title: Sirtuin 7 regulates dosage compensation and safeguards the female X-chromosome [ChIP_SIRT7_TTF]

first: Joshua

last: Thackray

overall_design: ChIP-seq profiling of SIRT7 enrichment in WT and SIRT7-KO hybrid (mus/cas) mouse TTF cells.

first: Nicolas

last: Simonet

first: Barry

last: Kesner

first: François

last: Serra

first: Maria

last: Espinosa-Alcantud

first: Anna

last: Lappala

last: Guitart

first: Laia

last: Wang


title: Pulmonary fibroblast activation during Aspergillus fumigatus infection enhances lung defense via immunomodulation and tissue remodeling [RNA-Seq]

status: Public on Apr 07 2026

city: Columbus

first: Jose

middle: P

last: Guirao-Abad

state: OH

address: 2255 Kenny Rd

department: Emergency Medicine

country: USA

institute: Ohio State University

type: Expression profiling by high throughput sequencing

summary: Aspergillus fumigatus is the etiologic agent of invasive aspergillosis, a life-threatening fungal pneumonia that is initiated by the inhalation of conidia (spores) into the lung. If the conidia are not cleared, they secrete large quantities of hydrolytic enzymes and toxins as they grow, resulting in extensive damage to pulmonary tissue. Stromal fibroblasts are central responders to tissue damage in many organs, but their functional response to pulmonary injury caused by A. fumigatus has not been explored. In this study, we employed cell lineage tracing, targeted cell ablation, and single-cell RNA sequencing to monitor the dynamics of fibroblast behavior upon exposure to A. fumigatus in both immunocompetent and immunosuppressed hosts. The results demonstrate that a subset of pulmonary fibroblasts becomes activated in an immunocompetent host in response to a challenge with A. fumigatus conidia, acquiring a gene expression program reflecting the acquisition of new immunomodulatory properties as well as enhanced extracellular matrix (ECM)-secreting ability. Remarkably, through targeted ablation of fibroblasts that express the profibrotic activation marker periostin, we demonstrate that the progression of an invasive A. fumigatus infection in an immunosuppressed host is accelerated by the absence of periostin lineage cells and is accompanied by severe alveolar hemorrhage and angioinvasion. These findings uncover a novel protective role for fibroblasts in limiting the severity of A. fumigatus-induced pulmonary injury and emphasize the importance of the pulmonary stroma in host defense against this invasive fungal infection.

overall_design: Immunocompetent PostnLin mice were administered a single inoculum of A. fumigatus conidia to induce fibroblast activation and Pdgfrα-GFP mice were given a mock inoculation with saline. The induced PostnLin cells and the resident Pdgfrα-GFP fibroblasts were then isolated on day 7 post-inoculation by sorting tdTomato+ or GFP+ cells. Total RNA was extracted from the two populations and bulk RNA-Seq was performed to compare the transcriptional profile of A. fumigatus-induced PostnLin cells to that of Pdgfrα+ resident fibroblasts.

first: Shannon

last: Shearer

first: Jon

last: Bowden

first: Daniel

middle: A

last: Kasprovic

first: Christina

last: Grisham

first: Mustafa

last: Ozdemir

first: Elliot

last: Lowe

first: Michael

last: Tranter

first: Yunguan

first: David

middle: S

last: Askew

first: Onur

last: Kanisicak


title: Sirtuin 7 regulates dosage compensation and safeguards the female X-chromosome [RNA-seq ES]

city: Piscataway

first: Joshua

middle: K

last: Thackray

state: NEW JERSEY

address: 145 Bevier Road

department: Human Genetics Institute of New Jersey

institute: Rutgers University

summary: Sirtuins are deacetylases implicated in stress responses and longevity in mammals. Although their differential impact on the two sexes has been noted, underlying causes for sex biases in aging and cancer are not known. Here, using Sirtuin 7 (SIRT7) as a model in mice, we probe mechanisms leading to sex differences. We find that Sirt7-/- females have decreased fitness during embryogenesis and throughout life. Intriguingly, SIRT7 preferentially localizes to the X-chromosome, suggesting a potential defect in X-inactivation — the arm of dosage compensation that equalizes X-chromosome expression between XX females and XY males. Indeed, SIRT7 ablation causes increased Xist RNA levels, H3K27me3 enrichment, and gene repression on the inactive X (Xi) chromosome. Surprisingly, however, loss of SIRT7 exerts greatest effects on the active X (Xa). In Sirt7-/- females, the Xa demonstrates 3D structural disorganization, increased chromatin mixing, and acute propensity to break and form chromosome fusions in an Xa-specific manner. Disproportional to autosomes and Xi, the Xa becomes super-acetylated on H3K36 and super-activated in gene expression, resulting in an imbalance in the 2nd arm of dosage compensation — known as “Xa-hyperactivation” — which evolved to balance X-linked gene expression against the genome. These data reveal a crosstalk between sirtuins and dosage compensation and identify a novel regulator of Xa-hyperactivation.

overall_design: RNA-seq profiling of WT and SIRT7-KO hybrid (mus/cas) mouse ES cells at day 0 (LIF treated) and 7 days after LIF withdraw.

first: Nicolas

middle: G

last: Simonet

last: Marin

first: Uri

last: Weissbein

first: Danni

first: Josema

last: Castelló

first: Llorenç

last: Rovirosa

first: Jay

last: Tischfield

first: Lourdes

last: Serrano

first: Berta

middle: N

last: Vazquez

first: Aurora

last: Ruiz-Herrera

first: Bernhard

last: Payer

first: Biola

middle: M

last: Javierre

first: Alejandro

last: Vaquero

first: Jeannie

middle: T

last: Lee


title: Sirtuin 7 regulates dosage compensation and safeguards the female X-chromosome [H3K27me3_ES]

type: Genome binding/occupancy profiling by high throughput sequencing

overall_design: ChIP-seq profiling of H3K27me3 enrichment in WT and SIRT7-KO hybrid (mus/cas) mouse ES cells at day 0 (LIF treated) and 7 days after LIF withdraw.

first: Barry

last: Kesner

first: François

last: Serra

first: Maria

last: Espinosa-Alcantud

first: Anna

last: Lappala

last: Guitart

first: Laia

last: Wang


title: Subcellular mRNA localization patterns across tissues resolved with spatial transcriptomics

status: Public on Apr 06 2026

city: Rehovot

first: Roy

last: Novoselsky

address: Herzl 234

department: Molecular cell biology

country: Israel

institute: Weizmann Institute of Science

summary: Subcellular RNA localization, including nuclear retention and apical-basal compartmentalization in polarized epithelia plays a central role in post-transcriptional regulation. However, methods for high-throughput mapping of mRNA localization within intact tissue sections remain limited. Here, we apply high-resolution spatial transcriptomics to systematically resolve intracellular mRNA localization across diverse mammalian tissues. We introduce a computational approach that leverages image-derived features to extract subcellular information from spatial data and quantifies transcript localization patterns. Using this framework, we map apical-basal mRNA localization and nuclear retention in gastrointestinal epithelia and in liver hepatocytes. Our analyses reveal conserved and tissue-specific localization signatures that can be readily obtained from standard high-definition spatial transcriptomics experiments. This approach broadens the scope of spatial transcriptomics by enabling routine investigation of intracellular RNA distributions in both healthy and diseased tissues.

overall_design: 10x VisiumHD of FFPE liver samples  from two healthy WT C57BL/6 mice housed under normal conditions. The experiment included fluorescence staining for Atp1a1, Cd31, autofluorescence and DAPI.

first: Ofra

last: Golani

first: Tal

last: Barkai

first: Merav

last: Kedmi

first: Inna

last: Goliand

first: Shalev

last: Itzkovitz


title: Sirtuin 7 regulates dosage compensation and safeguards the female X-chromosome [H3K27me3_TTF]

first: Joshua

last: Thackray

overall_design: ChIP-seq profiling of H3K27me3 enrichment in WT and SIRT7-KO hybrid (mus/cas) mouse TTF cells.

first: Nicolas

last: Simonet

last: Marin

first: Uri

last: Weissbein

first: Danni

first: Josema

last: Castelló

first: Llorenç

last: Rovirosa

first: Jay

last: Tischfield

first: Lourdes

last: Serrano

first: Berta

last: Vazquez

last: Ruiz-Herrera


title: A Tunable Cas12a Platform for Single-Cell Perturbation Screening and CRISPRi

city: South San Francisco,

first: Carolina

last: Galan

state: CA

address: 1 DNA Way

institute: Genentech

summary: Single-cell perturbation (Perturb-seq) screens have primarily relied on Cas9 for inducing loss-of-function phenotypes, whereas Cas12a, despite its unique effectiveness for multiplex guide expression, remains underexplored. This may be due to Cas12a’s guide RNA array (pre-crRNA) self-processing activity and the subsequent challenges associated with pre-crRNA sequence recovery during single-cell RNA sequencing library preparation. To overcome the self-processing constraint, we optimized pre-crRNA expression vectors and established a degron-based, enhanced Cas12a system for gene knock-out. As demonstrated across cell types, target genes, and with a minimized guide RNA library, this new platform allows for accurate detection of pre-crRNAs and gene editing-induced effects on the transcriptome in single cells. Additionally, we show that HyperLbCas12a outperforms other existing variants for multiplexed gene suppression. While the rapid reversibility of this repressor highlights specific kinetic constraints for degron-based single-cell recording, the system provides a potent, modular tool for contexts requiring tunable, transient silencing. Together, this suite of technologies greatly expands the possibilities for future Perturb-seq efforts and broader application of Cas12a for genetic disruption at-scale.

overall_design: Plasmid libraries consisting of dual pre-crRNAs were transduced into H358-enAsCas12a or H358-enAsCas12a-FKP12F36V cells. In the case of H358-enAsCas12a-FKBP12F36V, prior to 10x processing, cells were cultured in 1µM dTAGV-1 (Tocris Bioscience, 6914) for 24h or 48h. dTagV1 was continuously supplied until the step of reverse transcription. Gene expression libraries were prepared according to the 10X Genomics according to the manufacturer’s protocol  (CG000316 or CG000418); CRISPR libraries for direct-capture Perturb-seq and direct-capture CROP-seq constructs were created according to 10x genomics user manuals (CG000316 or CG000418).

first: Valentina

last: Snetkova

first: Romain

last: Lopez

first: Antonio

middle: R

last: Rios

first: Takamasa

last: Kudo

first: Kristel

last: Dorighi

first: Søren

last: Warming

first: Benjamin

middle: J

last: Haley


title: Cardiac tertiary immune niches drive immune activation in immune checkpoint inhibitor myocarditis

city: New York

first: Carly

last: Tymm

state: New York

institute: Columbia University Medical Center

summary: Using integrated spatial and single-cell analyses in a pharmacological murine model, we identified regional infiltration of inflammatory myeloid cells and PD-1⁺CD8⁺ T cells in the heart that organize into fibroblast-rich immune structures, which we term Tertiary T Cell Niches (TTCNs). TTCNs serve as hubs for T cell activation, sharing features of tertiary lymphoid structures. Complementary TCR analyses revealed clonal expansion of cardiac T cells following ICI treatment. Together, these findings suggest that cardiac tertiary immune structures play a central role in activating and recruiting immune cells in ICI myocarditis and highlight pathways that could mitigate ICI cardiotoxicity.

overall_design: For the single-cell RNA-seq and TCR sequencing, all data were generated using the 10X Genomics Chromium OCM platform.  Extravascular cardiac leukocytes isolated from ICI-treated mice included three independent samples, each consisting of cells pooled from two mice (AC001, AC002, AC004). Intravascular cardiac leukocytes from 8 mice were pooled into 1 sample: AC0014IV. OCM samples were multiplexed and sequenced across two lanes for both gene expression (GEX) and TCR libraries.                                                                                Sample ID with OCM Barcode: AC001 OB1 AC002 OB2 AC0014IV OB3 AC004 OB4

first: Tymm

last: Carly

first: Bukhari

last: Syed

first: Mor

last: Adam


title: Single-Cell Transcriptomic and Chromatin Accessibility Atlas of Peripheral Blood Mononuclear Cells Reveals Immune Cell Heterogeneity and Breed- Specific Characteristics in Duroc and Meishan Pigs

city: Yangzhou

first: Cui

last: Du

state: Jiangsu

address: NO.88 University South Road

country: China

institute: Yangzhou University

summary: In this study, we applied scRNA-Seq to profile the PBMCs from Meishan and Duroc pigs which are two strains with distinctive immune traits, and applied scATAC-Seq to Duroc PBMCs. We expect this study will improve the understanding about the immune heterogeneity and inter-breed immune variations in pigs, and reveal novel immune cell populations, immune genes and regulators that would facilitate future biomedical studies. MS(Meishan pig); DR(Duroc pig).

first: Wenbin

last: Bao

first: Hairui

last: Fan


title: Sirtuin 7 regulates dosage compensation and safeguards the female X-chromosome [H3K36Ac_TTF]

first: Joshua

last: Thackray

overall_design: ChIP-seq profiling of H2AX and gammaH2AX enrichment in WT and SIRT7-KO mus/cas hybrid TTF cells.

first: Nicolas

last: Simonet

first: Barry

last: Kesner

first: François

last: Serra

first: Maria

last: Espinosa-Alcantud

first: Anna

last: Lappala

last: Guitart

first: Laia

last: Marin

first: Uri

last: Weissbein

first: Danni

last: Wang

first: Josema

last: Castelló

first: Llorenç

last: Rovirosa

first: Jay

last: Tischfield

first: Lourdes

last: Serrano

last: Vazquez

first: Aurora

first: Bernhard

last: Payer

first: Biola

last: Javierre

first: Alejandro

last: Vaquero

first: Jeannie

last: Lee


title: Sirtuin 7 regulates dosage compensation and safeguards the female X-chromosome [H3K36Ac_ES]

first: Joshua

last: Thackray

overall_design: ChIP-seq profiling of H3K36Ac enrichment in WT and SIRT7-KO hybrid (mus/cas) mouse ES cells at day 0 (LIF treated) and 7 days after LIF withdraw.

first: Nicolas

last: Simonet

last: Kesner

first: Berta

last: Ruiz-Herrera


title: Adjuvant personalized multivalent neoantigen DNA vaccination for MGMT unmethylated glioblastoma: a phase I trial (bulk TCR-seq)

first: Katherine

middle: E

last: Miller

address: 700 Children's Dr.

department: Institute for Genomic Medicine

institute: Nationwide Children's Hospital

summary: Glioblastoma is a fatal disease with a median prognosis of 12-18 months. Recent studies have shown encouraging results using neoantigen-based vaccines to stimulate glioblastoma-directed immune responses, but overall immunogenicity has been low. Here, we report the results of an open-label, single-arm, phase I clinical trial (GT-20; NCT04015700) to evaluate safety and feasibility (primary endpoints), as well as immunogenicity and preliminary clinical activity (secondary endpoints) of GNOS-PV01 monotherapy, a DNA-based personalized therapeutic cancer vaccine administered following surgical resection and radiation for patients with MGMT unmethylated glioblastoma. The GT-20 study vaccinated 9 patients utilizing up to 40 neoantigens per patient (range 17- 40) without causing any serious adverse events, unexpected toxicities, or dose limiting toxicities. The vaccine induced activation and expansion of circulating peripheral T cells in all evaluated patients, except one being treated with dexamethasone. The secondary endpoint was to evaluate six-month progression-free survival and twelve-month overall survival; each observed in 66.7% of patients. Median progression-free survival was 8.5 months, median overall survival was 16.3 months, and survival at 24 months was 33%, including one long-term survivor still alive four years from the time of initial surgery. This study met the pre-specified endpoints and supports the use of GNOS-PV01 as a potentially impactful component of glioblastoma immunotherapy.

overall_design:
We performed bulk TCR-sequencing (Adpative Biotechnologies immunoSEQ) on multiple tumor regions and PBMCs from different timepoints collected from three patients on the GT-20 protocol.
*** Adaptive Biotechnologies does not release raw data ***

first: Elizabeth

last: Garfinkle

first: Gavin

last: Dunn

first: Tanner

last: Johanns


title: HyDRA: a pipeline for integrating long- and short-read RNAseq data for custom transcriptome assembly

city: Brisbane

first: Juliet

last: French

address: 300 Herston Rd

country: Australia

institute: QIMRB

summary: Short-read RNA sequencing (RNAseq) remains a cornerstone for transcriptome profiling, but is limited in reconstructing full-length transcripts and capturing transcript diversity. While long-read RNAseq spans entire transcripts and resolves complex structures, this technology is hindered by its high error rates. In parallel, noncoding RNA transcripts remain underrepresented in current references. Here, we present HyDRA (Hybrid de novo RNA Assembly), a pipeline that integrates the accuracy of short reads with the structural resolution of long reads to produce more complete de novo transcriptome assemblies. Benchmarking showed HyDRA to outperform existing methods by up to 40%. Using the HyDRA human ovarian metatranscriptome, we identified >50,000 high-confidence long noncoding RNAs, most of which have not been previously detected using traditional methods. Although long-read RNAseq is advancing, the vast availability of short reads ensures HyDRA’s ongoing role in capturing high-confidence, cell-type specific transcripts and advancing our understanding of transcriptomic complexity and the noncoding genome.

overall_design: Short-read RNAseq data from primary and immortalized fallopian tube secretory epithelial cells (FT194, FT282, FC-0081-2839, FC-0081-4853, and ABC-TC5521) and ovarian surface epithelial cells (IOSE397, IOSE7576, ABC-TC3743, 7310-donor1-24593, and 7310-donor2-7859) and long-read RNAseq data from immortalized fallopian tube secretory epithelial cells (FT194) and ovarian surface epithelial cells (IOSE397)

first: Isabela

last: Almeida

first: Xue

last: Lu

first: Stacey

last: Edwards

first: Mainá

last: Bitar


title: Systematically characterizing the roles of E3-ligase family members in inflammatory responses with massively parallel Perturb-seq

city: Palo Alto

first: Basak

last: Eraslan

address: 2360 Saint Francis Drive

summary: E3 ligases regulate key processes, but many of their roles remain unknown. Using Perturb-seq, we interrogated the function of 1,130 E3 ligases, partners and substrates in the inflammatory response in primary dendritic cells (DCs). Dozens impacted the balance of DC1, DC2, migratory DC and macrophage states and a gradient of DC maturation. Family members grouped into co-functional modules that were enriched for physical interactions and impacted specific programs through substrate transcription factors. E3s and their adaptors co-regulated the same processes, but partnered with different substrate recognition adaptors to impact distinct aspects of the DC life cycle, several of which we validated experimentally. Genetic interactions were more prevalent within than between modules, and a deep learning model, comBVAE, leveraged modularity to predict the outcome of combinations. We experimentally validated 91 combinations inferring molecular mechanisms producing non-aditivity. The E3 regulatory network was associated with heritable variation and aberrant gene expression in immune cells in human inflammatory diseases. Our study provides a general approach to dissect gene function.

overall_design: Mouse bone marrow-derived macrophages (BMDCs) were transduced on day 2 with CRISPR-delivering lentivirus. Cells were activated with LPS for 3 hours and harvested on day 9. Cells then underwent antibody staining for cell hashing (HTOs) and transduced cells were enriched by Fluorescence Activated Cell Sorting (FACS). There are three experiments included in this study: two unique perturbation experiments and an unperturbed baseline study looking at gene expression changes upon LPS activation.

first: Kathryn

last: Geiger-Schuller

first: Olena

last: Kuksenko

first: Anugraha

last: Rajagopalan

first: Ana

last: Meireles

first: Markus

last: Brown

first: Kushal

last: Dey

first: Karthik

last: Jagadeesh

first: Pratiksha

middle: I

last: Thakore

first: Ozge

last: Karayel

first: Andrea

last: Yung

first: Simone

last: Haag

first: Karren

last: Dai Yang

first: Liat

last: Amir-Zilberstein

first: Toni

last: Delorey

first: Devan

last: Phillips

first: Raktima

last: Raychowdhury

first: Christine

last: Moussion

first: Alkes

middle: L

last: Price

first: Nir

last: Hacohen

first: John

last: Doench

first: Caroline

last: Uhler

first: Orit

last: Rozenblatt-Rosen

first: Aviv

last: Regev


title: Essential lncRNAs in the human transcriptome [CRISPR]

first: Wen-Wei

last: Liang

state: NY

address: 101 Avenue of the Americas

institute: New York Genome Center

summary: Mammalian genomes host a diverse array of RNA that includes protein-coding and noncoding transcripts. However, the functional roles of most long noncoding RNAs (lncRNAs) remain elusive. Using RNA-targeting CRISPR-Cas13 screens, we probed how loss of ~5,500 lncRNAs impacts cell fitness across five human cell lines and identified 788 lncRNAs with context-specific or broad essentiality. We confirm their essentiality with individual perturbations and find that the majority of essential lncRNAs operate independently of their nearest protein-coding genes. Using transcriptome profiling in single cells, we discover that loss of essential lncRNAs impair cell cycle progression and drive apoptosis. Many essential lncRNAs demonstrate dynamic expression across tissues during development. Using ~9,000 primary tumors, we pinpoint those lncRNAs whose expression in tumors correlates with survival, yielding new biomarkers and potential therapeutic targets. This transcriptome-wide survey of functional lncRNAs advances our understanding of noncoding transcripts and demonstrates the potential of transcriptome-scale noncoding screens with Cas13.

overall_design: To profile transcriptomic change after essential lncRNA perturbation, we engineered two human cell lines, HAP1 and MDA-MB-231, to express the nuclear-localized RfxCas13d effector under doxycycline-inducible control. Next, we transduced each cell line with the lentiviral gRNA library at a low multiplicity-of-infection (<0.2) to ensure each cell received only a single perturbation and induced Cas13 expression via doxycycline addition. Cells were harvest after 3 days and loaded into a Chromium Controller (10X Genomics). Gene expression (GEX) libraries were prepared using Chromium Single Cell 3ʹ v3 protocol (10x Genomics). bcgRNA (BCG) libraries from CaRPool-seq experiments were prepared using the standard feature barcoding Chromium Single Cell 3ʹ v3 (10x Genomics) workflow and additional steps noted in carpool-seq.com.


title: Fate mapping reveals a prenatal-to-neonatal wave of ILC2s with a history of Cd3g expression - RNA-Seq

city: Oklahoma City

first: Aneta

last: Pankow

state: Oklahoma

address: 825 NE 13th St

department: Arthritis and Clinical Immunology

institute: Oklahoma Medical Research Foundation

summary: Group 2 innate lymphoid cells (ILC2s) are thought to develop in the bone marrow and fetal liver. However, increasing evidence supports the presence of thymic ILC2s in mice and humans. In this study, we introduce a novel fate-mapping mouse model designed to track thymic ILC2s in peripheral tissues. Thymic ILC2s are labeled by ZsGreen whose expression is induced by Cd3giCre. While they are present in the thymus and peripheral tissues, they are not found in the bone marrow. These cells are predominantly abundant during the neonatal stage, when the intense alveolarization and tissue remodeling occur. RNA sequencing revealed elevated expression of cytokine genes, Il13 and Il4, as well as Klrg1 in these cells. They also harbor TCR gene rearrangements suggesting common developmental pathways with T cells. Furthermore, we demonstrate that neonatal ILC2s play a critical role in macrophage polarization in a naphthalene-induced injury lung model. These findings help explore the differences between pediatric and adult immunity and future therapeutic strategies.

overall_design: Gene expression profile was determined by bulk RNA sequencing of 14 days old RORaf/f (C) or RORaf/fIl7riCre (KO) mice. RNA isolated from whole lung tissue from untreated (Untr-2) mice and mice after 2 days (Tr-2), 5 days (Tr-5) and 7 days (Tr-7) days after treatment with naphthalene.

first: Xiao-Hong

last: Sun


title: Infiltration of monocyte-derived cells perturbs CNS metabolism, licensing arginine catabolism augmenting neuroinflammation

city: Vienna

first: Omar

last: Sharif

address: Schwarzspanier Strasse

department: Centre for Physiology and Pharmacology, Vascular Biology and Thrombosis research

country: Austria

institute: Medical University of Vienna

summary: Inflammatory responses are associated with recruitment of monocyte-derived cells (Mdcs) into tissues. While tissue-specific Mdc reprogramming is well-established, how Mdc inflitration alters tissue metabolism remains unclear. Here, using a murine neuroinflammation model coupled with genetic fate-mapping, metabolomics, and metabolite imaging, we identify that central nervous system (CNS) Mdc infiltration is associated with substantial metabolic changes, assigning disease-linked metabolites (DLMs) therein. Particularly, we found increased arginine catabolism driven by lesion-associated Arginase-1 (Arg1)-expressing Mdcs promoted oxidative damage, lipid accumulation and Mdc dysfunction. Genetic Arg1-deficiency within Mdcs during neuroinflammation increased extracellular arginine and was associated with rewiring of the CNS metabolic landscape, including attenuated DLMs. This was accompanied by enhanced Mdc-driven anti-inflammation, regulatory T cell expansion, and improved disease outcome. Opposing effects were observed upon dietary arginine deficiency.  Together, our work highlights key roles for Mdcs in CNS metabolism and reveals the pleiotropic beneficial effects of arginine in neuroinflammation.

overall_design: viable CD3-Ly6G-CD45+CX3CR1+Arg1+ cells were isolated and FACS sorted from spinal cords of EAE-induced Arg1tm1Lky animals and compared to viable CD3-Ly6G-CD45+CX3CR1+Arg1-

first: Martina

last: Kerndl

first: Andrea

last: Vogel

last: Komljenovic

first: Gernot

last: Schabbauer


title: A novel triple-action Pt(IV) prodrug presents multiple anti-cancer effects and overcomes cisplatin resistance

city: Jerusalem

first: Sheera

last: Adar

state: Israel

address: Botnar Building Room 001, Faculty of Medicine, Hebrew University, Ein Kerem

department: Department of Microbiology and Molecular Genetics

institute: Hebrew University of Jerusalem

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

overall_design: Refer to individual Series


title: Discovery of a primed endothelial progenitor that requires VEGF/ERK inhibition to complete vein differentiation [kairos_knockdown_scrnaseq]

city: Stanford

first: Lay Teng

last: Ang

address: Institute for Stem Cell Biology & Regenerative Medicine, Lokey Stem Cell Bldg, 265 Campus Drive

department: Urology

institute: Stanford University

summary: Extracellular signals and cell-fate trajectories during vein development remain elusive, despite trailblazing insights into artery development. Here we exploit human pluripotent stem cell differentiation and mouse embryology to present a model that answers longstanding questions: vein endothelial cell (EC) differentiation unfolds in two steps driven by opposing extracellular signals. First, VEGF differentiates mesoderm into “primed” ECs, newly-defined progenitors that co-express certain arterial (SOX17) and venous (APLNR) markers. Second, primed ECs execute vein differentiation upon VEGF/ERK inhibition; however, upon VEGF activation they can instead form artery ECs. The arteriovenous plasticity of primed ECs was supported by intersectional lineage tracing. Future venous genes including NR2F2 harbor poised chromatin in primed ECs, but are only transcribed upon VEGF/ERK inhibition. SOXF transcription factors, including SOX17, confer primed ECs with vein differentiation competence. Collectively, this two-step vein differentiation model—entailing primed EC intermediates and VEGF/ERK inhibition to trigger vein differentiation—has implications for VEGF-modulating therapies.

overall_design: HPSCs were differentiated into primitive streak, lateral mesoderm, artery ECs, primed ECs, vein ECs.

first: Sherry Li

last: Zheng

first: Kevin J

last: Liu

first: Anastasiia

last: Masaltseva

first: Kyle M

last: Loh


title: Lymphomatoid papulosis vs. advanced-stage cutaneous T-cell lymphomas: Single-cell profiling reveals markers of self-limited vs. aggressive disease behavior

first: Katharina

last: Rindler

address: Waehringer Guertel 18-20

department: Dermatology

summary: Cutaneous T-cell lymphomas (CTCL) encompass a broad spectrum from highly indolent to aggressive systemic disease, but underlying mechanisms remain only poorly understood. Using single-cell RNA sequencing (scRNA-seq), we profiled samples from patients with self-limited, spontaneously regressing lymphomatoid papulosis (LyP) lesions of CD4+, CD8+ or TCR-γδ+ clonal phenotypes, and compared results with cases of advanced-stage CTCL (aCTCL) variants, namely CD4+ MF, CD8+ AECTCL, and TCR-γδ+ MF, that all led to a documented lethal disease outcome. Within T cells, transcriptomic differences between LyP and the aCTCL group were primarily found among malignant clones, with only relatively minor differences within the polyclonal T-cell infiltrates. When compared to all aCTCL diagnoses, LyP top clones consistently exhibited a hyperactivated cytotoxic phenotype, characterized by enrichment of TNF-alpha, IL2/STAT5, interferon gamma, hypoxia, and complement activation pathways, irrespective of their CD4, CD8 or TCR-γδ lineage. In line, LyP lesions contained SPP1+ macrophages expressing M1-associated markers (INHBA, TNF and FBP1), consistent with a cytotoxic, type 1-skewed immune microenvironment, that were largely absent across aCTCL diagnoses. LyP samples further expressed markers of vascular dysfunction and stress (C2CD4B, IL6, CEBPD), corroborated by morphological signs of lymphocytic vasculitis and necrosis in a subset of patients. Along with signs of an insufficient coping response to hypoxia, these stromal reactions might be critical contributors to spontaneous lesion regression in LyP. Taken together, this study reveals mediators associated with the self-limited behavior of LyP lesions, which might be relevant for future immunomodulatory treatment strategies in CTCL.

overall_design:
single-cell RNA-seq investigation of skin lesions in CD30+LPD and advanced-stage CTCL
>>>Raw data are unvailable due to patient privacy concerns<<<

first: Shannon

last: Meledathu

first: Sumanth

last: Chennareddy

first: Malini

last: Naidu

first: Shane

last: Meehan

first: Jonas

last: Adalsteinsson

first: Natalia

last: Alkon

first: Emry

last: Cohenour

first: Grace

last: Christensen

first: Lauren

last: Port

first: Abigail

last: Fleischli

first: Amelia

last: Burnett

first: Constanze

last: Jonak

first: Patrick

last: Brunner


title: Single-cell RNA sequencing of iALI epithelial and mesenchymal cell derived from UHOMi003-A cell line

city: Montpellier

first: Said

last: Assou

state: Languedoc-Roussillon

address: 80 av Augustin fliche, IRMB

country: France

institute: IRMB, Hôpital Saint Eloi

summary: This study aims to specify the effect of air pollution (PM2.5 developed by NIST), on bronchial epithelium generated from induced pluripotent stem cells, called iALI. The exposition was performed during 15 days on basal side and media was changed everyday. We performed single-cell RNA-sequencing (10x Genomics Chromium) to capture 10 000 cells on EpCAM+ cells (epithelial cells) and on EpCAM- cells (mesenchymal cells), in both control and exposed condition. Then, analyse was perfomed in R by using Seurat pipeline, to identified each cluster cells and induced and repressed genes.

overall_design: Transcriptomic analysis of iALI at single cell resolution leads to the identification of cluster cells based on literature known markers, and then visualized by Uniform Manifold Approximation and Projection (UMAP) using Seurat.

first: John

last: De Vos

first: Arnaud

last: Bourdin

first: Amel

last: Nasri


title: Sirtuin 7 regulates dosage compensation and safeguards the female X-chromosome [CHART-seq]

first: Joshua

last: Thackray

overall_design: CHART-seq profiling of Xist transcript binding in WT and SIRT7-KO hybrid (mus/cas) mouse Tail Tip Fibroblasts (TTF).

first: Nicolas

last: Simonet

first: Barry

last: Kesner

first: François

last: Serra

first: Maria

last: Espinosa-Alcantud

first: Anna

last: Lappala

last: Guitart

first: Laia

last: Marin

first: Uri

last: Weissbein

first: Danni

last: Wang

first: Josema

last: Castelló

first: Llorenç

last: Rovirosa

first: Jay

last: Tischfield

first: Lourdes

last: Serrano

first: Berta

last: Vazquez

first: Aurora

last: Ruiz-Herrera

first: Bernhard

last: Payer

first: Biola

last: Javierre

first: Alejandro

last: Vaquero

first: Jeannie

last: Lee


title: Sirtuin 7 regulates dosage compensation and safeguards the female X-chromosome [RNA-seq TTF]

first: Joshua

last: Thackray

overall_design:
RNA-seq profiling of WT and SIRT7-KO hybrid (mus/cas) mouse ES cells at day 0 (LIF treated) and 7 days after LIF withdraw.
RNA-seq profiling of  WT and SIRT7-KO, male or female, primary mouse MEF cells.

first: Nicolas

last: Simonet

first: Barry

last: Kesner

last: Serra


title: Cardiac tertiary immune niches drive immune activation in immune checkpoint inhibitor myocarditis [CosMx]

first: Carly

last: Tymm

overall_design: Run 1/Slide 1  • Three pieces of cardiac tissue across 3 mice:   1. Isoproterenol (ISO)-treated A/J mouse (160 mg/kg, twice for 2 weeks, endpoint one week after last injection)   2. ISO + ICI (ISO 160 mg/kg twice for 2 weeks, then biweekly 400 µg aPD1 + 400 µg aCTLA4 for 2 weeks)   3. ICI-only (biweekly 400 µg aPD1 + 400 µg aCTLA4 for 2 weeks)   Run 2/Slide 2  • Eight pieces of tissue across 6 A/J mice:   o ICI-only: biweekly 400 µg aPD1 + 400 µg aCTLA4 for 2 weeks   o ISO + ICI: ISO 160 mg/kg twice for 2 weeks, then biweekly 400 µg aPD1 + 400 µg aCTLA4 for 2 weeks   FOV assignments  • AJ ICI M1 (Run 1): FOVs 72–143   • AJ ICI M2 (Run 2): FOVs 1, 6–8, 16–19, 28–32, 41–46, 54–58, 64–68, 76–80, 90–91, 105–118   • AJ ICI M3 (Run 2): FOVs 2–5, 9–15, 20–27, 33–40, 47–53, 59–63, 69–75, 81–89, 92–104, 192–239   • AJ ICI M4 (Run 2): FOVs 119–191   • ISO M1 (Run 1): FOVs 1–71   • ISO + ICI M1 (Run 1): FOVs 144–177   • ISO + ICI M2 (Run 2): FOVs 240–271   • ISO + ICI M3 (Run 2): FOVs 272–352   • ISO + ICI M4 (Run 2): FOVs 353–428

first: Tymm

last: Carly

first: Bukhari

last: Syed


title: Glabridin Promotes Melanosome Degradation and Alleviates Melanosome-Induced Mitochondrial Dysfunction in Keratinocytes via Autophagy

city: Guangzhou

first: xin

last: nie

address: Room 1101-03, Building 22, CAS Changchun Institute of Applied Chemistry Huangpu Advanced Materials Research Institute Park, No. 388 Lianyun Road, Huangpu District

institute: Guangzhou Qingnang Biotechnology Co., Ltd.

summary: Background: The excessive accumulation of melanosomes within keratinocytes contributes to skin hyperpigmentation and disrupts normal epidermal proliferation. Glabridin, an isoflavone derived from Glycyrrhiza glabra L., is a potent skin-lightening agent with tyrosinase-inhibitory activity. This study aims to further elucidate the skin-lightening mechanism of glabridin by investigating its effects on melanosome degradation. Methods: We investigated the effect of glabridin on melanosome degradation by measuring melanin content and PMEL17 protein levels. Autophagy activity was evaluated by mRNA sequencing, Western blot, and immunofluorescence. In vivo depigmenting efficacy was examined using epidermal skin equivalents and zebrafish embryos. Additionally, ATP content and mitochondrial membrane potential were measured to determine whether glabridin ameliorates melanosome-induced mitochondrial dysfunction. Results: Glabridin promoted melanosome degradation in keratinocytes and exhibited depigmenting effects in both pigmented epidermal skin equivalents and zebrafish embryos. Mechanistically, glabridin activated autophagy, and pharmacological inhibition of autophagy blocked its depigmenting effects. Additionally, we found that melanosome accumulation impaired mitochondrial function in keratinocytes, an effect that was reversed by glabridin. Conclusions: These findings demonstrate that glabridin promotes melanosome degradation via autophagy activation, which is associated with improved mitochondrial function, providing mechanistic insight into its skin-lightening effects.

overall_design: To explore the mechanism by which glabridin promotes melanosome degradation, mRNA sequencing was conducted on three groups: control HaCaT cells, melanosome-laden HaCaT, and melanosome-laden HaCaT treated with glabridin.

first: Nie

last: Xin


title: RNA-seq of mouse suprarenal aorta from VAMP8-knockout and wild-type mice with AngII or saline infusion

city: Lexington

first: Shayan

last: Mohammadmoradi

state: KY

address: 741 South Limestone, Saha CVRC BBSRB B364B

department: Saha Cardiovascular Research Center

institute: University of Kentucky

summary: VAMP8 (vesicle-associated membrane protein 8) is a SNARE protein implicated in exocytotic secretion from immune and vascular cells. To investigate the role of VAMP8 in the vascular transcriptomic response to AngII infusion, we profiled gene expression in suprarenal aortae of VAMP8-knockout (Vamp8-/-) and wild-type C57BL/6 mice receiving 5-day continuous subcutaneous infusion of angiotensin II (AngII, 1,000 ng/kg/min) or saline via osmotic minipump. Four experimental groups were generated: WT+Saline, WT+AngII, V8KO+Saline, and V8KO+AngII (n=4 per group). Differential expression analysis identified transcriptomic changes associated with AngII treatment, VAMP8 deficiency, and their interaction in the aortic wall.

overall_design: Male Vamp8-/- and wild-type C57BL/6 mice received subcutaneous osmotic minipump implantation delivering saline or AngII (1,000 ng/kg/min) for 5 days (n=4 per group, 4 groups). Suprarenal aortae were isolated and processed for poly-A enriched paired-end mRNA sequencing on the Illumina NovaSeq X Plus. Differential expression was performed using DESeq2.

first: Mohammadmoradi

last: Shayan

first: Whiteheart

last: Sidney


title: Discovery of a primed endothelial progenitor that requires VEGF/ERK inhibition to complete vein differentiation [kairos_timecourse_bulkrnaseq]

first: Lay Teng

last: Ang

overall_design: HPSCs were differentiated into primitive streak, lateral mesoderm, artery ECs, primed ECs, vein ECs. Several EC samples were FACS for CD144+ before RNA-seq

first: Sherry Li

last: Zheng

first: Kevin J

last: Liu

first: Anastasiia

last: Masaltseva

first: Kyle M

last: Loh


title: Discovery of a primed endothelial progenitor that requires VEGF/ERK inhibition to complete vein differentiation [kairos_timecourse_scrnaseq]

overall_design: HPSCs were differentiated into primitive streak, lateral mesoderm, artery ECs, primed ECs, vein ECs. Several ECs samples were FACS for CD144+ before scRNA-seq.


title: Transcriptomic and Functional Analysis of Fibroid Extracellular Vesicles

city: torrance

first: Omid

last: Khorram

address: 1124  w carson st

institute: The lundquist Institute at UCLA medical center

summary: This study aimed to characterize large and small RNA transcriptomic changes in extracellular vesicles from cultures of fibroid explants and matched myometrial explants.

overall_design: Analysis of large and small RNA transcriptomic changes from RNA-seq data in three sets of extracellular vesicles from cultures of fibroid explants and matched myometrial explants.

first: Tsai-Der

last: Chuang

first: Abigail

last: Wiseman

first: Gabriela

last: Alfaro

first: Shawn

last: Rysling

first: Nhu

last: Ton

first: Daniel

last: Baghdasarian


title: Salmonella SopB suppresses post-transcriptionally regulated cytokine  release to reduce early tissue inflammation and delay disease progression

city: Aachen

first: Kaiyi

last: Zhang

address: Pauwelsstraße 30

department: Medicine Microbiology

country: Germany

institute: Uniklinik RWTH Aachen

summary: Salmonella enterica subsp. enterica serovar Typhimurium (S. Typhimurium) manipulates cellular processes through the translocation of effector molecules into the host cell cytosol. Using a recently established neonatal S. Typhimurium infection model, we provide functional insights into how Salmonella outer protein B (SopB) suppresses early mucosal tissue inflammation and prolongs host survival. Mechanistically, SopB prevents a disintegrin and metalloprotease 17 (ADAM17) activation, plasma membrane translocation and the release of membrane-bound TNFa from enterocytes and reduces epithelial secretion of IL-18 via mTOR-controlled secretory autophagy. This abolishes the early epithelial transcriptional response and reduces immune cell recruitment and programmed cell death-mediated mucosal barrier disruption delaying disease progression. The immunosuppressive effect of SopB is independent of the C-terminally encoded phosphatidylinositol phosphatase and phosphotransferase activity but requires an intact N-terminal domain. Thus, SopB suppresses the early, post-transcriptional regulation of epithelial cytokine release in an inositol phosphatase-independent manner likely promoting pathogen transmission.

overall_design: Tnfr1 ko mice are infected on day 1 after birth with 100 CFU wild-type or SopB-deficient S. Tm. (both strains are provided by Prof. Michael Hensel at Osnabrück Univerisity, Germany). Enterocytes (IECs) were isolated from total small intestinal tissues using a previously established EDTA-based protocol at d1p.i. (Lotz et al., 2006). RNA was isolated from the epithelial cell pellet or homogenized intestinal tissue using TRIzol® according to manufacturer’s recommendations. The RNA samples were handed in to Genomics Core Facility at UKA.

first: Nour

last: Diab

first: Matthias

last: Schmitz

first: Mathias W.

last: Hornef


title: ROS-sensing Transcription Factor YchJ Regulates RssB-RpoS Pathway to Protect Salmonella Against Oxidative Attack by Macrophages

city: Jinan

first: wei

middle: wei

last: wang

address: No. 6699 Qingdao Road, Huaiyin District, Jinan City, Shandong Province

institute: Shandong First Medical University & Shandong Academy of Medical Sciences

summary: Bacterial pathogens must overcome oxidative stress to survive within host phagocytes. Although canonical systems such as OxyR are well characterized, alternative pathways remain poorly understood. Here, we identified YchJ, a conserved yet uncharacterized protein, as a central redox-sensitive transcription factor that coordinates a major antioxidant defense system in Salmonella independently of OxyR. Deletion of ychJ severely impaired bacterial survival under H2O2 stress and within macrophages. Proteomic analysis revealed that YchJ represses rssB, leading to RpoS accumulation and upregulation of key antioxidant enzymes, including SodC and KatE. Our results show that YchJ directly binds the rssB promoter as a transcription factor. Structural analysis revealed that ROS sensing by YchJ is achieved through reversible dimerization mediated by an intermolecular disulfide bond. This conformational switch enables a C-terminal basic-rich region of the dimer to recognize a palindromic sequence in the rssB promoter and repress rssB transcription. Dual-transcriptome analysis further confirmed that YchJ directly activates antioxidant defenses in Salmonella and significantly disrupts host pathways during intramacrophage infection. Our findings elucidate a previously unrecognized redox-sensing pathway essential for bacterial virulence and uncover a novel transcriptional mechanism controlling RpoS stability, thereby expanding our understanding of the stress-response regulation system in pathogenic bacteria.

sample_organism:
Mus musculus
Salmonella enterica

platform_organism:
Mus musculus
Salmonella enterica

overall_design: Dual RNA-seq of mus_musculus and  salmonella_enterica experiment and control group


title: Citraconate preserves T cell stemness and antitumor immunity

city: Suzhou

first: Ziyi

last: Luo

address: suzhou

institute: Chinese Academy of Medical Sciences & Peking Union Medical College

summary: Metabolic perturbations within the tumor microenvironment profoundly compromise the stem-like properties and functionality of CD8+ T cells. Elucidating the metabolic circuitry that maintains T cell stemness is therefore essential for rejuvenating tumor-infiltrating lymphocytes and enhancing immunotherapeutic efficacy. Here, we identify citraconate, an itaconate isomer, as a critical metabolite markedly depleted in CD8+ T cells under chronic antigen stimulation or hypoxia. Citraconate supplementation preserves stem-like properties, mitigates ferroptosis, and potentiates T cell-mediated antitumor efficacy. Mechanistically, citraconate sustains intracellular cAMP pool by suppressing phosphodiesterase (PDE1A/C) expression and maintaining mitochondrial integrity, thereby activating protein kinase A (PKA) signaling. This activation represses arachidonate-5-lipoxygenase (ALOX5) transcription, reducing arachidonic acid peroxidation. Clinically, reduced ALOX5 or PDE1A expression correlates with decreased T cell exhaustion and enhanced responses to immune checkpoint blockade (ICB) therapy. Our findings reveal the citraconate-mediated PDE1-cAMP-ALOX5 axis as a promising therapeutic target for potentiating cancer immunotherapy.

overall_design: RNA-seq profiling of CD8+ T cells with or without citraconate  to assess the impact of citraconate on the transcriptional regulation of CD8+ T cells in the presence or absence of OVA peptide stimulation

first: Wenhui

last: Li

first: Minmin

last: Ge

first: Minju

last: Ni

first: Lianjun


title: NAT10-mediated N4-acetylcytidine modification drives RNA splicing of PML to alleviate adipose-derived stem cell senescence and promote diabetic wound healing

city: Beijing

first: wuhan

last: wei

address: No. 33, Badachu Road, Shijingshan District

department: Plastic Surgery Hospital

institute: Chinese Academy of Medical Sciences

summary: Cellular senescence in adipose-derived stem cells (ADSCs) limits their application in diabetic wounds. Alternative splicing generates multiple isoforms that play critical roles in modulating stem cell senescence. N-acetyltransferase 10 (NAT10) regulates N4-acetylcytidine (ac4C) modification, which has been implicated in RNA splicing. As the role of NAT10 in ADSCs remains elusive, this study aimed to investigate the function of NAT10 in ADSC senescence and RNA splicing.

overall_design: Cellular senescence was assessed using senescence-associated β-galactosidase staining, western blot for senescence markers p21 and p16, and qRT-PCR analysis of senescence-associated secretory phenotype (SASP) expression. RT-PCR analysis was performed to explore the potential role of NAT10 in RNA splicing.

first: Wuhan

last: Wei


title: Sex-dependent alterations in motor cortex linking spinal cord injury to long-term neurodegeneration risk

city: West Lafayette

first: Chongli

last: Yuan

state: Indiana

address: 480 Stadium Mall Drive

institute: Purdue University

summary: Patients with spinal cord injuries often suffer from multiple complications, including cognitive impairments after recovering from the primary wound. The molecular mechanism linking spinal cord injury and brain abnormality, however, remains elusive and has been largely attributed to neuroinflammation. In this work, we assessed persistent compositional and molecular changes in the motor cortex at 30 days post injury, a commonly perceived chronic stage, using a rat T10 contusion injury model. Brain slices and neurons isolated from the motor cortex were assessed using immunohistochemistry and RNA-sequencing, respectively. We observed significant changes in cellular compositions in the motor cortex including a decreased neuron to astrocyte ratio, and alterations in the morphology of neurons and astrocytes, indicating changes in functionality. Comparison of transcriptomic data collected at a sub-acute stage, namely 7 days post injury, with that at 30 days post injury, identified persistent and de novo (genes uniquely altered at 30 dpi.) changes that occur primarily after recovery of the spinal cord injury, enriched for neuronal and synaptic function related pathways. Strikingly, male and female rats showed distinct transcriptomic alterations at 30 days post-injury, with males showing neuron-related changes and females showing alterations in metabolism- and extracellular matrix–related pathways. These molecular differences were accompanied by better functional recovery in females, as reflected by significantly higher BBB scores, suggesting sex-dependent molecular changes induced in the motor cortex. Collectively, our study lays the foundation for understanding sexual dimorphism in brains after spinal cord injury and provides a plausible connection between spinal cord injury and increased risk for neurodegeneration later in life.

overall_design: RNA-seq profiling of primary neurons enriched from rat motor cortex at Day7 and Day 30 post injury.

first: Junkai

last: Xie

first: Seth

last: Herr

first: Siyuan

last: Sun

first: Shichen

last: Wu

first: Han

last: Zhao

first: Xingjie

last: Ping

first: Xiaoming

last: Jin

first: Riyi

last: Shi


title: Pleuromutilins alleviate psoriasis via inhibition of hnRNP-U/TNF-α

city: Nanjing

first: Shuya

address: No.12 Jiangwangmiao Street, Xuanwu District, Nanjing, Jiangsu Province, China

institute: Hospital for Skin Diseases, Institute of Dermatology, PUMC

summary: We find that upregulation of hnRNP-U promotes psoriasis-associated inflammation, whereas epidermis-specific knockout of hnRNP-U in mice markedly attenuates psoriasiform dermatitis.

overall_design: Ccmparetive gene expression profing anelysis of RNA-seq data of Imiquimod-induced mice, including epidermis-specific Hnrnp-u knockout mice (n=4) and contrast mice (n=4).

first: Ruiyu

last: Xiang

first: Weijian

last: He

first: Hao

last: Shen

first: Yong

last: Yang


title: Enhanced endocrine–metabolic support and axonemal assembly in high-sperm-motility geese: insights from testicular cellular heterogeneity by scRNA-seq

city: Daqing

first: Hechuan

last: Wang

state: Heilongjiang

address: No.5 Xinfeng Road

institute: Heilongjiang Bayi Agricultural University

summary: We used 10× Genomics single-cell RNA sequencing to investigate the cellular and transcriptional mechanisms underlying sperm motility differences in goose testes. Spermatogenesis depends on the coordinated interaction between germ cells and somatic cells in the testicular microenvironment, but the cellular basis of sperm motility variation remains unclear in geese. Here, we generated a single-cell transcriptomic atlas of testes from high- and low-sperm-motility groups and identified the major germ-cell and somatic-cell populations involved in spermatogenesis. These findings provide new insights into the molecular regulation of avian spermatogenesis and establish a valuable resource for understanding the mechanisms underlying male fertility in geese.

overall_design: Following phenotypic evaluation of sperm motility, testicular cells from geese in the high- and low-motility groups were isolated and profiled using 10× Genomics single-cell RNA sequencing.

first: Kexin

last: Cong

first: Jiaxin

last: Yin

first: Ke

last: Jiang

first: Yunuo

last: Liu

first: Xiaofang

last: Ren

first: Ying

last: Zhang

first: Shengjun


title: Aβ suppresses oligodendrocyte differentiation in vivo or vitro via inflammatory activation and cytoskeletal contraction [RNA-seq2]

city: shanghai

first: Wenxiu

last: Dai

address: No. 746, Zhongshan Middle Road, Songjiang District, Shanghai

institute: Shanghai Jiao Tong University

summary: Aβ disrupts oligodendrocyte lineage cells through a dual mechanism inflammatory activation and cytoskeletal compromise leading to maturation blockade  and hypomyelination independent of axonal damage. These findings position oligodendrocyte dysfunction as a key component of AD white matter pathology and identify potential therapeutic targets.

overall_design: RNA-seq analysis of primary OPC in wild-type developing rats treated and untreated with Aβ.

first: Daopeng

last: Li

first: Liang


title: Simultaneous orthogonal cell engineering by a single CRISPR-Cas9 polyfunctional editor [amplicon-Seq]

city: Milano

first: Sara

last: Valsoni

state: MI

address: via Olgettina 60

department: TIGET

country: Italy

institute: SR-TIGET

summary: The parallel disruption of multiple genes coupled with targeted transgene insertion offers a powerful strategy for more effective and precise cell engineering. However, such orthogonal editing involves the induction of multiple DNA breaks, raising safety concerns related to the risks of chromosomal translocations. Here, we present a polyfunctional CRISPR-Cas9-based strategy that enables both transgene insertion and epigenetic silencing at distinct genomic loci in a single treatment without inducing reciprocal chromosomal translocations. This is accomplished through an optimized all-in-one epigenome editor equipped with a catalytically active Cas9, whose endonuclease activity is selectively disabled at epigenetically silenced loci by the use truncated gRNAs. As a proof of concept, we demonstrated that this platform enables efficient multi-locus editing, including functional replacement of the endogenous TCR with a tumor-selective one, targeted insertion of a prototypic CAR with either a selectable marker or an immunomodulatory receptor into a TCR locus or a ubiquitously expressed gene, and durable, multiplexed epigenetic silencing of clinically relevant genes in primary human T cells. Polyfunctional editing establishes a versatile and safe framework for orthogonal editing, broadening the scope of genome and epigenome engineering in cancer immunotherapy and beyond.

overall_design: Amplicon-seq for indels analysis of B2M TdTomato K-562 and  T cells treated with a poly-editor.

first: Deborah

last: Cipria

first: Tania

last: Baccega

first: Miriana

last: Rizzo

first: Piergiuseppe

last: Quarato

first: Alice

last: Reschigna

first: Rita

last: El Khoury

first: Martino

last: Cappelluti

first: Sandra

last: Ammann

first: Valeria

last: Mollica Poeta

first: Matteo

last: Conti

first: Pietro

last: Spinelli

first: Ivan

last: Merelli

first: Toni

last: Cathomen

first: Monica

last: Casucci

first: Angelo

last: Lombardo


title: Simultaneous orthogonal cell engineering by a single CRISPR-Cas9 polyfunctional editor [RNA-seq T-cells B2M/TET2/TGFBR2]

overall_design: Comparative analysis of transcriptome from T cells treated with dCas9-based fusion ETR, targeting the promoter of B2M or TET2 or TGFBR2 genes with  full length or truncated gRNAs.


title: Simultaneous orthogonal cell engineering by a single CRISPR-Cas9 polyfunctional editor [RNA-seq T-cells CD3D]

overall_design: Comparative analysis of transcriptome from T cells treated with dCas9-based fusion ETR, targeting regulatory regions of the CD3D gene with  full length or truncated gRNAs.


title: Simultaneous orthogonal cell engineering by a single CRISPR-Cas9 polyfunctional editor [bisulfite-Seq]

type: Methylation profiling by high throughput sequencing

overall_design: Targeted BS-sequencing for methylation analyses of selected genes in B2M TdTomato K-562 and T cells treated with a poly-editor.


title: IL-17 Signaling Mediates Inflammation-Induced Capillary Leakage in addition to TNFa

first: Christopher

middle: Thomas

last: Neullens

address: Pauwelsstraße 30, 52074

institute: RWTH Aachen University Medical Faculty

summary: Capillary leak is a hallmark of acute and chronic inflammatory conditions and represents a critical driver of organ dysfunction and poor clinical outcomes. Despite its clinical relevance, the mechanisms governing endothelial destabilization under inflammatory conditions remain incompletely understood. In this study, we investigated the endothelial response to inflammation using a dynamic human in vitro model, in which human umbilical vein endothelial cells (HUVECs) were cultured in the presence of lipopolysaccharide (LPS)–stimulated human peripheral blood mononuclear cells (PBMCs) to mimic systemic inflammation. Under these conditions, endothelial cells exhibited a pronounced inflammatory response characterized by loss of barrier integrity and the formation of capillary leak–like structures. Separated transcriptomic pathway analyses of HUVECs and PBMCs identified a strong activation of cytokine-driven signalling networks, with the IL-17 pathway emerging as a dominant regulator of endothelial dysfunction, alongside TNF and JAK–STAT signalling. Targeted pharmacological inhibition demonstrated that blockage of IL-17 signalling most effectively stabilized the endothelial layer and mitigated leak-like morphology compared to inhibition of other pathways. However, endothelial barrier function was only partially restored, indicating persistent dysfunction despite IL-17 pathway suppression. Together, these findings position IL-17 as a key driver of inflammation-induced capillary leak while highlighting the multifactorial nature of endothelial destabilization. This work underscores the necessity of addressing complex immune–endothelial interactions to develop effective strategies for preventing inflammation-induced vascular leakage.

overall_design: 3'mRNA Seq. from HUVECs and PBMCs that were incubated in presence and absence from each other for 24 hours in combinations including LPS-stimution, TNF-, anti-TNF- and anti-CTLA-4 treatments.

first: Maria

last: Cheremkhina

first: Stefan

last: Jockenhoevel

first: Tiana

last: Tietz

first: Christian

last: Cornelissen

first: Anja

last: Thiebes

first: Klaus

last: Tenbrock


title: Enhancer-directed gene delivery for digit regeneration based on conserved epidermal factors [scRNA-seq]

city: Madison

first: Jianhong

last: Ou

state: Wisconsin

address: 330 N Orchard Street

institute: Morgridge Institute For Research

summary: Limb loss remains a significant clinical challenge, but regenerative medicine approaches such as gene therapy offer a promising strategy to trigger endogenous regeneration programs. Optimal vector configurations and molecular targets for appendicular skeletal repair are not well defined. Here, we leveraged insights from species with a high endogenous capacity for appendage regeneration to design an enhancer-directed gene delivery platform that functions during mouse digit regeneration, a well characterized model for partial limb regeneration in mammals. Single-cell RNA sequencing of zebrafish caudal fin regeneration, combined with expression data in regenerating salamander limbs and mouse digit tips, implicated the SP family of transcription factors as conserved, epidermally-expressed mediators of appendage regrowth. Null mutants of Sp8 demonstrated impaired limb regeneration in salamanders, while conditional knockout of Sp6 and/or Sp8 in the mouse basal epidermis resulted in defective bony digit tip regeneration, involving an IL-17 mediated osteoclastogenic program. Spatiotemporally focused expression of FGF8, a known target of SP factors, using a zebrafish-derived tissue regeneration enhancer element via adeno-associated viral vectors, could partially rescue digit tip regeneration in SP knockout mice and accelerate digit regeneration in wildtype mice. Our results demonstrate a contextual gene therapy approach to address limb loss based on genes like Sp transcription factors conserved across multiple contexts of appendage regeneration.

overall_design: Cells were dissociated from caudal fin regenerates in krt1‑19e:h2az2a‑mCherry, krt4:LY‑EGFP zebrafish and sorted for endogenous fluorescence, followed by single-cell RNA-sequencing using the 10x Genomics Chromium Single Cell 3’ Reagents Kit v.2

first: David

last: Brown

first: Kenneth

middle: D

last: Poss


title: Expression of Atg8a in Somatic Follicle Cells Prevents Age-Associated Decline in the Drosophila Ovary

city: San Francisco

first: Todd

last: Nystul

address: 513 Parnassus Ave, Box 0412

institute: UCSF

summary: The ovary is one of the first organs to lose functionality with age. We found that aging of the Drosophila ovary is characterized by an accumulation of phenotypes in the somatic compartment, including failure of the follicle cells to encapsulate germ-cell cysts, an extended S phase, and increased DNA damage. In aged ovaries, follicle encapsulation defects are associated with the lack of a germ-cell cyst checkpoint in early oogenesis. Single-cell RNA sequencing revealed that, across all cell types in the ovary, cells in the follicle lineage have the highest number of differentially expressed genes. Overexpression of Atg8a, a key autophagy machinery gene homologous to mammalian LC3, specifically in follicle cells prevents age-associated decline in the follicle epithelium and loss of reproductive capacity. Collectively, these findings demonstrate that genetic manipulation of a small population of ovarian somatic cells is sufficient to improve both cell-autonomous and non-autonomous features of reproductive aging.

overall_design: Single-cell RNA sequencing of Drosophila ovaries at 1 week and 6 weeks post-eclosion.

first: Wolfgram

last: Emily

first: Nystul

last: Todd


title: Enhancer-directed gene delivery for digit regeneration based on conserved epidermal factors

overall_design: Regenerating mouse digit tips were harvested and trimmed to exclude all tissue proximal to the distal interphalangeal joint. Digits were flash frozen in a mortar immersed in liquid nitrogen, then pulverized with a pestle. Powderized samples were then transferred to a Dounce homogenizer with Trizol (Invitrogen, CA) for an additional stage of homogenization. RNA was extracted from the Trizol mixture according to the manufacturer’s instructions, followed by clean up with an RNA Clean & Concentrator Kit (Zymo Research, CA).


title: Mutation-dependent responses to sleep and exercise in clonal hematopoiesis

city: New York

first: Darwin

last: D'Souza

address: 1470 Madison Avenue

department: ImmunoBio

institute: Human Immune Monitoring Center

summary: Somatic mutations that cause the clonal outgrowth of mutant hematopoietic stem cells (HSCs), known as clonal hematopoiesis (CH), decay myeloid heterogeneity, activate inflammation, and increase the risk of atherosclerosis. However, whether lifestyle alters myeloid cell diversity by either modulating CH clone expansion or the phenotypic programming of CH mutant cells, thereby impacting atherosclerosis, is unknown. Here, in humans and mice and across mutations in Jak2, Tet2, p53 and Dnmt3a, we demonstrate mutation-dependent responses to lifestyle in CH and show that mutant cells are uniquely and selectively sensitive to lifestyle exposures. Sleep and exercise restrict Jak2 and Tet2 CH and its atherogenic consequences by diminishing the clonal expansion of mutant HSCs and reprogramming mutant bone marrow (BM) HSCs and aortic macrophages, but not their neighboring wildtype (WT) counterparts, through divergent brain-body axes. In two large human datasets, moderate-to-vigorous physical associated with less non-DNMT3A CH. In atherogenic mice with Jak2 or Tet2 but not p53 or Dnmt3a CH, uninterrupted sleep or exercise curtails clone expansion. In Jak2 CH sleep and exercise do so by selectively reprogramming mutant, but not WT, BM HSCs towards anti-proliferative and metabolically healthy phenotypes by tempering BM macrophage-HSC IL-1β signaling. Sleep or exercise also lessens Jak2-, Tet2-, and p53-, but not Dnmt3a-, driven atherosclerosis, partly independent of BM clonal dynamics, by locally reprogramming mutant macrophages in atherosclerotic lesions. In Jak2 mutant, but not adjacent WT, aortic macrophages, uninterrupted sleep reduces cell death, selectively blunting CLEC4E-dependent inflammasome activation and IL-1β generation, consequently diminishing lesions. Exercise, meanwhile, activates PAC1+ neurons in the brain’s locus coeruleus (LC), raising peripheral norepinephrine (NE), and preserves the expression of NE’s adrenergic receptor (ADR) β2 in Jak2V617F, but not cohabitant WT, aortic macrophages. Exercise-mediated NE signaling from the brain’s LC to Jak2V617F aortic macrophages selectively represses their inflammatory programing and atherosclerosis. Together, our findings uncover gene-lifestyle coaction and establish that healthy lifestyles gene-specifically diminish CH and selectively reprogram mutant HSCs and macrophages to maintain myeloid heterogeneity and cardiovascular health.

overall_design: Female Ldlr-/- mice were irradiated and reconsituted with 90% CD45.1WT BM cells and either 10% CD45.2 WT BM cells or 10% Jak2V617Ffl/wtMx1Cre BM cells. Eight weeks later, mice were injected with Poly(I:C) and fed a high cholesterol diet. Ldlr-/-WTBM mice remained sedentary while Ldlr-/-Jak2V617FBM mice remained sedentary or were exposed to SF or exercise for 12 weeks. Aortas were isolated and CD45+ cells were sorted. BM was isolated and mature leukocytes depleted. Cell suspentions were oligotagged for CD45.1 and CD45.2 before scRNAseq. Protocol details can be found in the published manuscript.

first: Teresa

last: Gerhard

first: Zhihong

last: Chen

first: Cameron

last: McAlpine


title: Sex- and hepatocyte Pparg-dependent hepatic effects of a high fat diet containing 60% kcal from fat and 2% cholesterol, which was supplemented with 10% fructose in the drinking water (HFC+Fr) to induce diet-induced obesity in male and female mice.

city: Chicago

first: Jose

last: Cordoba-Chacon

state: IL

address: 835 S Wolcott Ave

department: Medicine

institute: University of Illinois at Chicago

summary: Most western diets or Amilyn-based diets induce MASLD and MASH in mice but do not fully reproduce metabolic dysfunction in male and female mice. In this study, a high fat diet containing cholesterol (D12120101, Research Diets, Inc) supplemented with 10% w/v fructose in the drinking water (HFC+Fr diet) was provided to control and hepatocyte specific Pparg knockout male and female mice for 24 weeks. Both HFC+Fr-fed male and female mice developed obesity and insulin resistance. Only male mice developed clear signs of MASH with fibrosis. Hepatocyte specific Pparg had a significant effect in the regulation of hepatic transcriptomics in HFC+Fr-fed male mice.

overall_design: In this study, we used a cohort of chow-fed nine to eleven-week-old Pparg floxed littermate male and female mice. These mice were treated with AAV vectors to generate control or hepatocyte specific Pparg knockout mice. Two weeks later, a subset of control and hepatocyte specific Pparg knockout mice were switched to a HFC+Fr diet for 24 weeks. A group of low-fat fed control male and female littermate mice served as baseline controls. We have six groups of mice, and their liver samples were included in this report as follow: Low fat diet male livers (n=4; M-LF-C); Low fat diet female livers (n=5, F-LF-C); High fat, cholesterol diet male control liver (n=6, M-HF-C); High fat, cholesterol diet female control liver (n=5, F-HF-C); High fat, cholesterol diet male hepatocyte Pparg knockout liver (n=5, M-HF-KO); High fat, cholesterol diet female hepatocyte Pparg knockout liver (n=5, F-HF-KO).

first: Marta

last: Sierra-Cruz

first: Izabela

last: Hawro

first: Samuel

last: Lee

last: Muratalla


title: The intestinal microbiota impacts nutritional immunity and resistance to Acinetobacter baumannii pneumonia

city: Nashville

first: Jennifer

last: Sucre

state: TN

address: 2215 B Garland Ave, 1125 Light Hall

department: Division of Neonatology, Department of Pediatrics

institute: Vanderbilt University Medical Center

summary: Broad-spectrum antibiotics are frequently administered prophylactically to intensive care unit patients as part of empiric care. This treatment has been associated with subsequent infections by the emerging nosocomial pathogen Acinetobacter baumannii; however, the mechanisms underlying this association remain unclear. Here, we demonstrate in a murine model that prophylactic broad-spectrum antibiotic administration drives susceptibility to intranasal infections with A. baumannii, and that reconstitution of the intestinal microbiota by fecal microbiota transplant restores control of infection, implicating microbiota dysbiosis as a key driver of pulmonary disease. Using single-cell RNA sequencing, we determine that prophylaxis suppresses phagocyte effector functions in the lung, including nutritional immunity pathways that restrict pathogen access to essential nutrient metals. Depletion studies identify neutrophils and inflammatory monocytes as central mediators of microbiota-dependent protection, and infection of mice lacking nutritional immunity components lipocalin-2 or calprotectin abrogates the effects of prophylaxis, establishing a causal linkage between microbiota dysbiosis and impaired phagocyte-mediated nutritional immunity. Together, these findings provide a mechanism for the increased severity of A. baumannii pneumonia following antibiotic exposure and highlight the intestinal microbiota as a potential therapeutic target to prevent future nosocomial infections with this and other healthcare-associated pathogens.

overall_design: Single Cell RNAseq of dissociated adult C57BL/6 mouse lungs (8-12 weeks of age). Mice were in one of four treatment groups: 1. Mock infection, normal water. 2. Mock infection, antibiotic prophylaxis (ampicillin, vancomycin, gentamicin) prior to infection. 3. Infected with Acinetobacter baumannii, nornal water. 4. Infected with  Acinetobacter baumannii, with antibiotic prophylaxis. Suspensions were enriched for cells that were CD45 negative, Ter119 negative, and viable.

first: Erin

last: Green

first: Nicholas

last: Negretti

first: Nicolas

last: Shealy

first: Tess

last: Brunner

first: Felipe

last: Moser

first: Sydney

last: Drury

first: Kacie

last: Traina

first: Valeria

last: Reyes Ruiz

first: Tzushan

last: Yang

first: Eric

last: Pamer

first: Mariana

middle: X

last: Byndloss

first: Raf

last: van de Plas

first: Joseph

last: Zackular

middle: H

last: Light

last: Skaar


title: Bulk RNA sequencing of liver NK cells and ILC1s from T-bet WT vs. inducible Ncr1-T-bet Δ/Δ mice

city: Saint Louis

first: Todd

last: Fehniger

state: Missouri

address: 660 South Euclid Avenue, Campus Box 8007

institute: Washington University School of Medicine

summary: The role of the T-box transcription factors T-bet and Eomes in innate lymphoid cells (ILCs) beyond their development is not well understood. We generated an inducible, NKp46 (Ncr1)-specific T-bet floxed knock-out mouse (Ncr1-T-betΔ/Δ) model. Bulk RNA sequencing of liver NK cells after tamoxifen treatment from T-bet WT (NKp46-CreERT2 x T-bet fl/fl) vs. Ncr1-T-bet Δ/Δ (NKp46-CreERT2 x T-bet fl/fl) mice had minimal transcriptional changes, whereas T-bet-deficient ILC1 exhibited significant gene expression changes. This sequencing data along with their rapid loss in vivo in Ncr1-T-betΔ/Δ mice reveal the requirement for continuous T-bet expression in liver ILC1.

overall_design: T-bet WT mice and T-bet Δ/Δ mice were treated with 1mg tamoxifen by oral gavage for 2 days (Day 1-3), then on Day 3, liver NK cells (YFP+NK1.1+CD49a-CD49b+) and ILC1 (YFP+NK1.1+CD49a+CD49b-) from the same mice were flow sorted. Qiagen RNA easy micro kit was used to extract RNA of cells directly sorted into Buffer RLT + β-mercaptoethanol, following manufacturer’s protocol. Sequencing library was prepared using the SMARTer Ultra Low RNA kit for Illumina Sequencing (Takara-Clontech) and sequenced on Illumina NovaSeq X Plus.

first: Fehniger

last: Todd

first: Wong

last: Pamela

first: Paik

last: Christy

first: Foltz

last: Jennifer


title: MRE11 proximal polyadenylation site-mediated looping impacts transcription and genomic stability

city: Boston

first: Dipanjan

last: Chowdhury

state: MA

address: 4 Blackfan Circle

institute: Dana-Farber Cancer Institute

summary: Alternative polyadenylation (APA) generates transcript isoforms with variable 3' UTR lengths, but its regulatory role in DNA damage response (DDR) genes remains poorly understood. Here, we demonstrate that the proximal polyadenylation site (pPAS) of MRE11 engages in PAS-promoter looping to facilitate RNA polymerase recycling and maintain high promoter activity, a mechanism not well characterized in mammals. Deletion of the MRE11 pPAS disrupts this looping, reduces MRE11 transcription, impairs MRN complex levels, and phenocopies hypomorphic MRE11 mutations. MRE11pPAS-/- cells exhibit ectopic DNA replication and reduced viability under overgrowth conditions. EdU-seq revealed that aberrant DNA synthesis in MRE11pPAS-/- cells occurs primarily at intronic and intergenic regions. MRE11 ChIP-seq showed decreased binding at these regions, correlating with elevated DNA replication. Furthermore, several DDR genes with multiple PASs form PAS-promoter loops, suggesting a broader regulatory mechanism. Together, our findings establish the MRE11 pPAS as a critical noncoding element that maintains genome stability through transcriptional regulation via PAS-promoter looping.

overall_design: Crosslinked cells were sonicated and diluted 10-fold with ChIP dilution buffer and incubated with protein A and protein G dynabeads (1:1 mix) and primary antibodies at 4°C overnight. Antibody-bound DNA was subsequently washed with low salt wash buffer, high salt wash buffer, LiCl wash buffer once, and TE wash buffer twice, respectively. ChIPed DNA was reverse-crosslinked and purified for ChIP-qPCR analysis.

first: Kaimeng

last: Huang


title: Microarray expression profiling data for premalignant mammary epithelial cells with a K8+ luminal origin

first: Zhe

last: Li

address: 77 Avenue Louis Pasteur, Room 466

department: Medicine, Genetics Division

institute: Brigham and Women's Hospital

summary: This dataset contains gene expression profiles of premalignant mammary epithelial cells (MEC) with loss of either p53 alone, or loss of both p53 and RUNX1, or loss of both p53 and BRCA1. The premalignant MECs were generated in female mice carrying their corresponding conditional knockout alleles as well as a conditional YFP-reporter, upon intraductal injection of Cre-expressing adenovirus under the control of the Keratin 8 (K8) promoter (Ad-K8-Cre) to their mammary glands.

overall_design: Total RNAs from YFP+ mammary epithelial cells sorted from Trp53L/L or Trp53L/L;Runx1L/L or Trp53L/L;;Brca1 female mice carrrying the Rosa26-Stop-YFP conditional reporter 10-14 days after intraductal induction of Ad-K8-Cre adenovirus were prepared and subjected to microarray expression profiling.

first: Dongxi

last: Xiang


title: Phenylalanyl-tRNA synthetase balances longevity and immunity by downregulating endogenous mitochondrial double-stranded RNAs

city: Daejeon

first: Seung-Jae

middle: V

last: Lee

address: 291, Daehak-ro, Yuseong-gu, Daejeon, Republic of Korea

country: South Korea

institute: Korea Advanced Institute of Science and Technology

summary: Endogenous double-stranded RNAs (dsRNAs) are immunogenic self-molecules that drive aberrant immune activation under pathological conditions. Here, we show that dsRNAs and their regulation by dsRNA-binding proteins are key determinants of the fine balance between aging and immunity in Caenorhabditis elegans and cultured human cells. We find elevated levels of dsRNAs with organismal aging and cellular senescence. We identify a moonlighting function for phenylalanyl-tRNA synthetase, FARS-1/FARSA, as a key factor necessary and sufficient for extending lifespan by directly binding to and downregulating dsRNAs, in particular, mitochondrial dsRNAs. FARS-1/FARSA possesses a previously unrecognized dsRNA-binding domain that mediates these effects separately from its canonical role in translation. Genetic inhibition of fars-1/FARSA upregulates immune response-related genes and enhances innate immunity through regulating the levels of mitochondrial dsRNAs. Our study establishes that FARS-1/FARSA is an evolutionarily conserved dsRNA-binding protein that delays aging and promotes longevity by counteracting the accumulation of dsRNAs.

overall_design: RNA-seq profiling of wild-type N2 under control or fars-1 RNAi conditions

first: Jooyeon

last: Sohn

first: Moonhyeon

last: Jeon

first: Seokjin

last: Ham

first: Hanseul

first: JinA

last: Lim

first: JiSoo

last: Park

first: Jongsun

first: Yoonji

last: Jung

first: Gee-Yoon

first: Eunseok

last: Kang

first: Dajeong

last: Bong

first: Yoosik

last: Kim


title: The PLAGL1-KLF4-IRX5 axis promotes the osteogenesis of periosteal progenitors during mandible regeneration [ATAC-seq]

city: Shanghai

first: Jiahui

last: Du

address: Zhizaoju Road 639

summary: Transcription factor (TF) networks are pivotal in regulating the fate of stem/progenitor cells. However, the intricate regulatory mechanisms mediated by key TFs in adult periosteal stem/progenitor cells (PSPCs) remain poorly understood, hindering the development of targeted therapies for craniofacial bone regeneration. Here, we show that the imprinted TF PLAGL1 is critical for osteoblast differentiation in PSPCs. Furthermore, we identify that PLAGL1 transcriptionally activates TF Irx5 in collaboration with TF KLF4, which, in turn, induces the expression of downstream osteogenic genes.

overall_design: ATAC-seq profiling of periosteal stem/progenitor cells (PSPCs) from control and Ctsk-iCre;Plagl1fl/fl mice after 3 days of osteogenic induction


title: Bulk RNA-seq profiling of p53-deficient mammary tumors driven by Runx1 and/or Rb1 loss

summary: This data set includes gene expression profiles of Runx1/p53-null, Rb1/p53-null, and p53/Runx1/Rb1-null mammary tumors developed in their respective female mouse models carrying the corresponding conditional knockout alleles (for Runx1, Rb1, and p53). The tumors were induced from luminal mammary epithelial cells after intraductal injection of adenovirus expressing Cre from the Keratin 8 promoter.

overall_design: Gene expression profiles were obtained from mammary tumors from three groups of mice as descirbed in the summary. These tumors represented the final product of lineage-tracing experiments targeting luminal mammary epithelial cells carrying different conditional knockout alleles for Runx1, Rb1, and p53. The lineage tracing was initiated by intraductal delivery of luminal keratin 8 (K8) promoter-controlled Cre-expressing adenovirus (Ad-K8-Cre) into the mammary glands of mice harboring the corresponding conditional knockout alleles.

first: Sen

last: Han

first: Dongyi

last: Zhao


title: A stem cell-derived neuronal culture model for mouse species variation in neurite growth

city: Berkeley

first: Rachel

middle: B

last: Brem

address: 321 Koshland Hall

department: Plant and Microbial Biology

institute: UC Berkeley

summary: Stem cell-derived neurons represent a powerful experimental system for the study of axon growth and regeneration, cutting cost and technical challenges. We developed an in vitro system using stem cell-derived motor neurons to investigate phenotypic differences in neurite growth between mouse subspecies. Previously, neurons in CAST/EiJ mice (Mus musculus castaneus) were reported to regrow CNS axons after injury to a greater degree than other strains. Here, we assayed neurite outgrowth from stem cell-neurons in a fully in vitro system, in the abscence of injury signals. In this system, CAST/EiJ neurons grew longer axons than those of C57BL/6J and 129S6/SvEv origin. Additionally, CASTx129 hybrid neurons exhibit comparable neurite growth to CAST/EiJ, indicating a dominant genetic effect.

sample_organism:
Mus musculus
Mus musculus castaneus
Mus musculus domesticus

overall_design: To assess the transcriptional identify of the various cell lines used, we sequenced differentiated motor neurons. Samples were sequenced from CAST/EiJ, 129S6/SvEv, and hybrid CASTx129 lines.

first: N

last: Simon

first: M

last: Freier

first: J

last: Valencia

first: Y

first: D

last: Bautista

last: Dutton

first: R


title: Engineered cytokine-secreting cells to prevent foreign body response against implants

city: Houston

first: H. Courtney

last: Hodges

state: TX

address: 1 Baylor Plaza, BCM 229

department: Department of Molecular & Cellular Biology

institute: Baylor College of Medicine

summary: Foreign body response hinders the long-term functionality of implants such as microencapsulated cell therapies. To address this issue, we delivered encapsulated engineered cells that secrete targeted cytokines to mitigate fibrosis on microcapsules. Here, we quantify immune profile change due to these encapsulated cytokine-secreting cells in mice. We identified that IL10 prevents inflammatory response while IL12 impairs the TGF-beta pathway to mitigate fibrosis.

overall_design: Single-cell 5' RNA-seq (scRNA-seq) of the surrounding implant cells in the intraperitoneal cavity and spleens from C57BL/6 mice that were implanted with encapsulated engineered cells that secrete IL10 or IL12 cytokines (RPE-IL10 or RPE-IL12), encapsulated non-engineered cells (RPE-null), or microcapsules alone (sham).

first: Boram

first: Yuen San

last: Chan

first: Dilrasbonu

last: Vohidova

first: Amanda

last: Nash

first: H

middle: C

first: Omid

last: Veiseh


title: A local sympathetic-immune axis inhibits melanoma growth in mice by dictating adrenergic control

first: Daniel

last: Kutsovsky

address: 1300 York Ave

department: Biochemistry

institute: Weill Cornell Medicine

summary: The nervous system drives tumor growth directly through intra-tumoral axons and indirectly through the systemic action of hormones. Yet contexts where the nervous system inhibits tumor growth are less defined. Here, we performed optical reconstruction of axonal innervation in mouse models of cutaneous melanoma, revealing progressive innervation by sympathetic axons. Local depletion of these axons accelerates while local optogenetic activation slows melanoma growth, together consistent with these axons acting as a physiological growth brake. The sympathetic nervous system is typically associated with driving tumor growth through activation of β adrenergic receptors (ARs). Here we find that the initial tumor seeding conditions sensitizes melanomas from β AR driven growth promotion towards a2 AR driven growth inhibition. Mechanistically, axonal activation of a2 ARs restricts the number and distribution of pro-tumor myeloid cells, independent of T cell activity. Together, our data reveal context-dependent, bidirectional neural control of tumor progression.

overall_design: 3 melanomas each from mice treated with vehicle (DMSO) and resiniferatoxin (RTX); vehicle (ascorbic acid), and 6-hydroxydopamine (6OHDA). Tumors were dissociated and sent for scRNA-seq.

first: Tingting

last: Liu

middle: Y

first: Ethan

last: Earlie

first: Liangliang

last: Ji

first: Michael

last: Iskols

first: Shakti

last: Ramsamooj

first: Xavier

last: Dawkins

first: Marwa

last: Zerhouni

first: Alexander

last: Birbrair

first: Elena

last: Piskounova

first: Ming

middle: O

last: Li

first: Ashley

last: Laughney

first: David


title: The PLAGL1-KLF4-IRX5 axis promotes the osteogenesis of periosteal progenitors during mandible regeneration [CUT＆tag-Seq]

first: Jiahui

last: Du

overall_design: CUT&Tag-seq profiling of PLAGL1_HA, PLAGL1_IgG, H3K4me3, and KLF4_FLAG in periosteal stem/progenitor cells (PSPCs) following 3 days of osteogenic induction, with KLF4_FLAG analysis performed in both control and Ctsk-iCre;Plagl1fl/fl mice.


title: Colon cancer resistance to Integrated Stress Response (IRS) at a clonal level

city: Bethesda

first: Thomas

middle: Joshua

last: Meyer

state: Maryland

address: 41 Center Drive, Building 41, Room B620

department: National Cancer Institute

institute: National Institutes of Health

summary: We explored a gene-editing approach to deplete ephrinB2 from colorectal cancer cell clones using a Doxycyline (Dox)-inducible lentiviral vector encoding a short hairpin (sh) RNA for ephrinB2 silencing via RNA interference. We discovered that shRNA-mediated gene silencing induced massive cell death in 596-7 cells (Ha T, et al. Antisense transcription from lentiviral gene targeting linked to an integrated stress response in colorectal cancer cells. Mol Ther Nucleic Acids. 2022;28:877-891.), with a small portion of cells maintaining viability.

overall_design: RNA expression in a clone of HT-29 cells (596-7 and derived 596-7-R1 clone)

first: Giovanna

last: Tosato

first: Taekyu

last: Ha

first: Hidetaka

last: Ohnuki

first: Lili

first: Parthav

last: Jailwala

first: Justin

last: Lack


title: FTO Inhibition Depletes Cysteine Metabolism and Sensitizes NSCLC to Radiation Therapy

city: Stanford

first: Erinn

last: Rankin

state: California

address: 269 Campus Dr

department: Rad Onc

institute: Stanford University

summary: Non-small cell lung cancer (NSCLC) remains a leading cause of cancer mortality, underscoring the urgent need for novel therapeutic strategies. Metabolic rewiring is a major cause of therapeutic resistance in tumors. Targeting cysteine metabolism is an attractive strategy due to its crucial role in maintaining oxidative stress. Here we demonstrate that FTO promotes NSCLC growth and survival by orchestrating cysteine metabolism through transcriptional regulation of the cystine transporter SLC7A11 and the de novo transsulfuration pathway enzymes cystathionine β-synthase (CBS) and cystathionine γ-lyase (CTH). Genetic and pharmacological inhibition of FTO reduced cystine uptake and de novo cysteine synthesis, leading to depleted intracellular glutathione (GSH), elevated reactive oxygen species (ROS), and reduced tumor growth. We further show that FTO inhibition increases radiation-induced DNA damage and sensitizes NSCLC cells and xenograft tumors to radiation therapy (RT). These findings establish FTO as a pivotal epitranscriptomic regulator of cysteine metabolism and radiation response in NSCLC, revealing a novel metabolic vulnerability that can be exploited therapeutically. Targeting FTO presents a promising strategy to enhance the efficacy of RT and improve outcomes for patients with NSCLC.

overall_design: H1299 cells were transduced with shRNA containing virus particles (twice) and generated stable KD and WT lines (1 week post infection). RNA was extracted and sent it for sequencing at Novogene.

first: Nishanth

last: Kuganesan


title: Simultaneous orthogonal cell engineering by a single CRISPR-Cas9 polyfunctional editor [RNA-seq K562]

first: Sara

last: Valsoni

address: via Olgettina 60

summary: The parallel disruption of multiple genes coupled with targeted transgene insertion offers a powerful strategy for more effective and precise cell engineering. However, such orthogonal editing involves the induction of multiple DNA breaks, raising safety concerns related to the risks of chromosomal translocations. Here, we present a polyfunctional CRISPR-Cas9-based strategy that enables both transgene insertion and epigenetic silencing at distinct genomic loci in a single treatment without inducing reciprocal chromosomal translocations. This is accomplished through an optimized all-in-one epigenome editor equipped with a catalytically active Cas9, whose endonuclease activity is selectively disabled at epigenetically silenced loci by the use truncated gRNAs. As a proof of concept, we demonstrated that this platform enables efficient multi-locus editing, including functional replacement of the endogenous TCR with a tumor-selective one, targeted insertion of a prototypic CAR with either a selectable marker or an immunomodulatory receptor into a TCR locus or a ubiquitously expressed gene, and durable, multiplexed epigenetic silencing of clinically relevant genes in primary human T cells. Polyfunctional editing establishes a versatile and safe framework for orthogonal editing, broadening the scope of genome and epigenome engineering in cancer immunotherapy and beyond.

overall_design: Comparative analysis of transcriptome from K-562 cells treated with dCas9-based fusion ETR, targeting the B2M promoter with full length or a truncated gRNA.

first: Deborah

last: Cipria

first: Tania

last: Baccega

first: Miriana

last: Rizzo

first: Piergiuseppe

last: Quarato

first: Alice

last: Reschigna

first: Rita

last: El Khoury

first: Martino

middle: A

last: Cappelluti

first: Sandra

last: Ammann

first: Valeria

last: Mollica Poeta

first: Matteo

last: Conti

first: Pietro

last: Spinelli

first: Ivan

last: Merelli

first: Toni

last: Cathomen

first: Monica

last: Casucci

first: Angelo

last: Lombardo


title: Ribonuclease DIS-3 promotes longevity by generating tRNA halves that inhibit translation via binding ribosomal proteins

first: Seung-Jae

last: Lee

summary: Transfer RNA (tRNA) halves (tRHs)  are generated via the cleavage of tRNAs and affect various biological processes, but their roles in aging and longevity remain poorly understood. Here, we determined the function of tRHs in aging by demonstrating that DIS-3/DIS3/RRP44 ribonuclease cleaved tRNAs for generating tRNA halves that promoted longevity. By performing a targeted genetic screen and subsequent biochemical analysis, we found that DIS-3/DIS3 catalyzed the generation of representative tRHs. We showed that dis-3 was crucial for longevity conferred by dietary restriction by upregulating SKN-1/NRF in the nematode Caenorhabditis elegans. Overexpression of 5′-tRH-Gln, a DIS-3 target, increased lifespan by reducing translation via binding ribosomal proteins. We further showed that mammalian DIS3 contributed to the generation of several tRHs and the prevention of premature cellular senescence. Overall, our data indicate that DIS-3/DIS3 is an evolutionarily conserved tRH-generating ribonuclease that counteracts aging.

overall_design: RNA-seq profiling of wild-type, dis-3(L75Q) mutant, eat-2(ad1116) mutant, and eat-2(ad1116); dis-3(L75Q) mutants.

first: Seokjun

last: Ha

first: Jisoo

last: Park

first: Hanseul

first: Donghun

first: Gee-Yoon

first: Seokjin

last: Ham

first: Seung-Jae V.


title: The placentalia specific histone H3.4 promotes germ cell development and reproductive fitness [RNA-seq]

city: Basel

first: Antoine

last: Peters

address: Maulbeerstrasse 66

country: Switzerland

institute: Friedrich Miescher Institute

summary: Genes involved in chromatin remodeling during sperm development are under strong evolutionary pressure to optimize reproductive fitness In mice, the histone H3.4 variant, encoded by the single copy H3f4 gene, is essential for spermatogenesis. H3.4 is highly expressed in spermatogonia and replaces progressively canonical H3 histones during spermatogonial expansion and differentiation. Phylogenetic analysis shows that H3f4 is specific to the placental lineage and originates from an ancestral H3.2 gene existing prior to the divergence of placental and marsupial mammals over 100 million years ago. Situated in a majorly truncated histone cluster, placental H3f4 orthologs show increased synonymous and non-synonymous substitution rates compared to analogous marsupial H3.2 genes. H3f4 carries four non-synonymous substitutions relative to H3.2 genes. To understand the impact of evolutionary sequence divergence on reproductive fitness, we reverted three of four non-synonymously substituted residues in H3f4 to those present in H3.1 (H3f4V24A, H3f4H42R, H3f4S98A) while leaving H3f4C96 intact which is identical to H3.1C96. When expressed from both alleles, neither single nor triple reversion of diverged residues affected spermatogenesis much. In contrast, hemizygous expression of the triply reverted H3f4H3.1 allele on a H3f4-deficiency background caused an >40% reduction in testis weight associated with death of pachytene spermatocytes, impaired differentiation of spermatids and aberrant expression of thousands of genes during spermatid elongation. Hemizygous expression of individual residue substitution alleles revealed that residues V24 and H42 of H3.4 promote spermatogenesis, while residue S98 is neutral. Together, our study shows that H3f4 has been subject to positive evolutionary selection, promoting male reproductive fitness.

overall_design: RNA-seq from FACS-isolated spermatogenic populations (ScLZ, ScPD, rSt and elSt) from mice with H3f4 gene being mutated to either encode for H3.1, or to contain single substitution mutations (H3.4V24A and H3.4H42R)

last: F.M. Peters

first: Pavel

last: A. Komarov


title: The placentalia specific histone H3.4 promotes germ cell development and reproductive fitness [ChIP-seq]

overall_design: ChIP-seq for histone variants (H3.4-MYC and H3.3) and for histone hodifications (H3K4me3 and H3K27me3) in FACS-isolated sparmetogenic populations (SgU, SgD, ScLZ, ScPD and rSt)


title: Compromised Transcriptional Elongation Stimulates Initiation in Human Cells [4sU-DRB-seq]

first: tian

last: tian

address: Dongchuan Road

institute: Shanghai Jiao Tong University

summary: P-TEFb is a well-established positive regulator of pause release of Pol II. The human PAF1 complex (PAF1C), consisting of PAF1, CTR9, CDC73, CTR9 and SKI8, plays critical roles in transcription and RNA processing, yet whether and how it regulates pause release is under debate. To address these issues, we further analyzed effects of PAF1 loss on pause release while using P-TEFb inhibition as a positive control. We found that besides increasing pause of Pol II, P-TEFb inhibition decreases and increases chromatin occupancy of PAF1C and Integrator, respectively. In contrast, acute PAF1 degradation increases the occupancy of Integrator and ARMC5, suggesting a positive role of the PAF1C in pause release by preventing promoter-proximal termination and Pol II degradation. Moreover, combined P-TEFb inhibition and PAF1 degradation increase pause of Pol II more than either treatment alone does, suggesting a cooperativity between them in pause release. Importantly, either P-TEFb inhibition or PAF1 degradation increases occupancy of initiation factors, suggesting that elongation perturbation may stimulate initiation, contrary to conclusions of three previous studies. Altogether, our results firmly established the PAF1C as a critical positive regulator of pause release and advanced current understanding of the connection between pause and initiation.

overall_design: 10 million cells were used in each 4sUDRB-seq experiment. To determine if the global reduction of transcription output was due to impaired productive elongation, PAF1-dTAG cells were treated with 100 µM DRB for 3 hours to clear elongating Pol II from gene body, and dTAGV-1 or DMSO was added 1 hour after the addition of DRB to initiate acute PAF1 degradation. Subsequently, the cells were washed twice with pre-warmed PBS to remove the inhibitory effect of DRB. Finally, the cells were treated with 1 mM 4-thiouridine (4sU) and cultured for 0, 10 and 20 min, respectively.

first: Ming

last: Yu

first: Ruoyu

last: Zhao

first: Tian

last: Tian


title: Single cell RNA sequencing of hippocampus and hypothalamus in AD mice with fructose-induced MetS and DHA and NR supplementation

city: Los Angeles

first: Xia

last: Yang

email: xyang123@ucla.edu, jading@ucla.edu

state: CA

address: 610 Charles E Young Dr S

institute: University of California Los Angeles

summary: Alzheimer’s disease (AD) is a pervasive neurodegenerative disorder with hallmarks of beta-amyloid (Aβ) plaques and neurofibrillary tangles. In light of the suboptimal benefit of Aβ clearing drugs to patients, the association between metabolic disorders and AD provides an alternative angle to understand the role of metabolic dysfunction in AD pathogenesis and the potential of modulating metabolism for improved therapeutic development. We investigated the effects of metabolic and nutritional modulators, namely, high fructose consumption as a potential risk, and docosahexaenoic acid (DHA) and nicotinamide riboside (NR) as potentially beneficial, on cell type specific transcriptomic responses in the hippocampus and hypothalamus of an AD mouse model. Fructose-induced metabolic syndrome increased expression of complement component 3 in ependymal cells and inflammatory genes in microglia, which were normalized by DHA and NR. NR and DHA led to reversals of Aβ signatures across cell types and targeted different aspects of microglial reactivity between AD and metabolically challenged AD. Our study supports the impact of metabolic regulation on neuroinflammation and provides molecular support for utilizing metabolic modulators for AD intervention.

overall_design: The 5xFAD mouse model of AD (MMRRC strain 34840) was used. At 2 months of age, 5xFAD mice were treated with high fructose (F), DHA, NR, DHA+F, or NR+F. A group of 5xFAD animals and wild type controls were untreated. scRNAseq was performed at 5 months.

first: Jessica

last: Ding