, tissue-remodelling states. Ligand–receptor analysis identified TGFB-centred signalling as the dominant inter-compartmental communication axis linking epithelial stress to immune modulation and fibroblast activation. Conclusions: FFA exhibits preserved cellular architecture but extensive, compartment-specific transcriptional reprogramming. Chronic epithelial stress and TGFB-driven inter-compartmental signalling, rather than overt cytotoxic inflammation, emerge as central disease mechanisms.

overall_design: Scalp skin biopsies were obtained from patients diagnosed with frontal fibrosing alopecia (FFA) and from healthy control subjects. Single-cell suspensions were generated from each biopsy and processed independently. Single-cell RNA sequencing libraries were prepared using the 10x Genomics Chromium Single Cell 3′ platform, enabling transcriptomic profiling at single-cell resolution. Libraries were sequenced using paired-end Illumina sequencing. Raw sequencing reads were processed using Cell Ranger with alignment to the human reference genome (GRCh38), and filtered feature-barcode matrices were generated for each sample. The study design allows comparative analysis of cell-type–specific transcriptional profiles between FFA and control scalp skin.

first: Viviana

last: Dávila-Flores

first: Jesús

last: Gay-Mimbrera

first: Carmen

last: Mochón-Jiménez

first: Irene

last: Rivera-Ruiz

last: de Luque-Fernández

first: Pedro

last: Gómez-Arias

first: Beatriz

last: Isla-Tejera

first: Benjamin

last: Ungar

last: Hu

first: Helen

last: He

first: Fernando

last: Leiva-Cepas

first: Emma

last: Guttman-Yassky


title: ER CUT&RUN profiling in MCF7 cells following abemaciclib treatment

city: Melbourne

first: Shom

last: Goel

address: 305 Grattan St

institute: Peter MacCallum Cancer Centre

summary: The retinoblastoma protein (Rb) is a tumour suppressor best known for repressing E2F transcription factors and halting cell cycle progression. In hormone receptor-positive (HR+) breast cancer, CDK4/6 inhibitors activate Rb by preventing its phosphorylation, forming a key component of current endocrine therapy regimens. How pharmacologically activated Rb remodels chromatin and influences transcription beyond cell cycle arrest remains poorly understood. Here we show that CDK4/6 inhibition induces redistribution of hypo-phosphorylated Rb to promoters and enhancers. While Rb predictably binds to cell cycle gene promoters to repress transcription, at other sites it unexpectedly promotes expression of oestrogen-responsive genes by integrating into oestrogen receptor (ER)-rich transcriptional hubs. CDK4/6 inhibition enhances ER target gene expression in breast cancer cells, patient-derived xenografts, and clinical HR+ breast cancer samples in an Rb-dependent manner. This reprogramming is mediated in part by KDM5A, whose interaction with Rb contributes to gene regulation at these loci. Critically, components of this Rb-driven ER transcriptional program are pro-proliferative. In endocrine-sensitive tumours, this can be neutralised with anti-oestrogen therapy, explaining therapeutic synergy. In endocrine-resistant settings such as ESR1-mutant breast cancer, the program persists, limiting therapeutic efficacy. These findings reframe Rb as a dual-function transcriptional regulator that, while enforcing cell cycle arrest, can also activate programs that counteract its tumour suppressor function.

overall_design: CUT&RUN for ER in estrogen receptor-positive cell line MCF7 treated with DMSO control or abemaciclib 500 nM for 2 days. One replicate per group.


title: Type I IFN Dynamics Orchestrate Protective ICAM1⁺ Neutrophil Heterogeneity as an Early Checkpoint for Survival in Lethal Viral Infection [RNA-Seq]

city: Tokyo

first: Riho

last: Saito

address: 7-3-1 Hongo Bunkyo-ku

department: Graduate School of Pharmaceutical Sciences

institute: University of Tokyo

summary: The early host response influences outcomes following viral infection. Here, we leveraged variability in survival after intranasal vesicular stomatitis virus (VSV) challenge in genetically identical mice to retrospectively profile systemic immune responses associated with survival or lethality. Survival was strongly associated with a robust systemic type I interferon (IFN) surge within 24 hours of infection, and blockade of type I IFN signaling during this narrow early window markedly reduced survival, establishing early IFN induction as a key determinant of outcome. This protective IFN surge rapidly remodeled the systemic immune landscape, inducing a specialized ICAM1⁺ neutrophil subset, primed within the bone marrow and characterized by a pro-inflammatory signature and enhanced phagocytic activity. Importantly, neutrophil-specific deletion of ICAM1 significantly reduced survival following viral infection. Together, these findings identify an early type I IFN-dependent checkpoint that shapes protective ICAM1+ neutrophil heterogeneity and disease trajectories, with implications for prognostic strategies and host-directed therapies.

overall_design: RNA-seq profiling of blood circulaing CXCR2+ neutrophils from wildtype c57b6j mice at 1 day after intranasal infection with VSV.

first: Hiroki

last: Sugishita

first: Yukiko

last: Gotoh

first: Tomohiko

last: Okazaki


title: Fibroblasts resist Staphylococcus aureus on the skin surface by responding to interleukin-1 and recruiting neutrophils

city: La Jolla

first: Jared

middle: S

last: Simmons

address: 9500 Gilman Dr

institute: University of California, San Diego

summary: Staphylococcus aureus (SA) can cause serious infections, yet it often resides on the skin without causing disease. This apparent paradox suggests a strong host defense system which prevents SA from invading the dermis. In this study, we investigate how the skin detects and responds to superficial SA exposure. Using unbiased transcriptomic, biochemical, and phosphoproteomic analyses, followed by validation of the identified activation pathways in human and mouse models, we discovered that fibroblast recognition of interleukin-1 is essential for skin inflammation in response to topical SA. Deletion or blockade of the interleukin-1 receptor in fibroblasts in vitro abolished the change in gene expression and reduced chemokine production. Furthermore, PdgfraΔIl1r1mice that lack interleukin-1 receptor 1 only on fibroblasts decreased neutrophil infiltration and allowed increased bacterial growth on the surface of mouse skin after topical SA application. These findings show fibroblasts actively participate in innate immunity and highlight an interleukin-1-dependent communication pathway from keratinocytes to fibroblasts to neutrophils. Understanding this pathway provides new insights into mechanisms that initiate neutrophil recruitment to the skin and may help develop new approaches to therapy.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Data from 2 independent experiments: 1. RNA-seq on cultured human preadipocytes after 4h in either PBS or KC-CM (dialyzed NHEK-CM) with or without anakinra (24h pretreatment). 2. scRNA-seq on Il1r1-fl/fl control or Pdgfra-cre/Il1r1-fl/fl mouse skin (C57BL/6) 14h after treatment with vehicle or S aureus (USA300).

first: Richard

last: Gallo


title: Type I IFN Dynamics Orchestrate Protective ICAM1⁺ Neutrophil Heterogeneity as an Early Checkpoint for Survival in Lethal Viral Infection [scRNA-Seq]

overall_design: Single cell RNA-seq profiling of blood circulaing immune cells from wildtype c57b6j mice before and after intranasal infection with VSV.

first: Akisawa

last: Satomi


title: Type I IFN Dynamics Orchestrate Protective ICAM1⁺ Neutrophil Heterogeneity as an Early Checkpoint for Survival in Lethal Viral Infection [BM RNAseq]

overall_design: RNA-seq profiling of peripheral bone marrow cells from wildtype c57b6j mice at 1 day after intranasal infection with VSV.


title: Type I IFN Dynamics Orchestrate Protective ICAM1⁺ Neutrophil Heterogeneity as an Early Checkpoint for Survival in Lethal Viral Infection [ICAM1 RNAseq]

overall_design: RNA-seq profiling of blood circulaing ICAM1+ and ICAM1- neutrophils from wildtype c57b6j mice at 1 day after intranasal infection with VSV.


title: Microbial volatile organic compound 2,5-D enhances Arabidopsis growth through activation of nitrate and ammonium transport

city: Saijo

first: Theint

last: Thida

state: 広島県

address: 2081-1, Joy Seven, room no. 213

department: Hiroshima University

institute: Hiroshima University

summary: Nitrogen is an essential nutrient for plant growth. We report here that 2,5-dimethylpyrazine is a key regulator of plant nitrogen and ammonium uptake through its regulation of AtNRTs and AtAMTs in the roots of Arabidopsis thaliana (Col-0) after 24 hours of exposure. In addition, 2,5-dimethylpyrazine encompassed plant hormone-related genes, such as SAURs and NRT2 family proteins, as well as defense-related transcription factors, including CNGCs, MAPK cascade, and WRKYs. Taken together, our data provide that 2,5-dimethylpyrazine is a potential candidate for promoting plant growth by regulating nitrate and ammonium transporters.

overall_design: RNA-seq profiling of wildtype Arabidopsis thaliana (Col-0) and expression of ammonium and nitrate transport genes for 24 hours exposure to 2,5-dimethylpyrazine.

first: Mami

last: Nampei

first: Matias

last: Siueia Júnior

first: Li

last: Jiacheng

first: Hisashi

last: Ômura

first: Akihiro

last: Ueda


title: Transcriptomic profiling in human breast cancer patient-derived xenograft following treatment with palbociclib and fulvestrant

overall_design: PDX KCC-P-3837 estrogen receptor-positive patient-derived xenograft tumours were transplanted into the mammary fat pads in NSG mice and treated with palbociclib, fulvestrant, and/or vehicle for 19 days. 3'RNA-seq was performed on flash frozen tumour tissues. Two tumours per treatment arm.


title: Transcriptomic profiling in human ovarian cancer cell line OVCAR-3 following CDK2 inhibitor treatment

overall_design: 3'RNA-seq profiling of OVCAR-3 cells treated with DMSO control or INX-315 300 nM for 2 days. Three replicates per group.


title: Transcriptomic profiling in MCF7 cells following KDM5A knockdown and treatment with abemaciclib

overall_design: MCF7 cells were transfected with non-targetting or KDM5A-targetting siRNA and treated with abemaciclib or DMSO control for 2 days for RNA extraction. Three replicates per arm.

first: Antonio

last: Ahn

first: Rhiannon

last: Coulson

first: April

last: Watt


title: Independent cell type-specific expression and distal regulation of the 9p21 locus cell cycle regulators: p14ARF, p16INK4A, p15INK4A, and ANRIL [scRNA-seq]

city: Cambridge

first: Neva

middle: Cherniavsky

last: Durand

address: 75 Ames St

institute: Broad Institute

summary: The 9p21 locus encodes tumor suppressors p16INK4A and p14ARF (both encoded by CDKN2A), p15INK4B (CDKN2B), and a long non-coding RNA, ANRIL (CDKN2B-AS1). The ~1 megabase locus is notable for a high density of single nucleotide polymorphisms (SNPs) associated with aging-related diseases and traits. Despite clear importance, our understanding of the the cell-type specific expression dynamics and cis-regulatory mechanisms of the 9p21 transcripts has been constrained by the complexity of the transcript structures, low expression, and severe alterations to the locus in cancer cell lines. Here, we innovated custom computational and molecular tools to overcome these technical challenges and fill critical knowledge gaps in our understanding of 9p21 locus architecture and regulation of the 9p21 transcripts. Across tissues, we found that p15INK4b is the predominant transcript. We deconvolved the expression of the CDKN2A transcripts, p16INK4A and p14ARF, revealing stark differences in the expression of these overlapping but functionally distinct transcripts across tissues and cell types. Comparative analysis in a model of fibroblast senescence revealed a striking switch from preferential p14ARF expression in cycling cells, to p16INK4A and p15INK4b as cells age and senescence. Perturbation of putative cis regulatory elements nominated a network of promoters and enhancers that regulate p16INK4A, p14ARF, and p15INK4b and ANRIL. The identified elements are accessible in vivo, and can be used to guide studies into variant interpretation. Our systematic characterization of 9p21 transcript isoforms, promoters, and distal elements across cell states, in vivo tissues and senescence offers new mechanistic insights and a framework for future studies of this vital but poorly understood locus.

overall_design: This study systematically profiles the 9p21 locus to dissect transcript expression, chromatin architecture, and regulatory element function across multiple experimental modalities. Dial-up scRNA-seq (Gray et al. and Chan et al.) captures per-cell transcript expression, including new experiments outside published datasets. Region Capture Micro-C (RCMC) maps 3D chromatin contacts at the locus. ChIP-seq for CTCF and H3K27ac identifies regulatory protein binding and active enhancer marks. A pooled CRISPRa screen with CROP-seq and single-cell RNA-seq tests candidate regulatory elements for effects on gene expression. Complementary total RNA-seq in WI-38 cells across senescence provides temporal expression dynamics. Reference and supporting files, including custom annotations and curated target region sets, facilitate reproducibility and interpretation.

first: Elena

last: Torlai Triglia

first: Tyler

middle: E

last: Miller

middle: C

first: Kyung Lock

last: Kim

first: Salva

last: Casani Galdon

first: Joshua

last: D’Antonio

first: Noam

last: Shoresh

first: Kristin

last: Ardlie

first: David

last: Hendrickson

first: J Graham

last: Ruby

first: Bradley

last: Bernstein

first: Elizabeth

last: Gaskell


title: Transcriptomic profiling in MCF7 parent and RB1 knockout cells following treatment with abemaciclib

first: Shom

last: Goel

overall_design: 3'RNA-seq profiling of estrogen receptor-positive cell line MCF7 parental cells and isogenic RB1 knockout cells, treated with DMSO control or abemaciclib 500 nM for 5 days. Three replicates per group.


title: Independent cell type-specific expression and distal regulation of the 9p21 locus cell cycle regulators: p14ARF, p16INK4A, p15INK4A, and ANRIL [Hi-C]


title: Independent cell type-specific expression and distal regulation of the 9p21 locus cell cycle regulators: p14ARF, p16INK4A, p15INK4A, and ANRIL [ChIP-seq]


title: Rb CUT&RUN profiling in ZR-75-1 cells following treatment with palbociclib

overall_design: CUT&RUN for Rb in estrogen receptor-positive cell line ZR-75-1 parental and RB1 KO cells, treated with DMSO control or palbociclib 500 nM for 4 days. Two replicates per group in parental cells, and one replicate for RB1 KO cells.


title: DNMT3B plays antagonistic roles with SMARCB1 and is a targetable vulnerability in rhabdoid tumors [Methylation array]

city: Paris

first: Franck

last: Bourdeaut

address: 26 rue d'ulm

institute: Institut Curie

type: Methylation profiling by genome tiling array

summary: Purpose: Rhabdoid tumors (RTs) are highly aggressive pediatric cancers driven by the biallelic inactivation of the SMARCB1 tumor suppressor gene, the sole recurrent genetic alteration. SMARCB1 encodes a core subunit of the SWI/SNF chromatin remodeling complex; its loss disrupts epigenetic gene regulation, supporting the classification of RTs as prototypical epigenetically driven cancers and highlighting the therapeutic potential of targeting epigenetic modifiers. Notably, previous studies have reported the overexpression of DNMT3A and DNMT3B, enzymes responsible for de novo DNA methylation, in RTs. Experimental Design: Using patient samples, cell lines, and an ex vivo brain organoid system, combined with immunohistochemistry and bioinformatics, we investigated the role of DNMT3 enzymes in RT progression. Results: In a composite tumor case, SMARCB1-deficient and -proficient regions displayed distinct methylation profiles. SMARCB1 loss correlated with increased DNA methylation and DNMT3A/B overexpression. To assess their respective roles in RTs, we used CRISPR-Cas9 to knock out DNMT3A/B in a SMARCB1-inducible RT cell line. DNMT3B loss impaired viability more strongly than DNMT3A. DNMT3B knock-out and SMARCB1 re-expression regulated overlapping gene programs related to development and cell adhesion at methylation and transcriptional levels. We next demonstrated that the cytotoxicity of the DNMT inhibitor decitabine, which impairs RT cell growth in human iPS-derived cerebral organoids, is primarily mediated by DNMT3B. Conclusions: These results show that DNMT3B plays a key role in the cascade of epigenetic effects following SMARCB1 loss and is pivotal in the RT sensitivity to decitabine; our study therefore supports the development of DNMT3B-specific inhibitors for RT.

overall_design: DNA from I2A rhabdoid tumor cells under basal condition (day 0), SMARCB1-reexpressing derivatives (days 10 and 14), DNMT3A- and DNMT3B-knock-out derivatives, and from 70 ATRT and 1 meningioangiomatosis human patient samples were analysis using Infinium MethylationEPIC arrays.

first: Céline

last: Chauvin

first: Ryszard

last: Wimmer

first: Ludovica

last: Brunetti

first: Joanna

last: Cyrta

first: Ana

last: Boranijasevic

first: Mamy

last: Andrianteranagna

first: Julien

last: Masliah-Planchon

first: Kévin

last: Beccaria

first: Sandrina

last: Turczynski

first: Grégory

last: Thomson

first: Maëva

last: Veyssière

first: Rachida

last: Bouarich-Bourimi

first: Zhi-Yan

last: Han

first: Owen

last: Hoare

first: Aurore

last: Coulomb

first: Mylène

last: Bohec

first: Sylvain

last: Baulande

first: Gaëlle

last: Pierron

first: Olivier

last: Delattre

first: Arnault

last: Tauziède-Espariat

first: Nicolas

last: Servant

first: Alexandre

last: Baffet


title: Mitochondrial profiling across macrophage states reveals inhibition of IL-4/IL-13 reprogramming by the integrated stress response

city: Epalinges

first: Alexis

last: Jourdain

address: Boveresses 155, CP51

department: Immunobiology

country: Switzerland

institute: University of Lausanne

summary: Mitochondria drive cellular reprogramming by integrating metabolism and signaling. In macrophages, mitochondria are central to immunometabolic responses to external cues, but the extent to which they are remodeled and participate in macrophage reprogramming remains unclear. Here, we integrate transcriptomics with whole-cell and purified mitochondrial proteomics to profile LPS/IFNγ- and IL-4/IL-13-stimulated macrophages. We reveal a striking disconnect between mitochondrial transcript and protein levels following either stimulus, and a STAT6-dependent increase in mitochondrial DNA (mtDNA) expression and intramitochondrial translation in IL-4/IL-13 macrophages. We demonstrate that pharmacological inhibition of mitochondrial translation or individual respiratory chain complexes variably impairs reprogramming, whereas ATP synthase inhibition uniquely triggers an HRI-dependent integrated stress response (ISR) through mitochondrial hyperpolarization, thereby preventing IL-4/IL-13 reprogramming. Mechanistically, we show that restoring mitochondrial membrane potential or inhibiting the ISR rescues IL-4/IL-13-mediated reprogramming. Together, we identify mtDNA expression, intramitochondrial translation, and mitochondrial membrane potential as critical, drug-sensitive determinants of the IL-4/IL-13 response.

overall_design: RNAseq profiling of WT C57BL/6J bone marrow derived macrophages (BMDM). BMDM were activated with either nothing (naive) or 20 ng/mL of interleukin-4 (IL-4) (Peprotech, 214-14) and 20 ng/mL of IL-13 (Peprotech, 210-13) for 24 hours. For the inhibitors, cells were pre-treated with the inhibitors for 1h and then the media was changed to media containing IL-4/IL-13 and the respective inhibitors. Oligomycin was used at 1 uM, ISRIB at 1 uM, Sal003 at 10 uM and tunicamycin at 5 ug/mL.

first: Joan

last: Blanco-Fernandez

first: Miriam

last: Lisci

first: Mads

last: Foged

first: Chloé

last: Chapuis

first: Tim

last: Pflästerer

first: Tatjana

last: Kleele


title: Rb CUT&RUN profiling of ZR-75-1 cells following treatment with abemaciclib

first: Shom

last: Goel

overall_design: CUT&RUN for Rb in estrogen receptor-positive cell line ZR-75-1 parental cells, treated with DMSO control or abemaciclib 500 nM for 4 days. Two replicates per group.


title: Transcriptomic profiling of MCF7 ESR1-PARENT and ESR1-Y537S cells cultured under estrogen-depleted or estradiol-supplemented conditions following abemaciclib treatment and CCND1 knockdown

overall_design: RNA-seq profiling was performed on MCF7 ESR1-PARENT and MCF7 ESR1-Y537S mutant cells cultured in charcoal-stripped serum (CSS) with or without estradiol (E2), transfected with either a non-targeting control siRNA (siCTRL) or siRNA targeting CCND1, and treated with either DMSO vehicle control or 500 nM abemaciclib prior to RNA extraction and sequencing. Three biological replicates were included for each experimental group.

first: Antonio

last: Ahn

first: April

last: Watt

first: Rhiannon

last: Coulson


title: Stress induces DCP2 translation via a stalling-dependent mechanism

city: Heidelberg

first: Mykola

last: Roiuk

address: Im Neuenheimer Feld 580

department: Signal Transduction in Cancer and Metabolism

institute: German Cancer Research Center (DKFZ)

summary: The integrated stress response (ISR) globally suppresses protein synthesis while selectively permitting translation of a small subset of stress-responsive mRNAs, many of which contain upstream or overlapping open reading frames (uORFs/oORFs). Although translational induction of transcripts such as ATF4 has classically been attributed to delayed re-initiation caused by reduced ternary complex availability, the mechanisms by which uORFs and oORFs allow ISR-selective translation remain incompletely understood. Here, using ribosome profiling during early ISR activation combined with reporter assays, we identify DCP2, encoding a major mRNA decapping enzyme, as a previously unrecognized ISR-induced transcript. We show that translational induction of DCP2 depends on an overlapping ORF whose conserved 3′ region, corresponding to a ribosome pausing site, acts as a potent inhibitory element. Both the DCP2 oORF and main ORF increase in translation during stress, indicating that stress relieves repression by this inhibitory element. This reveals a mode of ISR-dependent gene regulation in which inhibition by a nascent peptide or stalling element embedded either in a uORF or an oORF is relieved upon stress to induce translation.

overall_design: To determine whether the presence of uORFs affects the translation of all transcripts equally during stress, we performed ribosome profiling 2 hours after treatment with either 1 µg/mL tunicamycin or DMSO as a control. To account for stress-induced transcriptional changes, we also performed total RNA sequencing on the same samples. Because the Illumina rRNA depletion kit was discontinued during the course of the study, each ribosome profiling sample was split into two aliquots, and libraries were prepared either with or without the rRNA depletion step, to assess if ribosome profiling can be carried out without rRNA depletion.

first: Aurelio

last: Teleman


title: RNA-seq analysis of MDA-MB-231 and MDA-MB-453 cells treated with abemaciclib and/or fulvestrant

overall_design: 3'RNA-seq profiling of ER-negative breast cancer cell lines MDA-MB-231 and MDA-MB-453 cells treated with DMSO control, abemaciclib 500 nM, fulvestrant 5 nM, or a combination of abemaciclib and fulvestrant for 5 days. Three biological replicates per group.


title: Bulk RNA sequencing of mono-culture and tri-culture microglia

city: STANFORD

first: Takeshi

last: Uenaka

state: California

address: 265 Campus Drive G3145, SIM1 Bldg - 3rd floor

institute: Stanford University

summary: Microglia are the immune cells of the central nervous system and are thought to be key players in both physiological and disease conditions. Several microglial features are poorly conserved between mice and human, such as the function of the neurodegeneration-associated immune receptor Trem2. Induced pluripotent stem cell (iPSC)-derived microglia offer a powerful opportunity to generate and study human microglia. However, human iPSC-derived microglia often exhibit activated phenotypes in vitro, and assessing their impact on other brain cell types remains challenging due to limitations in current co-culture systems. Here, we developed fully defined brain microtissues, composed of human iPSC-derived neurons, astrocytes, and microglia, co-cultured in 2D or 3D formats. Our microtissues are stable and self-sufficient over time, requiring no exogenous cytokines or growth factors. In this experiment, we performed RNA-seq to compare microglia maintained in cytokine-supplemented mono-culture with microglia maintained in tri-culture without exogenous cytokines.

overall_design: RNA-seq profiling for microglia in mono-culture and the one in tri-culture, 2 weeks after they are plated

first: Wanhua

last: Li

first: Marius

last: Wernig


title: RamDA-seq analysis of CRISPRa-mediated RELA activation for proof-of-concept validation of CATaN

city: Shinjuku-ku

first: Kazuyoshi

last: Ishigaki

state: Tokyo

address: 9N3 Center for Integrated Medical Research, 35 Shinanomachi

department: Department of Microbiology and Immunology

institute: Keio University

summary: This dataset was generated to validate CATaN (Canonical correlation Analysis of Transcriptome and TF-gene regulatory Networks), an unsupervised framework that integrates transcriptomic data with transcription factor (TF) gene regulatory networks. To test whether CATaN correctly recovers the transcriptional program driven by a specific TF, we activated RELA, a subunit of NF-κB, in Jurkat cells using CRISPR activation (CRISPRa; dCas9-VP64 with a single guide RNA targeting RELA) and compared it with a control guide RNA. Transcriptomes were profiled by RamDA-seq. Transductions were performed in triplicate for each condition, and cells were harvested at five, six, and seven days post-transduction. Differential expression analysis showed concordant directional changes in RELA-regulated genes; CATaN applied to these data assigned higher CC1 loadings to RELA ChIP-seq samples than to other TFs. Together, these results provide proof-of-concept support for the method.

overall_design: Jurkat cells stably expressing dCas9-VP64 were transduced with either a single guide RNA targeting RELA (sgRELA) for CRISPR activation, or a control guide RNA (sgControl). Transductions were performed in triplicate for each condition. Cells were harvested at five, six, and seven days post-transduction and profiled by RamDA-seq. This design yielded 36 RNA-seq samples (18 sgRELA and 18 sgControl across the three time points).

first: Haruka

last: Takahashi

first: Hiroaki

last: Hatano

first: Michihiro

last: Kono

first: Koma

last: Haruta

first: Masahiro

last: Nakano

first: Reza

last: Bagherzadeh

first: Maximilian

last: Drees

first: Yuki

last: Oguma

first: Daichi

last: Harita

first: Tsugumi

last: Kawashima

first: Takahiro

last: Arakawa

first: Hajime

last: Inokuchi

last: Nishino

first: Kenichiro

last: Asahara

last: Itamiya

first: Jun

last: Inamo

first: Bunki

last: Natsumoto

first: Yumi

last: Tsuchida

first: Shuji

last: Sumitomo

first: Akari

last: Suzuki

first: Yuta

last: Kochi

first: Keishi

last: Fujio

first: Kazuhiko

last: Yamamoto

first: Tazro

last: Ohta

first: Eiryo

last: Kawakami


title: KDM5A CUT&RUN profiling of MCF7 cells treated with abemaciclib or DMSO control

first: Shom

last: Goel

state: Victoria

overall_design: CUT&RUN for KDM5A in estrogen receptor-positive cell line MCF7 treated with DMSO control or abemaciclib 500 nM for 2 days. Two replicates per group.

first: Antonio

last: Ahn

first: April

last: Watt


title: Spatial Transcriptomics Unveils Immune Microenvironment Heterogeneity and Response-related Subtypes in NSCLC Treated with Neoadjuvant Chemoimmunotherapy

city: Beijing

first: Rongzhen

last: Li

state: 北京Beijing

address: Department of Respiratory and Critical Care Medicine, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, 1 Shuaifuyuan, Dongcheng District, Beijing 100730, China

institute: Peking Union Medical College

summary: We conduct a single-arm phase 2 trial using pembrolizumab plus platinum-doublet chemotherapy to evaluate the efficacy and safety of perioperative immune-chemotherapy in patients with driver mutation-negative potentially resectable NSCLC. Further, we perform an exploratory analysis of spatial transcriptomic profiles to characterize the immune microenvironment landscape in primary tumor lesions and paired draining lymph nodes after chemoimmunotherapy, aiming to explore distinct pathological response features and potential mechanism.

overall_design: In this single-arm phase II trial (NCT05383716), patients with resectable NSCLC received neoadjuvant pembrolizumab plus platinum-based chemotherapy, followed by surgery and adjuvant immunotherapy. The primary endpoint was major pathological response (MPR). Digital spatial profiling was performed on resected tumors and paired draining lymph nodes from 22 patients to to characterize the immune landscape of response.

first: Minjiang

last: Chen

first: Zuolin

last: Cheng

first: Yuxi

last: Wei

first: Guoqiang

last: Yu

first: Ji

first: Yan

last: Xu

first: Mengzhao

last: Wang


title: Transcriptome profiling of TSPYL2 knockdown in decidualized human endometrial stromal cells

city: Nagoya

first: Jiali

last: Ruan

state: Aichi

address: 65 Tsurumai-cho

institute: Nagoya University

summary: To investigate the role of TSPYL2 during decidualization, MC1 cells, a human endometrial stromal cell line, were induced to decidualize with cAMP and MPA for 4 days and then transfected with siNC or siTSPYL2. RNA-seq was performed using three biological replicates for each group. Gene-level read counts were generated from STAR-aligned reads using featureCounts, and differential expression analysis was performed using DESeq2.

overall_design: Comparative transcriptome analysis of decidualized MC1 cells transfected with siNC or siTSPYL2. MC1 cells were induced to decidualize with cAMP and MPA for 4 days, followed by transfection with siNC or siTSPYL2. Six RNA-seq libraries were generated, including three siNC biological replicates and three siTSPYL2 biological replicates.

first: Ayako

last: Muraoka

first: Kosuke

last: Yoshida

first: Akira

last: Yokoi


title: Transcriptional profiling in esophageal adenocarcinoma

city: Ann Arbor

first: Laura

last: Kresty

email: lkresty@umich.edu, Kresty.1@osu.edu

address: 1500 E Medical Center Dr

department: Surgery

institute: University of Michigan

summary: The goal of this project is to perform transcriptomics profiling of treatment-naïve esophageal adenocarcinoma (EAC) patients to identify genes and pathways associated with esophageal adenocarcinoma. A total of 67 samples were collected from EAC patients.

overall_design: This study consists of 67 tissues from treatment or therapy-naïve esophageal adenocarcinoma cases who contributed esophageal tissue. Samples were fresh frozen, followed by RNA isolation. Transcriptomic profiling was performed to determine RNA expression levels for each patient sample.


title: Host transcriptional signatures associated with disease tolerance and environmental persistence in a mosquito-microsporidian system

city: Vancouver

first: Luis

middle: M.

last: Silva

state: British Columbia

address: 2212 Main Mall

department: Department of Zoology

institute: The University of British Columbia

summary: Experimental evolution and RNA sequencing were combined in the mosquito-microsporidian system Anopheles gambiae-Vavraia culicis to test whether parasite lineages selected for early or late transmission differ in the host transcriptional responses they elicit. Mosquitoes were infected with evolved parasite lineages and sampled at a common sporulating stage. Co-expression analyses identified host gene modules associated with infection background, virulence and environmental persistence.

overall_design: Forty-eight RNA-seq libraries were generated from pools of 10 adult female Anopheles gambiae. Samples included four uninfected control pools, four pools infected with an unselected reference Vavraia culicis lineage, twenty pools infected with five independently evolved early-selected parasite lineages, and twenty pools infected with five independently evolved late-selected parasite lineages. All infected mosquitoes were sampled on day 10 of adulthood, when infections were chronic and sporulating.

first: Luís


title: RB and KDM5A CUT&RUN profiling in MCF7 cells following KDM5A knockdown and abemaciclib treatment

first: Shom

overall_design: CUT&RUN for RB and KDM5A in MCF7 cells treated with non-targeting control siRNA or siRNA against KDM5A and DMSO control or abemaciclib. KDM5A knockdown was verified by profiling KDM5A in DMSO-treated cells. RB was profiled in samples that received control or KDM5A siRNA and treated with DMSO or abemaciclib, with two biological replicates per RB CUT&RUN condition. Mouse IgG2a and Rabbit IgG were CUT&RUN negative controls; their raw FASTQs are included.

first: Antonio


title: Twist1 Overexpression Models an Aggressive Subtype of Pancreatic Ductal Adenocarcinoma in vivo

city: Baltimore

first: yang

last: song

state: MD

address: HSF-3, 3rd Floor; 670 W Baltimore St.

department: Department: Institute for Genome Sciences

institute: University of Maryland in Baltimore

summary: Pancreatic ductal adenocarcinoma (PDAC) is a lethal cancer with very poor survival outcomes. Genomic analyses have identified distinct molecular subtypes in human PDAC with quasimesenchymal/squamous/basal-like (QSB) having the worse prognosis. However, no in vivo models exist for QSB subtypes. The epithelial-mesenchymal transition (EMT) transcription factor TWIST1 is upregulated in QSB PDAC. We hypothesized that the TWIST1-dependent plasticity program is a key regulator of QSB PDAC subtype development. Non-negative matrix factorization (NMF) of human PDAC subtypes identified 3 NMF clusters. NMF1 was dominated by QSB subtypes, had the worst overall survival, and exhibited high TWIST1 expression. We generated a novel tetracycline-inducible autochthonous pancreas specific KrasG12D-Twist1 genetically engineered tumor mouse model (Pdx1 promoter-Cre (P); LSL-KrasG12D (R); LSL-rtTA-IRES-GFP (G); Twist1-tetO-Luc (T)). PGRT mice showed accelerated pancreatic tumor development and decreased overall median survival compared to PGR (KrasG12D alone) mice (4.6 months vs 9.7 months, p< 0.0001). Histological analyses showed that PGRT pancreatic tumors spanned a spectrum of PDAC to QSB PDAC subtype, exhibited an EMT profile and squamous histology. Twist1 induction resulted in increased metastases (9.52 % PGR at 48 weeks vs 73.91% PGRT at 24 weeks, p<0.0001). PGRT tumors demonstrated gene expression programs related to collagen content, EMT and cell movement and was most concordant with human QSB PDAC subtypes and squamoid single cell subtype. Twist1 overexpression cooperated with KrasG12D for PDAC development in vivo and progression towards metastasis and mimicked the human QSB PDAC subtypes. This novel model may offer new perspectives for PDAC research and for the testing of novel therapeutic strategies in an aggressive PDAC subtype.

overall_design: The RNA sequencing profiling of different mouse alleles PGT (Control), KrasG12D (PGR), KrasG12D/+-p53R172H/+ (KPC), and KrasG12D-Twist1-overexpressing (PGRT) was compared to showed that Twist1 overexpression model the QSB variant of PDAC.

first: Muhammad Ajmal

last: Khan

first: Jinhee

last: Chang

first: Yang

last: Song

first: Amol

last: C. Shetty

first: Triet

last: Nguyen

first: Danielle

last: N. Waters

first: Dipanwita

last: Dutta Chowdhury

first: Elan

last: Simms

first: Aaron

last: Chan

first: Christine

last: Lam

first: Priyanshi

last: Patel

first: Lucas

last: T. Tran

first: Jiayu

first: Jarey

last: H. Wang

first: Katriana

last: Nugent

first: Matthew

last: Ballew

first: Ghali

last: Lemtiri-Chlieh

first: Francesca

last: Anna Carrieri

first: Reem

last: Malek

first: Hailun

first: Noelle

last: R. (Jurcak) Thielman

first: Keyu

first: William

last: L. Hwang

first: Lei

last: Zheng

first: Kathleen

last: Gabrielson

first: Phuoc

first: Audrey

last: Lafargue


title: scATACseq of sorted stromal cell compartment from patients with Crohn's disease

first: Simon

last: Koplev

address: Jeffrey Cheah Biomedical Centre Cambridge Biomedical Campus, Puddicombe Way

department: Cambridge Stem Cell Institute

country: United Kingdom

institute: University of Cambridge

overall_design: n=13 tissues (samples) pre-enriched by FACS sort of CD45-EPCAM- stromal cells

first: Matthias

last: Friedrich


title: scRNAseq of sorted stromal cell compartment from patients with Crohn's disease


title: bulk RNAseq of Ccd18-Co cell line treated with cytokine mix and/or epigenetic modulator

overall_design: Ccd18-Co cell line unstimulated or stimulated with cytokines, followed by vehicle or epigenetic modulators. N=4 replicates per treatment group.