title: Decoding interactions in a soil microbiome Streptomyces community through multi-omics: functional insights into siderophore ecology

city: Liverpool

first: Francesco

last: Del Carratore

address: Crown Street

department: Department of Biochemistry, Cell and Systems Biology

institute: University of Liverpool

summary:
Background
Streptomyces are key contributors to soil microbiome function, known for their biosynthetic diversity. While advances in -omics technologies have improved our understanding of microbiome composition and metabolic potential, the mechanisms underpinning interspecies interactions remain poorly resolved. Here, we investigate the molecular basis of interactions among four sympatric Streptomyces soil microbiome isolates, focusing on phenotypic, metabolomic and transcriptomic responses.
Results
Co-culture experiments revealed that one isolate, strain A, exhibited pronounced phenotypic changes when grown alongside each of the other three strains. Untargeted metabolomics and RNA-seq analyses showed that strain A undergoes distinct metabolic and transcriptional shifts depending on its partner, with the strongest response elicited by strain C. Despite all four strains possessing a conserved desferrioxamine biosynthetic gene cluster, only strain C constitutively produced desferrioxamine B (DFO-B), a hydroxymate siderophore, indicating a role of iron bioavailability in the interaction. Supplementation with DFO-B or iron mimicked the growth stimulation of strain A observed in co-culture with strain C, and CRISPR base editing of desD in strain C abolished both DFO production and the phenotypic induction of strain A. However, transcriptomic profiles of strain A varied significantly depending on the partner strain, with distinct sets of biosynthetic gene clusters and metabolic pathways activated in response to strains B and C, suggesting additional cues beyond DFO-B. In contrast, strain D did not elicit growth stimulation in its partners, and itself showed downregulation of amino acid and carbon metabolism when co-cultured with strain C. These findings indicate that Streptomyces interactions are not only mediated by siderophore piracy but also involve complex, strain-specific molecular responses.
Conclusions
Our findings demonstrate that Streptomyces interactions are highly strain-specific and only partly mediated by siderophore piracy, with DFO-B acting as a potent interspecies cue. The divergent molecular responses to different partners suggest nuanced mechanisms of microbial sensing and competition. These insights advance our understanding of microbial crosstalk and highlight the ecological and evolutionary complexity of siderophore-mediated interactions. By integrating transcriptomics, metabolomics, and biochemical assays, we present a robust framework for dissecting microbial interactions, with implications for microbiome engineering and synthetic community design.

overall_design: RNA-seq profiling for 4 Streptomyces spp. Isolated from soil. Samples were collected considering axenic and co-culturing experiments in petri dishes at 3 data points (day 2, day 3 and day 4). Each sample contains a mixuture of the extracted RNA of the 4 isolates.

first: F

first: J

last: Connolly

first: K

last: Schmidt

first: A

last: Bisesi

middle: N

last: Martinson

last: Chua

first: M

last: Kuhs

last: Boneza

first: S

last: Heinsch

first: L

last: Kinkel

last: Smanski

first: W

last: Harcombe

first: R

last: Breitling

first: E

last: Takano


title: LPC 18:2-Driven Apoptosis In Neutrophils Is Non-Inflammatory and Lipid Raft Dependent

status: Public on Feb 13 2026

city: La Jolla

first: Sonia

last: Sharma

state: CA

address: 9420 Athena Cir

department: Center for Autoimmunity and Inflammation

country: USA

institute: La Jolla Institute of Immunology

type: Expression profiling by high throughput sequencing

summary: Lysophosphatidylcholines (LPCs) are potent bioactive lipids whose fatty acid composition dictates their immunomodulatory effects. Here, we delineate how unsaturated LPC 18:2 and saturated LPC 16:0 differentially regulate neutrophil survival and inflammatory programs. LPC 18:2 markedly increased reactive oxygen species (ROS) generation and caspase-3/7 activation, accompanied by Annexin V positivity, mitochondrial membrane depolarization, and cytochrome C release, f. Features consistent with intrinsic apoptosis. In contrast, LPC 16:0 induced robust LDH and HMGB-1 release, indicating membrane rupture and pyroptosis-like death. Bulk RNA sequencing revealed that LPC 16:0 strongly upregulated inflammatory and cytokine genes. Disruption of lipid-raft integrity abolished LPC 18:2–induced ROS and apoptosis, underscoring the dependence of these effects on membrane organization. Collectively, these results identify LPC 18:2 as a non-inflammatory, mitochondria-dependent inducer of neutrophil apoptosis, whereas LPC 16:0 promotes inflammatory, lytic death programs. These findings highlight how lipid saturation determines neutrophil fate and immune tone, providing mechanistic insight into how distinct LPC species shape inflammation and tissue injury.

overall_design: The experiment was conducted using Homo sapiens primary peripheral blood neutrophils. Cells were subjected to three treatment conditions: unstimulated (vehicle control), LPC 16:0 at 10 µM for 45 minutes, and LPC 18:2 at 10 µM for 45 minutes. A single time point of 45 minutes post-treatment was analyzed. Each condition included three independent biological replicates derived from distinct healthy donors, resulting in a total of nine RNA-seq libraries. Each donor served as a unique biological replicate, and within a given donor, only one treatment was applied per sample without any technical pooling.

first: Priyanka

last: Saminathan

first: Alicia

last: Gibbons

first: Ian

last: Mathews

first: Maija

last: Corey

first: Ashmitaa

last: Logandha Ramamoorthy Premlal

first: Namratha

last: Nadig

first: Camille

last: Fang

first: Mousa

last: Vatanmakanian


title: Microglia gene-expression after a transient ischemic attack (TIA)

city: München

first: Gemma

last: Llovera

address: Feodor-Lynen-Straße 17

country: Germany

institute: Institut für Schlaganfall- und Demenzforschung (ISD)

summary: Analysis of more than 700 genes using the Neuroinflammation panel (NS_Mm_NeuroInflam_v1.0) of Nanostring technologies from isolated microglial cells in naive and TIA animals revealed that they have an activated phenotype even 3d after TIA.

overall_design: In the study presented here, 3 animals per group and 65ng of microglial RNA per sample were used.


title: Eurycomanone from Eurycoma longifolia Jack roots regulates dopamine secretion via transsynaptic GABA signalling in differentiated SH-SY5Y human neuroblastoma cells: a transcriptomic study

city: Subang Jaya

first: Bin Seng

last: Low

state: Selangor

address: No 1, Jalan Taylor's,

department: School of Medicine

country: Malaysia

institute: Taylor's University

summary:
We employed microarray-based transcriptomic analysis to uncover potential molecular pathways associated with the dopaminergic effect of the quassinoid compound from the plant. The transcriptomic insights generated from this work may provide a deeper molecular basis for the long-debated aphrodisiac properties attributed to the plant
We used microarrays to analyse the gene expression and identified distinct classes of up-regulated and down-regulated genes during this process.

overall_design: SH-SY5Y human neuroblastoma cells were differentiated for RNA extraction and hybridization on Affymetrix microarrays. The arrays were washed, stained, and then scanned. A total of 3 dose-specific treated cell intensity files were generated and imported to Transcriptome Analysis Console for gene expression analysis.


title: A simple liquid 3D cell culture paradigm models oxidative mitochondrial metabolism of epithelial breast cancer cells with relevance for lung metastases

city: Frederick

first: Yongmei

last: Zhao

state: MD

address: Post Office Box B

department: Advanced Biomedical Computational Science

institute: Frederick National Labroatory for Cancer Research

summary: Three-dimensional (3D) cell culture systems have emerged as powerful tools to model tumor biology ex vivo. However, the diverse array of 3D culture methods available presents challenges in selecting the most appropriate model for specific research questions. This study provides a comparative analysis of breast cancer cells (SUM149, IBC-3, MDA-MB-468) in the mammosphere culture (SphC) model or an “emboli” culture (EmC) model, which enrich for cancer stem cells and epithelial features, respectively. The EmC model, designed originally for inflammatory breast cancer, is characterized by media viscosity and mechanical rocking of the culture vessel. Notably, cells in EmC showed a distinct and durable reduction in cell proliferation while demonstrating increased capacity to establish experimental lung metastases. Quantitative analysis of electron microscopy images suggested that cells in EmC acquire nuclear and mitochondrial features that resemble those of tumor tissue. Proteomics, single-cell transcriptomics, and metabolic flux analyses showed that cells in EmC and SphC favor mitochondrial oxidative metabolism (OXPHOS) and glycolysis, respectively. EmC rendered cells hypersensitive to OXPHOS inhibition, yet more resistant to oxidative stress. Several genes associated with lung metastasis, including ID1, were specifically enriched in EmC. Given the increasingly recognized role of OXPHOS in cancer cell survival during dissemination and as established metastases, we propose that the EmC paradigm is a suitable ex vivo model to study signaling pathways relevant for tumor tissue and to assess drug sensitivities and resistance mechanisms of metastatic breast cancer cells ex vivo

overall_design: SUM149 and IBC-3 cells were cultured in 2D, "emboli" culture (EmC) or mammosphere culture (SphC) for 3 days.

first: Balamurugan

last: Kuppusamy

first: Esta

last: Sterneck


title: Identification of target genes and regulatory networks for bone mineral density GWAS loci through systematic targeting and inhibition [hFOB ATAC-seq]

city: Toronto

first: John

middle: A

last: Morris

state: Ontario

address: 160 College Street

department: Donnelly Centre for Cellular and Biomolecular Research

country: Canada

institute: University of Toronto

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Osteoporosis is characterized by low bone mineral density (BMD) and elevated fracture risk. Most BMD-associated GWAS variants lie in noncoding regions, complicating efforts to identify causal genes and mechanisms. To overcome this variant‑to‑function challenge, we developed STING‑seq, a framework integrating biobank‑scale GWAS with single‑cell CRISPR inhibition (CRISPRi) screening to directly connect noncoding cis‑regulatory elements (CREs) to their target genes. Applied to human fetal osteoblast (hFOB) cells across osteogenic differentiation, STING‑seq linked 86 CREs to 71 target genes at BMD loci. Arrayed CRISPRi and activation validated key CRE–gene relationships, including long‑range enhancers regulating CXCL12 (over 500 kb apart). We further uncovered trans‑regulatory networks and characterized the DAP3–YY1AP1 bidirectional promoter, demonstrating a role for DAP3 in mineralization via mitochondrial pathways. Together, these findings provide mechanistic insight into how noncoding GWAS variants shape osteoblast activity and highlight the genes and pathways that mediate genetic effects on BMD.

overall_design: ATAC-seq profiling of hFOB 1.19 cells at days 0, 2, 4, and 10 of osteogenic differentiation.

first: Larry

last: Mesner

first: Mohammadjavad

last: Kahlani

first: Vedavathi

last: Madhu

first: Lucia

last: Wagner

first: Kristyna

last: Kupkova

first: Hannah

last: Pitas

first: Ana

last: Mello

first: Samuel

last: Ghatan

first: Tuuli

last: Lappalainen

first: Neville

last: Sanjana

first: Charles

middle: R

last: Farber


title: Reduced levels of inositol hexakisphosphate kinase (IP6K) impair life-cycle transitions and the intracellular development of Trypanosoma cruzi within human cardiomyocytes

city: São Paulo

first: Eduardo

middle: Moraes

last: Reis

state: SP

address: Av. Prof. Lineu Prestes, 748

department: Biochemistry

country: Brazil

institute: University of São Paulo

summary: Trypanosoma cruzi is a protozoan parasite etiological agent of the vector-borne disease American trypanosomiasis, also known as Chagas disease. During its life cycle, T. cruzi undergoes some fundamental processes: metacyclogenesis (transition from epimastigote to metacyclic trypomastigote form) – occurring in the invertebrate host, amastigogenesis (transition from trypomastigote to amastigote form) and trypomastigogenesis (transition from amastigote to trypomastigote form) – both occurring within nucleated cells of the mammalian host, and finally epimastigogenesis (transition from trypomastigote to epimastigote forms) – which occurs as soon as the invertebrate host ingests trypomastigote forms during the bloodmeal. The precise mechanisms that regulate these processes remain elusive but certainly depend on kinases. The canonical function of the inositol hexakisphosphate kinases (IP6Ks) is to phosphorylate the phosphate groups from inositol hexaphosphate (IP6), leading to the formation of inositol pyrophosphates (PP-IPs). However, recent studies have been describing non-canonical functions for this kinase family. Here, after disrupting a single IP6K allele of T. cruzi and confirming its downregulation, we observed that the life cycle of this parasite was profoundly impaired. Epimastigote forms of T. cruzi IP6K-deficient showed morphological alterations, increased population presenting a dormant-like state (quiescence), and a reduced differentiation capacity during metacyclogenesis. The restricted metacyclic forms of T. cruzi IP6K-deficient that were able to differentiate had reduced infective potential (invasion rate) in human cardiomyocytes. Interestingly, 120 h after infection, the amastigote forms of T. cruzi IP6K-deficient showed an impaired ability to transform into trypomastigotes, with most of the population egressing from human cardiomyocytes without completing trypomastigogenesis. Transcriptomic data show that the levels of surface proteins (mucins and trans-sialidases) were altered in both epimastigote and trypomastigote forms in response to the low levels of IP6K, which helps explain our findings. Finally, we observed that the few trypomastigote forms of T. cruzi IP6K-deficient that egressed from human cardiomyocytes could not complete the epimastigogenesis. Together, our findings strongly suggest that IP6K is critical to sustain the T. cruzi life cycle. Thus, as the disruption of both IP6K alleles did not generate viable parasites and the similarity relative to its human homolog is ~15%, this kinase could be a potential target for drug development against Chagas disease.

overall_design: RNA-seq profiling of control T. cruzi (CL Brener) lineage (parental T7/Cas9) and its IP6K single-knockout lineage (using epimastigotes and trypomastigotes forms).

first: Bryan

middle: E

last: Abuchery

first: Thaise

middle: L

last: Teixeira

first: Vitor

last: da Silva

first: Rauni

middle: B

last: Marques

first: Fernanda

middle: M

last: Gerolamo

first: Carolina

last: Catta-Preta

first: Bruno

last: Santarossa

first: Marija

last: Krasilnikova

first: Craig

last: Lepslay

first: Yete

middle: G

last: Ferri

first: Maria Cristina

last: Motta

middle: C

first: Miguel

last: Chiurrilo

first: Noelia

last: Lander

first: Simone

last: Calderano

first: Richard

last: McCulloch

first: Marcelo

middle: S


title: Transcriptome changes in C57BL/6 mice after ischemia/reperfusion injury

city: Zhengzhou City

first: Yang

last: Bai

state: Henan Province

address: 1 Jianshe Dong Road, Erqi District, Zhengzhou City, Henan Pr

country: China

institute: Zhengzhou University

summary: Renal warm ischemia-reperfusion (I/R) injury drives acute kidney injury and transplant dys-function through endoplasmic reticulum (ER) stress. Although Activating Transcription Factor 6 (ATF6) modulates ER stress resolution via the unfolded protein response, its functional role in re-nal IRI remains undefined. Using murine model of warm renal I/R and hypoxia-reoxygenation (H/R)-treated HUVECs/HK-2 cells, we systematically investigated the expression of ATF6 and molecular mechanisms through RNA-seq, ChIP, and dual-luciferase assays. IRI significantly up-regulated the expression of ATF6 in renal tissues, especially in proximal tubule epithelial cells. Functionally, ATF6 activation reduced Cr, BUN, and inflammatory cytokines, whereas genet-ic/pharmacological suppression aggravated renal tubular damage. Mechanistic studies revealed ATF6 transcriptionally represses FHL2 by direct promoter binding. The interaction between FHL2 and TRAF6 promoted the TRAF6/NF-κB signaling pathway, and the phosphorylation levels of p65 and IκBα were increased. ATF6 overexpression effectively counteracted FHL2-mediated NF-κB hyperactivation, establishing a protective axis where ATF6 mitigates inflammation via FHL2 sup-pression. This study identifies ATF6 as a renoprotective factor that suppresses FHL2 to inhibit TRAF6/NF-κB inflammation during IRI. The ATF6/FHL2/NF-κB axis provides mechanistic in-sights and therapeutic targets for ischemic renal injury and transplant complications.

overall_design: We randomly assigned C57BL/6N mice to sham-operated (Sham, n=6) or I/R (n=6) groups. The I/R group received pentobarbital sodium anesthesia followed by renal pedicle occlusion. After 60 minutes of ischemia, we restored blood flow for 6h reperfusion before euthanasia and sample collection

first: Xuan

last: Wu


title: LncRNA NAS1 deficiency drives cisplatin resistance in non-small cell lung cancer

city: nanjing

first: Zhang

last: Jun

state: jiangsu

address: longmiandadao

institute: China Pharmaceutical University

summary: Cisplatin resistance remains a major obstacle in non-small cell lung cancer (NSCLC) treatment. Long non-coding RNAs (lncRNAs) have been implicated in chemotherapy resistance, yet their specific roles and mechanisms are not fully understood. This study reveals that low expression of lncRNA NAS1 drives cisplatin resistance via NR2F1-mediated TGFB1/NF-κB signaling in NSCLC.

overall_design: Total RNA-seq profiling of three distinct cisplatin-resistant NSCLC cell lines (H226/DDPr, H460/DDPr, PC9/DDPr) and their corresponding parental cells.

first: Xianrong

last: Lin

first: Wenjun

last: Tao


title: Estrogen Receptor α Protects Against Obesity-Induced Metabolic Dysfunction by Regulating VAT Tregs (oVAT)

city: Atlanta

first: Gregory

middle: K

last: Tharp

state: GA

address: 954 Gatewood Dr

department: Developmental and Cognitive Neuroscience

institute: Yerkes National Primate Research Center

summary: Estrogens are important for metabolic health. Individuals with low levels of circulating estrogens, including men and postmenopausal women, exhibit an elevated risk for developing obesity-associated metabolic syndromes. Chronic low-grade inflammation in the visceral adipose tissue (VAT) is a major contributor to metabolic dysfunction during obesity. Regulatory T cells (Tregs) in the VAT limit tissue inflammation and protect against obesity-associated metabolic disease. In this study, we identify opposing roles of estrogen receptor α (ERα) in regulating VAT Tregs in female mice under steady-state and obese conditions. At steady state, ERα restrained the age-dependent clonal expansion of specific VAT Treg subsets expressing the IL-33 receptor ST2. However, during obesity, ERα-deficiency predisposed females to the loss of ST2+ VAT Tregs, exacerbating VAT inflammation and insulin resistance. These findings indicate that ERα signaling protects against obesity-induced metabolic diseases by preserving metabolically protective ST2+ VAT Treg subsets, and the loss of this protection may contribute to the heightened metabolic risk in estrogen-deficient individuals.

overall_design: ovarian visceral adipose tissue (oVAT) was isolated from 27-week-old Era+/+ or Era-/- female mice fed an NCD or HFD for 17 weeks. Stromal vascular fractions from each group were stained with antibodies targeting appropriate surface markers along with corresponding TotalSeq™ hashtag antibodies for downstream demultiplexing. CD4+ CD25hi Tregs from all groups were sort-purified into one tube containing RPMI 1640 with 10% FBS for 10X genomics 5' single cell RNA/TCR sequencing analysis.

first: Cody

last: Elkins

first: Jianliang

last: Zhang

first: Chengyu

last: Ye

first: Samara

last: Moll

first: M

last: Weitzmann

first: Chaoran

last: Li


title: TGF-β and IL-4+IL-13 induce neuroplasticity in an in vitro model of hPSC-derived sensory neurons

city: Groningen

first: Reinoud

last: Gosens

address: Antonius Deusinglaan 1

department: Molecular Pharmacology

country: Netherlands

institute: University of Groningen

summary: Asthma is a chronic inflammatory disease of the airways characterized by airway hyperresponsiveness. Recent studies using mouse models, bronchial biopsies of asthma patients, and lung tissue of patients with fatal asthma have implicated neuroplasticity as a pathological feature of severe asthma, possibly contributing to airway hyperresponsiveness. Currently, we lack the models to study the mechanisms behind sensory neuroplasticity in vitro. Here, we describe the differentiation of sensory neurons from human pluripotent stem cells. Generation of sensory neurons was validated using immunofluorescence, flow cytometry and RNA-sequencing, as well as functional responsiveness to capsaicin using calcium imaging. We exposed these hPSC-derived sensory neurons to TGF-β or Th2-cytokines IL-4 together with IL-13, both of which have important roles in asthmatic airway remodelling. Both treatments induced neuroplasticity-related changes such as increased network density and neuronal sensitivity in the sensory neurons, albeit TGF-β more strongly than IL-4+IL-13. Our results show that hPSC-derived sensory neurons can be generated robustly and reproducibly and provide a good model to investigate the mechanisms behind neuroplasticity. Furthermore, our findings support a role for the asthmatic airway microenvironment, most notably TGF-β, in the development of neuroplasticity.

overall_design: H9WAO9 human Pluripotent Stem cell (hPSC)-derived sensory neurons were used to study neuroplasticity development. Mature hPSC-derived sensory neurons exposed to TGF-β or IL-4+IL-13 for 5-days were subjected to RNA-sequencing.

first: Carli

last: Koster

first: Sophie

middle: T

last: Bos

first: Chiara

last: Lavitola

first: Mihaly

last: Balogh

first: Barbro

last: Melgert


title: RNA-SEQ of HRECs depleted with Norrin/beta-catenin signaling components or KIF11 or PRDX1

city: Chengdu

first: Shujin

state: Sichuan

address: Chengdu, Sichuan province, China, first-ring road, No.34

institute: Sichuan Provincial People's Hospital, Sichuan Academy of Medical Sciences

summary: We performed RNA-Seq on human microvascular endothelial cells (HRECs) with lentivirus-mediated knockdown of CTNNB1, LRP5, FZD4, TSPAN12, KIF11, and PRDX1. This dataset includes the gene expression profiles that reveal the specific gene expression changes that occur in HRECs following the knockdown of these key genes implicated in FEVR.

overall_design: Processed RNA-seq data were generated from human retinal endothelial cells (HRECs) that had been depleted of CTNNB1, LRP5, FZD4, TSPAN12, KIF11, or PRDX1. A shared control group was used for the CTNNB1, LRP5, FZD4, and TSPAN12 knockdown groups, while the KIF11 and PRDX1 knockdown groups each had their own control group.


title: Single-Cell transcriptomic profiling of mouse retinal vascular cells reveals mechanisms of familial exudative vitreoretinopathy.

summary: Familial Exudative Vitreoretinopathy (FEVR) is a rare, inherited disorder characterized by a failure of peripheral retinal blood vessels to develop fully.To investigate the pathogenic mechanisms of FEVR, we conducted single-cell RNA-sequencing of retinal vascular cells from two distinct mouse models: Tspan12 knockout mice and mice with an endothelial-specific conditional knockout of Ctnnb1, which encodes β-catenin. Our analysis of the cellular and molecular profiles of these cells provides unprecedented insight into how disruption of key components of the Norrin/β-catenin pathway contributes to the pathology of FEVR.

overall_design: Three samples from  wild-type (WT) mice, Tspan12 knockout (KO) mice, and Ctnnb1 conditional knockout (cKO) mice.


title: FOXA1/FOXM1 ChIP-seq on giredestrant-resistant ER+ breast cancer cell lines

city: South San Francisco

first: Jackson

last: Liang

address: 1 DNA Way

department: Discovery Oncology

institute: Genentech

summary: Characterization of molecular features associated with giredestrant resistance in HR+ breast cancer cell line models

overall_design: HR+ breast cancer cells were cultured in giredestrant for 4-months until they acquired resistance to giredestrant. Cells were then profiled via ChIP-seq against FOXA1 and FOXM1.

first: Bence

last: Daniel

first: Vasumathi

last: Kameswaran

first: Ciara

last: Metcalfe


title: Protective efficacy and related mechanism of deinoxanthin in experimental animal and cell models of periodontitis

city: Jeonbuk

first: JungChae

last: Lee

address: 567, Baekje-daero, Deokjin-gu, Jeonju-si, Jeollabuk-do, Republic of Korea

country: South Korea

institute: Jeonbuk National University

summary: As periodontitis is a chronic inflammatory disease resulting in periodontal tissue destruction and tooth loss, studies have tried to development the biomaterials that effectively prevent inflammatory response and oxidative stress without side effects. In this work, we explored whether Deinococcus radiodurans-derived deinoxanthin (DEIX) protects against alveolar bone loss and connective tissue degradation in an experimental rat model of periodontitis investigated the related mechanisms. Oral supplementation with DEIX (25 mg/kg body weight, once per day for 14 consecutive days) protected rats against ligature-mediated loss of alveolar bone and connective tissues. That protection involved the DEIX-induced restoration of the ligature-stimulated disorders, including overproduction of inflammatory mediators, accumulation of reactive oxygen species, and imbalance between osteoclast and osteoblast activity in the inflamed periodontium. In vitro experiments using human periodontal ligament cells (hPDLCs) and THP-1 cells supported the mechanisms by which the direct addition of DEIX (20 μM) recovers lipopolysaccharide (LPS, 2 μg/mL)-stimulated inflammatory responses in the cells. RNA-sequence profiling from the hPDLCs further supported the potency and the related mechanisms of DEIX to protect cells from LPS-mediated inflammatory damages. Collectively, this study introduces the protective role of DEIX in inflammatory periodontal tissue destruction and supports its clinical usefulness for patients who suffering from chronic periodontitis.

overall_design: Total RNAs were isolated from human-derived periodontal ligament cells (hPDLCs) for RNA-sequence profiling. In this study, the hPDLCs were exposed to 2 μg/mL lipopolysaccharide (LPS) for seven days followed by the additional-14 days incubation in combination with 20 μM DEIX or not before the RNA extraction. There are four groups: 1) non-treated control hPDLCs (Control-C1-I, C1-II, C1-III), 2) LPS treatment alone (LPS-C2-I, C2-II, C2-III ), 3) LPS and DEIX treatment (LPS+DEIX-C3-I, C3-II, C3-III), and 4) DEIX treatment alone (Only DEIX-C4-I, C4-II, C4-III).

first: Jung-Chae


title: HE4 regulates anticancer immune evasion by upregulating PD-L1 expression on macrophages through the IFN-γR-STAT3 axis  [L1MKK]

first: Bo

last: Zeng

address: No.4, Section 2, North Jianshe Road, Chengdu

department: school of medicine

institute: University of Electronic Science and Technology of China

summary: Current immune checkpoint inhibitors (ICIs) targeting PD-L1/PD-1 have achieved great success in clinical cancer treatment. However, it still faces great challenges, such as a low overall response rate and immune-related adverse events (irAEs). PD-L1 is transcriptionally induced by immune cell-secreted cytokines, such as IFN-γ, in the tumor microenvironment (TME), but whether the expression of PD-L1 is regulated by cancer cell-derived molecules is still unclear. Here, we show that HE4 is another critical transcriptional regulator of PD-L1 on macrophages in the TME through the IFN-γR1/IFN-γR2-JAK-STAT3 signaling pathway and that blocking mouse or human HE4 with monoclonal neutralizing antibodies has a significant therapeutic effect on multiple mouse or human cancers by downregulating PD-L1 expression on macrophages and promoting CD8+ T-cell activation in the TME. Furthermore, we showed that the expression level of HE4 is high in multiple human cancer samples and that the level of HE4 is correlated with the efficiency of anti-PD-1 immunotherapy in human adenocarcinoma patients. Together, we not only identify a critical molecule and mechanism regulating the expression level of PD-L1 on myeloid cells in the TME but also identify a cancer-derived therapeutic target upstream of PD-L1 for immunotherapy, which may be more specific and has fewer irAEs.

overall_design: Bulk transcriptomic analysis was conducted to compare gene transcription in macrophages after stimulation with HE4 and IFN-γ. In brief, Raw264.7 cells were seeded in 6-well plates at a density of 5 × 105 cells/well and then stimulated with either HE4-Fc (20 μg/mL), IFN-γ (1 μg/mL), or HE4-Fc (1 μg/mL) plus IFN-γ (1 μg/mL) for 3 h. After washing twice with ice-cold PBS, total RNA was extracted and reverse-transcribed into cDNA. Library construction and sequencing were conducted via the Illumina HiSeq 4000 platform (Illumina, San Diego, CA, USA) at Majorbio Biopharm Technology Co., Ltd. (Shanghai, China). The sequencing data were analyzed via an online platform (https://cloud.majorbio.com/).

first: Chenhui

last: Wang


title: Intestinal plasmacytoid dendritic cells preferentially produce interferon lambda [scRNA-seq]

city: Portland

first: Timothy

middle: J

last: Nice

state: OR

address: 3181 SW Sam Jackson Park Rd

department: Molecular Microbiology and Immunology

institute: Oregon Health & Science University

summary: The healthy intestine mounts immune responses to microbiota to maintain homeostasis, which includes basal production of interferon cytokines.  Previous work showed that Type III Interferon (IFN-λ) stimulates localized pockets of interferon-stimulated genes (ISGs) in the adult mouse intestinal epithelium at homeostasis that provide preemptive protection from viral pathogens. Here, we demonstrate that a major source of homeostatic IFN-λ production in the intestine is a population of epithelium-associated plasmacytoid dendritic cells (pDC). These pDC are recruited to the intestine by bacterial microbiota colonization, and pDC depletion or bone marrow reconstitution with IFN-λ-deficient pDC results in reduced homeostatic ISGs in the intestinal epithelium. Notably, intestinal pDC preferentially produce IFN-λ over Type I IFNs whereas splenic pDC produce more Type I IFNs. Comparison of splenic and intestinal pDC reveal tissue-specific changes in gene expression and genomic accessibility, including evidence of response to transforming growth factor beta (TGF-β) in the intestine.  Isolated gut pDC produce more IFN-λ that splenic pDC upon stimulation, and pre-treatment of a human pDC cell line with TGF-β results in enhanced production of IFN-λ upon stimulation. This study implicates pDC as important sources of homeostatic IFN-λ in the intestine and defines the role of barrier cytokine TGF-β in regulating IFN types produced by pDC upon stimulation. Reprogramming of recruited pDC by tissue cytokines may have important implications for balancing effective antimicrobial responses with damaging inflammation at barrier tissues.

overall_design: Pooled myeloid cells from small intestinla epithelial fractions of wildtype mice were analylzed by single-cell RNA sequencing to identify cellular sources of interferon lambda gene produciton at homeostasis.

first: David

last: Constant


title: LGR5⁺ Stem Cells Maintain Apex Position in Regenerative Hierarchy of the Intestinal Epithelium During Homeostasis and Injury

city: Calgary

first: Arshad

last: Ayyaz

state: Alberta

address: BI238D, 507 Campus Dr nw

institute: University of Calgary

summary: The cellular origin of intestinal epithelial regeneration has been a subject of continued debate, with recent models challenging the primacy of WNT-dependent LGR5⁺ crypt base columnar (CBC) cells as the central regenerative population. Here, we revisit this question through quantitative integration of single-cell transcriptomic, chromatin accessibility, spatial, and lineage-tracing analyses across the proximal-to-distal axis of the small intestinal epithelium. Our data show that under homeostatic conditions, LGR5⁺ cells exclusively sustain epithelial self-renewal in nearly all crypt–villus units along the entire length of the small intestine. Following irradiation or chemotoxic injury, surviving LGR5⁺ CBC cells and their immediate progeny undergo transcriptional and epigenetic reprogramming into transient fetal-like cell states that initiate epithelial repair. Crucially, successful regeneration depends on the reactivation of canonical WNT/β-catenin signaling, as evidenced by increased TCF motif accessibility and upregulation of WNT target genes, including Lgr5. Accordingly, pharmacological inhibition of WNT signaling blocks the reconstitution of LGR5⁺ cells and crypt regeneration, leading to epithelial collapse. These findings reconcile prior controversies by demonstrating that epithelial regeneration throughout the small intestine, even following injury, is ultimately driven by LGR5⁺ CBC cells in a WNT-dependent manner.

overall_design: Mouse intestines were extracted at day 1 and 4 following 12Gy of irradiation and nuclei were isolated.

first: Julian

last: Chua

first: Lucy

last: Driver

first: Masahiro

last: Narimatsu

first: Mardi

last: Fink

first: Lixia

last: Luo

first: Jordan

last: Tran

first: Jatin

last: Roper

last: Kirsch

first: Jeff

last: Wrana

first: Chang-Lung


title: Intestinal plasmacytoid dendritic cells preferentially produce interferon lambda [ATAC-seq]

overall_design: Plasmacytoid dendritic cells were isolated from spleen and small intestinal epithelium of four mice for analysis of genomic accessability by ATACseq.


title: Intestinal plasmacytoid dendritic cells preferentially produce interferon lambda

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

overall_design: Refer to individual Series


title: scRNA-seq reveals transcriptional profiling of telocytes in synovial tissue

city: Rochester

first: Schwarz

last: Edward

state: NY

address: 601 Elmwood Ave

institute: University of Rochester Medical Center

summary: This study employs scRNA-seq to characterize Efhd1⁺ tdTomato⁺ telocyte-like cells isolated from knee synovial tissues. By comparing their transcriptional profiles to fibroblasts, we aim to delineate distinct cellular phenotypes and understand telocyte heterogeneity across tissue microenvironments. Our analysis reveals unique gene expression signatures defining telocytes providing insights into their potential roles in tissue homeostasis and fibrosis.

overall_design: Efhd1-CreER x Ai9 mice were treated with tamoxifen to induce tdTomato expression in Efhd1-expressing cells. Synovial tissues from knee joints synovial tissues were dissected and enzymatically digested to obtain single-cell suspensions. Viable tdTomato⁺ cells were isolated by fluorescence-activated cell sorting (FACS) and subjected to scRNA-seq. Sequencing data were processed with Cell Ranger for alignment and UMI counting, followed by downstream analysis using Seurat to identify cell clusters, marker gene expression, and differential gene signatures between telocytes and fibroblasts.

first: Yue

last: Peng

first: Edward

last: Schwarz


title: Human U2 snRNA BSL sequence mediates splicing efficiency driven by branchpoint sequence complementarity

city: Santa Cruz

first: Melissa

last: Jurica

address: 1156 High Street

institute: University of California Santa Cruz

summary: To investigate the global effects of perturbing U2 snRNA sequence on splicing and gene expression, we carried out transcriptomic analysis of HEK293T cell lines stably expressing U2 snRNA mutants C28U in the branchpoint interacting stem loop or T34A/ A35C/G36T to create an orthogonal branch point recognition sequence. With both mutants, the expression of many genes is generally repressed, potentially by nonsense mediated decay (NMD) arising from a global decrease in splicing efficiency. However, we also see upregulation of genes important for pre-mRNA processing, translation and protein folding. In several cases, the upregulation of certain genes can be linked to a shift in alternative splicing that favors the productive isoform relative to an NMD-targeted isoform.

overall_design: Short-read RNA-seq of HEK293T cell lines engineered for stable expression of empty vector (783) or RNU2-1 expression locus harboring either C28T (U2-C28U), TAG36ACT (U2-UCA) using lentiviral integration


title: PTMA Safeguards Mitochondrial Integrity to Sustain Metabolic Function and Antitumor Persistence of CD8+ T Cells

city: shanghai

first: keling

last: huang

address: south chongqing road

institute: shanghaijiaotong university

summary: While immune checkpoint blockade (ICB) has revolutionized cancer treatment, its clinical efficacy remains constrained by the progressive exhaustion of tumor-reactive CD8+ T cells - a dysfunctional state intimately linked to mitochondrial impairment. This underscores the urgent need to identify and target molecular regulators that govern mitochondrial metabolic fitness in CD8+ T cells, thereby preserving their effector function within the immunosuppressive tumor microenvironment (TME) to improve therapeutic outcomes. Through integrative single-cell TCR-seq and longitudinal transcriptomic profiling of reactive CD8+ T cell clones during clinical ICB treatment, we found PTMA levels in CD8+ T cells showed significant positive correlation with clinical ICB efficacy, establishing its potential as a predictive biomarker for immunotherapy outcomes. Importantly, we demostrated that Ptma expression is directly regulated by T cell factor 1 (TCF1), a transcription factor essential for T cell stemness. Moreover, genetic ablation of Ptma severely compromises the persistent effector function of CD8+ T cells and completely abrogates the in vivo efficacy of ICB. Mechanistically, we reveal that PTMA physically interacts with TFAM to safeguard mitochondrial DNA (mtDNA) integrity in CD8+ T cells under metabolic stress, thereby maintaining electron transport chain (ETC) function and oxidative phosphorylation (OXPHOS) capacity. Collectively, our findings delineate the TCF1-PTMA axis as a critical molecular nexus connecting mitochondrial metabolic competence to the stem-like properties of CD8+ T cells. This discovery provides both fundamental insights into the metabolic regulation of antitumor immunity and clinically actionable strategies to overcome ICB resistance.

overall_design: A comparative RNA-seq analysis of wild-type versus PTMA-overexpressing OT1 CD8+ T cells under 48-hour OVA peptide stimulation

first: Keling

last: Huang

first: Xiaoxue

last: LI

first: Feng

last: Wang


title: HE4 regulates anticancer immune evasion by upregulating PD-L1 expression on macrophages through the IFN-γR-STAT3 axis [LLC tumors, scRNA-seq]

first: Bo

last: Zeng

overall_design: Subcutaneous LLC tumors were harvested on the day after the last administration of the mAbs. Tissues were minced on ice and then digested at 37°C with a shaking speed of 50 rpm or 30 min. Cell suspensions were filtered through a 40-μm strainer, and the cell pellet was resuspended in red blood cell lysis solution and incubated on ice for 5 min. The cell pellet was washed and resuspended in PBS containing 0.5% FBS, followed by the evaluation of cell viability. The samples were then sent to Majorbio Biopharm Technology Co., Ltd. (Shanghai, China), for further sequencing and bioinformatic analysis. Briefly, library construction was performed according to the manufacturer’s protocol (DNBelab C TaiM4 V3.0) and quantified via a high-sensitivity DNA chip on a Bioanalyzer 2100 and a Qubit high-sensitivity DNA assay (Thermo Fisher Scientific). Libraries were sequenced on the MGI DNBSEQ-T7 platform (MGI-Tech, Shenzhen, China).

first: Chenhui


title: Bulk RNA-seq datasets supporting analysis of the MYC–USP10–SOX4 pathway in thymocytes and T-ALL

city: Changsha

first: Mengdi

last: Zhang

state: Hunan

address: 139 Middle Renmin Road, Furong District

department: National Clinical Research Center for Metabolic Diseases, Metabolic Syndrome Research Center, Key Laboratory of Diabetes Immunology, Ministry of Education, and Department of Metabolism and Endocrinology

institute: The Second Xiangya Hospital of Central South University

summary: This study presents bulk RNA-seq datasets generated from thymocytes of C57BL/6J mice to elucidate the transcriptional impact of thymus-specific loss of the deubiquitinase USP10. Total RNA was isolated from age-matched wild-type (WT) and USP10 conditional-knockout (USP10-cKO) thymus tissues, poly(A)+ fractions were enriched, and paired-end 150 bp libraries were sequenced on an Illumina NovaSeq 6000 platform. The resulting expression profiles allow direct comparison of global gene-expression programs between normal thymocyte development and USP10-deficient conditions, providing a resource for identifying USP10-regulated pathways, discerning stage-specific transcriptional alterations, and advancing our understanding of how ubiquitin signaling modulates early T-cell differentiation.

overall_design: RNA-seq profiling of thymocytes from thymus-specific USP10-knockout and wild-type C57BL/6J mice

first: Bin

last: Zhao


title: HE4 regulates anticancer immune evasion by upregulating PD-L1 expression on macrophages through the IFN-γR-STAT3 axis [RNA-Seq - cell line]

overall_design: Bulk transcriptomic analysis was conducted to compare gene transcription in macrophages after stimulation with HE4 and IFN-γ. In brief, Raw264.7 cells were seeded in 6-well plates at a density of 5 × 105 cells/well and then stimulated with either HE4-Fc (20 μg/mL) or IFN-γ (100 ng/mL) for 3 h. After washing twice with ice-cold PBS, total RNA was extracted and reverse-transcribed into cDNA. Library construction and sequencing were conducted via the Illumina HiSeq 4000 platform (Illumina, San Diego, CA, USA) at Majorbio Biopharm Technology Co., Ltd. (Shanghai, China). The sequencing data were analyzed via an online platform (https://cloud.majorbio.com/). Genes upregulated by HE4-Fc (compared with Fc stimulation) or IFN-γ (compared with the vehicle) are listed. Furthermore, KEGG pathway enrichment analysis was performed to systematically compare the signaling pathways modulated by HE4 and IFN-γ.


title: HE4 regulates anticancer immune evasion by upregulating PD-L1 expression on macrophages through the IFN-γR-STAT3 axis

overall_design: Fresh tumor and paired adjacent non-tumor tissues were collected from two lung adenocarcinoma patients immediately after surgical resection. Tissues were rinsed with ice-cold PBS three times to remove blood clots and then minced on ice. The tissue block was incubated with a digestion solution made of 0.25% trypsin and 10 µg/mL DNase I, at 37°C with a shaking speed of 50 rpm or 30 min. Cell suspensions were filtered using a 70-μm cell strainer and then through a 40-μm strainer again, and the filtrate was collected into a new centrifuge tube. The cell pellet was resuspended in 3 ml of red blood cell lysis solution and incubated on ice for 5 min. Then, 10 ml of precooled PBS was added and centrifuged at 250 × g for 5 min. The cell pellet was resuspended in PBS containing 0.5% FBS. Cells were stained with 0.4% AO/PI to evaluate cell viability. 10 × Genomics single-cell capture, Illumina library construction, preprocessing of scRNA-seq data, integration, dimensionality reduction and clustering were performed by Capital Bio-Tech (Beijing, China).


title: Transcriptomic Characterization of Liver Tissue Changes in Human and Mice with Acute-on-Chronic Liver Failure and Liver Cirrhosis/Fibrosis

city: Shanghai

first: Yun

last: Song

address: 1111 Xianxia Road, Shanghai 200336, China

institute: shanghai jiao tong university

summary: This study aims to investigate gene expression differences at the transcriptional level in liver tissue from acute-on-chronic liver failure (ACLF) compared to liver cirrhosis (CLD, human) or Fibrosis (mice) and healthy control groups.For human samples, they were categorized into ACLF, CLD, and healthy control (HC) groups. For mouse samples, they were divided into ACLF, Fibrosis, and HC groups.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Regarding human samples, liver tissue was collected. For mouse samples, fibrosis was induced by intraperitoneal injection of 10% CCl4 in olive oil at a dose of 0.2 mL/kg, twice a week for 8 weeks. Subsequently, ACLF model mice were established by inducing acute liver injury in the fibrosis mice with an intraperitoneal injection of CCl4 at 0.4 mL/kg, followed by an intraperitoneal injection of 100 μL Klebsiella pneumoniae bacterial solution 24 hours after the acute injury.

first: Panyu

last: Chen

first: Xiao

last: Lin

first: Zhaokai

first: Wenxi

last: Su

first: Yunyun

last: Ren

first: Chen

first: Kang

last: He

first: Min

last: Shi

first: Yugang


title: Caspases 3 and 7 restrict intestinal inflammation by regulating stem cell function and gsdmd-mediated pyroptosis

city: Ghent

first: Bruno

last: Verstraeten

state: Oost-Vlaanderen

address: Technologiepark 71

country: Belgium

institute: Ghent University

summary: Aberrant intestinal epithelial cell (IEC) death is a hallmark of inflammatory bowel disease (IBD), observed in patient samples and animal models of colitis. While multiple cell death pathways contribute to disease pathogenesis, the dominant modalities and their regulatory mechanisms in IBD remain poorly defined. In this study, we demonstrate that mice with intestinal epithelial cell-specific deletion of caspase-3 and -7 (Casp3/7 Tg) exhibit exacerbated colitis, increased mortality, and impaired epithelial regeneration, phenotypes not observed in other cell death-related knockout models. These findings highlight a previously unrecognized protective role of caspase-3/7 in controlling inflammation and tissue regeneration following intestinal injury. Mechanistically, caspase-3/7 deficiency negatively affects intestinal stem cell proliferation and promotes GSDMD-dependent pyroptosis, compromising barrier integrity and return to steady state conditions. Together, our results identify caspase-3/7 as key regulators of epithelial integrity and inflammation and suggest their modulation as a promising therapeutic strategy in IBD and colorectal cancer.

overall_design: Two sets of mRNA seq samples: The first set of samples are mural colon tissue samples from 4 conditions (0 or 8 days post DSS treatment of Casp3/7 transgenic and wild type mice). The second set of  samples were derived from mural intestinal epithelial cells of Casp3/7 transgenic and wild type mice. The samples were taken 0 or 3 days post DSS treatment.

first: Xie

last: Wei

first: Verstraeten

last: Bruno

first: Vandenabeele

last: Peter


title: Kidney Effects of Immune Checkpoint Inhibitors on a Humanized Chimeric Mouse Model.

first: Alexandra

last: Krivonos

state: MN

address: 200 1st St SW

department: Quantitative Health Sciences

institute: Mayo Clinic

summary: We have employed a single cell sequencing approach using 10x Genomics scRNA-seq to study immune checkpoint inhibitor effects on kidney cells. Immune checkpoint inhibitors leads to break of immune tolerance and potential adverse related event. It is known that interstitial nephritis is most common presentation. Because exact mechanism is not clear this model wants to induce inflammation in a chimeric mouse. Evaluation of genes over or under expressed in differents groups of treatment by scRNA-seq would allow to understand better the dinamic of inflammation and identified potential targets.

overall_design: Following ICI and ICI plus cytokines treatment, kidney cells from humanized PD1/PDL1 mice were isolated following 10x Genomics protocols and analyzed using scRNA-seq.

first: Sandra

last: Herrmann

first: Victor

middle: D

last: Cuenca Narvaez

first: Mohammad

middle: O

last: Al Refai

first: Coraima

last: Nava Chavez


title: Host-pathogen relationships in aoudad (Ammotragus lervia) and bighorn sheep (Ovis canadensis) with respiratory disease agents

city: Canyon

first: Matthew

middle: Adam

last: Scott

state: TX

address: 3201 Russell Long Blvd

department: Large Animal Clinical Sciences

institute: Texas A&M University

summary: To evaluate transmission propensity of Mycoplasma ovipneumoniae and leukotoxigenic Pasteurellaceae from aoudad and bighorn sheep, we assessed the transcriptomic signals from bulk lung and tracheobronchial lymph node samples of aoudad and bighorn sheep exposed to Mycoplasma ovipneumoniae in either the presence or absence of leukotoxigenc Pasteurellaceae. Our aim was to determine gene expression patterns related to respiratory pathogenic exposure within nonindigenous aoudad and native bighorn sheep to provide crucial information related to respiratory pathogenesis and identify potentially predictive patterns in individuals as it relates to interspecific contact and mortality events.

sample_organism:
Ammotragus lervia
Ovis canadensis

platform_organism:
Ammotragus lervia
Ovis canadensis

overall_design: RNA-Seq profiling of frozen lung (combined cranioventral and caudodorsal) and tracheobronchial lymph node samples collected at time of mortality.

first: Logan

last: Thomas

first: Robert

last: Valeris-Chacin

first: Walter

last: Cook


title: RNAseq on giredestrant-resistant ER+ breast cancer cell lines

first: Jackson

last: Liang

overall_design: HR+ breast cancer cells were cultured in giredestrant for 2- or 4-months until they acquired resistance to giredestrant. Cells were then profiled via bulk RNA-seq and ATAC-seq.

first: Jane

last: Guan

first: Ciara

last: Metcalfe


title: Matrix stiffness regulates telocyte differentiation into fibroblasts in vitro

first: Schwarz

last: Edward

summary: Primary telocytes from ankle joints of Efhd1CreER x Ai9 mice were purified by FACS based on tdTomato (tdT) expression. These tdT⁺ telocytes were cultured on collagen-coated CytoSoft® plates of varying stiffness (0.2, 8, and 64 kPa). After 24 hours, cells were harvested for bulk RNA sequencing using Illumina NovaSeq X Plus. The expression of telocyte markers and fibroblast/myofibroblast markers showed stiffness-dependent changes. Notably, stiff ECM induced a telocyte-to-myofibroblast transition, evidenced by downregulation of Efhd1 and Myoc and upregulation of myofibroblast markers(eg. Acta2 and Tagln). Additional analysis revealed significant changes in expression of Timp, Mmp, and Collagen genes. These findings suggest that ECM stiffness is a key regulator of telocyte fate and function at the PLV surface.

overall_design: Telocytes were isolated from ankle synovium of Efhd1CreER x Ai9 mice, and tdTomato⁺ cells were purified by flow cytometry. Purified cells were cultured for 24 hours on collagen-coated CytoSoft® 24-well plates with three levels of stiffness: 0.2 kPa (soft), 8 kPa (intermediate), and 64 kPa (stiff). Total RNA was extracted and subjected to bulk RNA sequencing. Gene expression changes were analyzed with significance set at p < 0.05.

first: Yue

last: Peng

first: Edward

last: Schwarz


title: ATACseq on giredestrant-resistant ER+ breast cancer cell lines


title: Estrogen Receptor α Protects Against Obesity-Induced Metabolic Dysfunction by Regulating VAT Tregs

first: Gregory

last: Tharp

summary: Estrogens are important for metabolic health. Individuals with low levels of circulating estrogens, including men and postmenopausal women, exhibit an elevated risk for developing obesity-associated metabolic syndromes. Chronic low-grade inflammation in the visceral adipose tissue (VAT) is a major contributor to metabolic dysfunction during obesity. Regulatory T cells (Tregs) in the VAT limit tissue inflammation and protect against obesity-associated metabolic disease. In this study, we identify opposing roles of estrogen receptor α (ERα) in regulating VAT Tregs in female mice under steady-state and obese conditions. At steady state, ERα restrained the age-dependent clonal expansion of specific VAT Treg subsets expressing the IL-33 receptor ST2. However, during obesity, ERα-deficiency predisposed females to the loss of ST2+ VAT Tregs, exacerbating VAT inflammation and insulin resistance. These findings indicate that ERα signaling protects against obesity-induced metabolic diseases by preserving metabolically protective ST2+ VAT Treg subsets, and the loss of this protection may contribute to the heightened metabolic risk in estrogen-deficient individuals.

overall_design: Spleen and/or ovarian visceral adipose tissue (oVAT) was isolated from 27-week-old Era+/+ female mice fed an NCD or Era-/- female mice fed an NCD or HFD for 17 weeks. Stromal vascular fractions from each group were stained with antibodies targeting appropriate surface markers along with corresponding TotalSeq™ hashtag antibodies for downstream demultiplexing. CD4+ CD25hi Tregs from all groups were sort-purified into one tube containing RPMI 1640 with 10% FBS for 10X genomics 5' single cell RNA/TCR sequencing analysis.

first: Cody

last: Elkins

first: Chengyu

last: Ye

first: Samara

last: Moll

first: Jianliang

last: Zhang

first: M. Neale

last: Weitzmann

first: Chaoran

last: Li


title: Epithelial-mesenchymal transition is associated with altered immune composition and cytotoxic function in triple-negative breast cancer

city: Lebanon

first: Brock

last: Christensen

state: NH

address: 1 Medical Center Dr

institute: Dartmouth College

summary: Epithelial–mesenchymal transition (EMT) is a dynamic process that contributes to breast cancer progression, metastasis, and therapy resistance. EMT also influences the tumor immune microenvironment, shaping immune cell infiltration and function. Although the immunological features of fully epithelial or fully mesenchymal tumor states have been characterized, the immune landscape associated with intermediate or partial EMT states remains poorly understood. To address this gap, we established five single-cell–derived clonal populations from the triple-negative 4T1 mouse mammary tumor cell line, representing a spectrum of EMT phenotypes, and analyzed tumors derived from these clones using single-cell RNA sequencing. Here, we show that tumors derived from these clones retained their relative EMT states in vivo and that EMT progression is associated with a graded reduction in tumor immunogenicity, accompanied by alterations in immune cell composition and function. Specifically, along the EMT spectrum, tumor cells exhibited progressive downregulation of MHC class I and II gene expression, along with decreased infiltration of cytotoxic CD8+ T cells and reduced expression of key effector and trafficking genes (Gzmb, Ccr5, Cxcr6). B cell composition also shifted, with decreased frequencies of IgG1-producing plasma cells and an enrichment of regulatory-like B cells. Natural killer (NK) cells similarly demonstrated progressive functional suppression, marked by reduced expression of cytotoxic molecules and downregulation of key effector pathways.   Together, these findings reveal that EMT is associated with immune cell recruitment and function in a graded manner, providing a rationale for integrating EMT phenotyping into therapeutic strategies to overcome immune resistance in breast cancer.

overall_design: Five single-cell-derived clonal populations representing epithelial, intermediate, and mesenchymal states were generated from the murine 4T1 triple-negative breast cancer cell line and implanted into BALB/c mice. Tumors were harvested, and both total tumor cells and CD45⁺ immune cells were isolated using magnetic sorting. One tumor per group was used for single-cell RNA sequencing using the 10x Genomics platform.

first: Hanxu

last: Lu

first: Meisam

last: Bagheri

first: Fred

last: Kolling

first: Lucas

last: Salas

first: Xiaofeng

last: Wang

first: Todd

last: Miller

first: Diwakar

last: Pattabiraman


title: Single cell multiomic characterizations of cART, SIV and morphine’s effect on brain myeloid cells

city: Omaha

first: Howard

last: Fox

state: Nebraska

address: 42nd and Emile

institute: University of Nebraska Medical Center

summary: The primary immune cells in the brain, microglia and CNS-associated macrophages (CAMs), are key targets for HIV in humans and SIV in nonhuman primates. Infection with these viruses can lead to neurological dysfunction collectively referred to as HAND. While antiretroviral therapy (ART) has proven effective in managing HAND, particularly the severe form known as HIV-associated dementia (HAD), its efficacy in preventing milder forms remains limited, leaving HAND a prevalent complication among PWH. Substance abuse, notably with opioids such as morphine and its synthetic product heroin, as well as pharmaceutical opioids, is relatively common in people with HIV (PWH) and the progression of HIV and its associated complications is exacerbated in opioid-using PWH. This study investigates the individual and combined effects of ART-treated SIV infection and morphine use on brain myeloid cells in rhesus macaques. Using single nucleus multiomic sequencing (integrating gene expression and chromatin accessibility), we examined the phenotypic, transcriptomic, and epigenomic changes in microglia and CAMs. Our findings confirm that ART significantly restores homeostasis in brain myeloid cells during SIV infection in macaques, paralleling its effects in HIV-infected humans. However, integration of data from ART-suppressed PWH with our macaque data suggested that there were more activation/inflammatory phenotypes of brain myeloid cells in PWH compared to ART-suppressed SIV-infected macaques. Additionally, we found that morphine exhibits immunosuppressive effects on brain myeloid cells, characterized by reduced antiviral phenotypes, downregulation of immune activation genes, including MHC class II and interferon-inducible proteins, and lack of the enrichment of the transcription factors in the AP-1 and ETS families activity. Gene co-expression network analysis further confirmed that the brain myeloid cell activation is associated with ART-treated infection, with no or negative correlation with morphine use. However, morphine exposures might indirectly affect the brain myeloid cells’ activation and functions, probably through endocrine receptors. These findings provide a deeper understanding of the effects of SIV infection on brain myeloid cells within the context of ART-treated SIV infection and opioid abuse, a critical consideration for PWH. Insights gained from this study could inform clinical approaches and therapeutic development for this vulnerable population.

overall_design: The 16 macaques were randomly assigned to four groups (n = 4 per group): Saline, SalineSIV, Morphine, and MorphineSIV. Animals in the Morphine and MorphineSIV groups received intramuscular morphine injections, Macaques in the SalineSIV and MorphineSIV groups were infected with SIVmac251. Five weeks post-infection, all SIV-infected animals began receiving a daily subcutaneous injection of antiretroviral therapy (ART) at 1 mL/kg, containing 40 mg/mL emtricitabine (FTC), 20 mg/mL tenofovir (TFV), and 2.5 mg/mL dolutegravir (DTG), which continued until necropsy.

first: Benjamin

last: Lamberty

first: Katy

last: Emanuel

first: Meng

last: Niu

first: Xiaoke

last: Xu


title: Identification of Functional Genetic Components modulating Cellular Toxicity Response to PFOS using Genome-wide CRISPR Screens

city: Gainesville

first: Chanhee

last: Kim

state: Florida

address: 2187 Mowry Road

department: Physiological sciences

institute: University of Florida

summary: Per- and polyfluoroalkyl substances (PFAS) are a group of chemicals that pose significant health and environmental risks due to their persistence and widespread use. These substances, known for their strong chemical bonds, are linked to various health issues such as liver damage, developmental disorders, and immune dysfunction. However, the molecular mechanisms behind their toxicity are not fully understood. Although several studies have identified molecular responses to PFAS exposure, there remains considerable uncertainty about the underlying mechanisms leading to PFAS-associated health effects. To help bridge this gap, we conducted genome-wide CRISPR-based screens in HepG2/C3A human liver cells to identify genes and pathways influencing cellular toxicity upon PFAS exposure. We focused on perfluorooctane sulfonate (PFOS), a widely used and studied PFAS compound with known toxic effects, yet unclear molecular mechanisms underlying PFOS-induced adverse outcomes. Using a genome-wide CRISPR knockout library targeting 18819 genes, we identified 340 candidate genes which modulate the cellular toxicity of PFOS when disrupted. Pathway and enrichment analyses of the corresponding gene products suggest a role for cell cycle, DNA damage response, and Wnt/-catenin pathways in mediating PFOS cytotoxicity, while further gene-phenotype association analysis using the comparative toxicogenomics database indicated enrichment of gene products previously associated with liver disease and neoplasia. We carried out individual targeted disruption of two candidate genes, SLC6A9 and CPSF2, in HepG2/C3A to confirm their roles in resistance to PFOS exposure. Additionally, molecular docking analysis predicts that PFOS may bind directly to GlyT1, the SLC6A9 gene product. Lastly, cross-species relevance analysis using gene targets identified in our screens suggested that molecular pathways of PFOS toxicity are highly conversed in several environmentally relevant species.

overall_design: Genome-wide CRISPR screen experiments using HepG2/C3A cells followed by sgRNA abundance profiling in control and PFOS exposure (192 uM) samples after 10 doubling times (= at Day 18)

first: Chris

last: Vulpe


title: Regulation of Epstein–Barr Virus latency by histone variant H2A.Z [ChIP-seq]

city: Philadelphia

first: Priyankara

last: Wickramasinghe

state: PA

address: 3601 Spruce Street

department: Bioinformatics

institute: The Wistar Institute

summary: Chromatin structure plays a central role in the regulation of Epstein-Barr Virus (EBV) latency. The histone variant H2A.Z.1 has been implicated in chromatin structures associated with initiation of transcription and DNA replication. Here, we examined the role of H2AZ.1 in the regulation of EBV chromatin and gene expression during EBV latent infection in two different EBV cancer cell models. We found that H2A.Z.1 is highly enriched with EBNA1 binding sites at oriP and Qp, and to a lesser extent with transcriptionally active CTCF binding sites on the EBV genomes in both Mutu I Burkitt lymphoma (BL) and SNU7-19 EBV-associated gastric carcinoma (EBVaGC) cell lines. RNA-interference depletion of H2A.Z.1 resulted in the reactivation of viral lytic genes (ZTA and EAD) and increases viral DNA copy numbers in both MutuI and SNU719 cells.  H2A.Z depletion also led to a decrease in EBNA1 binding to oriP and Qp, on the viral episome as well as on plasmids independently of other viral genes and genomes. H2A.Z.1 depletion also reduced peaks of H3K27ac and H4K20me3 at regulatory elements in the EBV genome.  In the cellular genome, H2A.Z.1 colocalized with only a subset of EBNA1 binding sites and H2A.Z.1 depletion altered transcription of genes associated with myc targets and mTORC1 signaling.  Taken together, these findings indicate that H2A.Z.1 plays an important regulatory role in the regulation of EBNA1 chromatin binding and function in the EBV episome and host chromosome, as well as the epigenetic programming of the latent episome.

overall_design: SNU-719 cells were treament with siControl and siH2A.Z, after 9 days the celles were recollected and processed to western blot, RNA-seq and ChIP-seq.

first: Leonardo

last: Castro-Muñoz

first: Davide

last: Maestri

first: Leena

last: Yoon

first: Bhanu

last: Karisetty

first: Italo

last: Tempera

first: Paul

last: Lieberman


title: CTCF regulates chromatin accessibility of the HBAD gene promoter during erythroid differentiation [ChIP-Seq]

city: Mexico City

first: Gustavo

last: Tapia

state: CDMX

address: Cto. Exterior s/n

department: Molecular Genetics

country: Mexico

institute: IFC-UNAM

summary: Chromatin structure undergoes dynamic rearrangements during cell differentiation. The architectural protein CTCF plays diverse roles in this process. Here, we demonstrate that a subset of gene promoters alters their chromatin accessibility profiles upon CTCF binding during differentiation of the chicken erythroblast cell line HD3. Furthermore, we show that CTCF contributes to establishing a specific 3D chromatin architecture in the α-globin multigenic domain prior to differentiation in HD3 cells.

overall_design: To characterize CTCF's role during in vitro chicken erythroid differentiation, we performed CTCF ChIP-seq in both uninduced HD3 cells and HD3 cells induced for 24 hours. We then generated a promoter-specific mutant lacking the CTCF binding site in the HBAD globin gene and analyzed resulting changes using ATAC-seq, to assess chromatin accessibility, and UMI-4C, to evaluate local chromatin interactions.

last: Tapia Urzúa

first: Hober

middle: N

last: Núñez Martínez

first: Martin

last: Franke

first: Sylvia

last: Garza Manero

first: Ángel

last: Cerecedo Castillo

first: Carlos

last: Peralta Álvarez

first: Georgina

last: Guerrero

first: José

last: Gómez Skarmeta

first: Félix

last: Recillas


title: Transcriptional profile in  SK-LM_S1 cells mediated by abrogation of  MAPK7 and VCAN gene expression [Part 1]

city: Madrid

first: Yosra

last: Berrouayel

address: Arturo Duperier, 4

country: Spain

institute: Universidad Autónoma de Madrid

summary: The ERK5 signaling pathway has emerged as a novel key player in soft tissue sarcoma biology, mediating tumour growth and progression. Here we show how the expression of VCAN, a member of the chondroitin sulphate proteoglycan family, is modulated by the ERK5 pathway at the transcriptional level and mediates biological processes such as migration, adhesion, proliferation and tumourigenesis in vivo.  This conclusion is based on genetic (shRNA, overexpression) and pharmacological approaches using  the human soft tissue sarcoma  cell lline Sk-LM-S1 a. In particular, VCAN knockdown mimics ERK5 knockdown in vitro and in vivo, whereas VCAN overexpression restores the phenotypes observed after ERK5 knockdown. In addition, transcriptomic studies show that VCAN is a central mediator in the ERK5-associated transcriptional landscape, supporting its role in diverse biological processes. Taken together, these data support VCAN as a new key target in the ERK5 pathway.

overall_design: Following effective knockdown of VCAN and MAPK7 gene expression in SK-LM-S1 cells using specific shRNA, samples were collected and transcriptomic profile analyzed by RNA-seq.

last: Berrouayel Dahour

first: Francisco J

last: Cimas

first: Luis

last: del Peso

first: Ricardo

last: Sanchez-Prieto


title: scRNA profiling of early postnatal thymus derived Vδ1+ gamma delta T cells.

city: Chicago

first: Erin

last: Adams

state: IL

institute: University of Chicago

summary: We focus on characterizing Vδ1 T cell development through determination of their effector profile, characterization of their paired TCR repertoire, and understanding of antigen reactivity towards CD1 molecules through single cell RNA analyses.

overall_design: To understand Vδ1 T cell developmental trajectories, gammadelta T cells from thymus samples were sorted on Vδ1, stained with immunophenotyping CITE-seq antibodies and barcoded tetramers for antigen specificity, and GEX, VDJ, and antibody libraries were sequenced using the 10X Genomics 5’ Single Cell Immune Profiling workflow

first: Mayuri

last: Viswanathan

first: Caitlin

last: Castro

first: Augusta

last: Broughton


title: Distinct roles of tip progenitor subpopulation in shaping the murine lung

city: San Francisco

first: Pao-Tien

last: Chuang

address: 555 Mission Bay Blvd. South

institute: University of California, San Francisco

summary: A gradient SOX9+ distal progenitor cell differentiation impacts the patterning of the lung during branching morphogenesis. Although the heterogeneity of this population has been suggested, the specific roles of these subgroups still largely obscure. Here, we explore at least two distinct functional subgroups: the tip progenitors residing at the tips are critical for bud extension and cell allocation, while the transition progenitors sitting immediately to them are indispensable for cell transition. It’s suggested Hippo pathway could regulate the gradient cell differentiation during lung branching. We observed the Cre recombinase driving by Sftpc promoter couldn’t delete Lats1/2 floxed alleles in majority of the tip of SOX9+ distal region, even though the Cre was activated in all the conducting airway. Genetic evidence implied the distal tip progenitors alone is sufficient to help construct the comparable lung structures, while cells at transition stage contributed to the conducting airways. At the expense of themselves, the distal tip progenitors would quickly switch to the transition state when activate YAP via LATS1/2 perturbation. The increasing of cells at transition state results in aggregation of AT1 cells and missing of AT2 cells. Lacking Lats1/2 in SOX9+ distal progenitors in 3D culture could also promote the cell differentiation toward the transition state. Thus, a lower YAP activity is benefit for tip progenitor cell reservation.

overall_design: Examine the difference of gene expression between Lats1/2 -deficient and their littermate control lungs

first: Kuan

last: Zhang

first: Madhuri

last: Basak

first: Erica

last: Yao


title: Stratifying IVF population endometria using a prognosis gradient independent of endometrial timing

city: Valencia

first: Patricia

last: Diaz-Gimeno

state: Valencia

address: EDIFICIO BIOPOLO - LA FE, Avenida Fernando Abril Martorell, 106 - Torre A, Planta 1ª

department: Genomics and Systems Reproductive Medicine

institute: IIS La Fe / IVIRMA Global Research Alliance

summary: This study aimed to stratify and characterize the disrupted WOI patterns using endometrial timing-corrected whole gene expression and artificial intelligence (AI) models in in vitro fertilization (IVF) patients undergoing hormone replacement therapy (HRT).

overall_design: A multicenter prospective study involving 195 IVF patients undergoing hormone replacement therapy (HRT) included the collection of endometrial biopsies during the mid-secretory phase.

last: Sebastian-Leon

first: Antonio

last: Parraga-Leo

first: Josefa Maria

last: Sanchez-Reyes


title: Antibiotic sensitization of pathogenic Pseudomonas aeruginosa via negative hysteresis in the cell envelope

city: Großhansdorf

first: Espen

last: Groth

address: Wöhrendamm 80

department: Department of Pulmonology

institute: LungenClinic Grosshansdorf

summary: Antimicrobial resistance (AMR) is a major challenge for global health. Multi-drug treatments can help to reduce AMR and be implemented through the sequential administration of antimicrobials (Roemhild and Schulenburg 2019). Such sequential treatments can achieve high efficacy through negative cellular hysteresis: the antibiotic-induced sensitization of bacterial cells towards a subsequently administered antibiotic (Roemhild et al. 2018). However, to date, the exact characteristics of antibiotic sensitization via negative hysteresis are unknown. Therefore, our study aimed at providing fundamental insights into the nature of hysteresis and its possible applicability to treatment, focusing on the high-risk human pathogen Pseudomonas aeruginosa. We found that negative hysteresis is robustly expressed across the species P. aeruginosa, particularly upon switches from β-lactam antibiotics to aminoglycosides, even if the β-lactam is administered at sub-MIC levels. We further characterized the molecular basis of hysteresis by a transcriptomic analysis of wildtype Pseudomonas PA-14 and a PA-14 CpxS T163P mutant that is described here. We demonstrate that the Cpx envelope stress response system is centrally involved in inducible cellular sensitization. Furthermore, negative hysteresis and the Cpx system are linked in several cases to the expression of synergistic drug interactions. Overall, our study reveals negative hysteresis to be associated with β-lactam-induced membrane stress, widely expressed across P. aeruginosa, thus identifying this phenomenon as a promising focus for effective antimicrobial therapy.

overall_design: Pseudomonas aeruginosa PA14 and a PA14 CpxS T163P mutant were grown to mid-exponential phase (OD600 = 0.065) at 37 °C, and then either treated with the beta-lactam antibiotic carbenicillin at sub-MIC, Minimum Inhibitory Concentration, (50 mg/L) for hysteresis induction or not treated. After 30 min, cells were resuspended in antibiotic-free medium and harvested at three sequential time points (0, 30 and 60 min) to investigate the expression dynamics induced by short-term carbenicillin/Beta-lactam treatment that could explain induced sensitivity towards other antibiotics/negative hysteresis.

first: Espen E.

first: Roderich

last: Roemhild

first: Sören

last: Franzenburg

first: Hinrich

last: Schulenburg


title: Scap Stabilizes PKM2 to Promote Glycolysis and Enhance Antifungal Immunity in Macrophages

city: Hangzhou

first: Jiaqi

last: Huang

state: Zhejiang

address: No. 866 Yuhangtang Road

institute: Zhejiang University

summary: Fungal infection induces substantial but poorly understood metabolic reprogramming in macrophages. To identify molecules that are potentially involved in the fungi infection of macrophages, RNA-Seq was conducted to compare the expression of mRNAs in Candida albicans infected BMDMs and control BMDMs from Scapf/f and Lyz2-cre-Scapf/f mice.

overall_design: RNA-seq was performed on bone marrow–derived macrophages (BMDMs) stimulated with Candida albicans for 6 hours, with untreated cells used as controls  from Scapf/f and Lyz2-cre-Scapf/f mice. Each group consisted of three samples, each containing 5 million cells. RNA extractions were sent to BGI (Shenzhen) for RNA-seq analysis.

first: Zhihua

last: Chen


title: Parathyroid hormone 1 receptor signaling links to craniosynostosis by regulating calvaria cartilage and the suture formation

city: Suita

first: Daisuke

last: Okuzaki

state: Osaka

address: Yamadaoka 3-1

department: Immunology Frontier Research Center

country: Japan

institute: Osaka univ.

summary: Our study uncovered a unique factor causing craniosynostosis studying calvaria chondrocyte differentiation and the regulation by Pth1r/Ihh signaling.

overall_design: We performed a comprehensive analysis using RNA-Seq. Calvaria tissue, including interparietal/occipital bone and excluding ear cartilages, was removed from both control and mutant newborns.

first: Katsuhiko

last: Amano

first: Mikihiko

last: Kogo


title: Mapping the Secondary Response to Traumatic Brain Injury Using Spatial Transcriptomics Shows Acute 4-aminopyridine Treatment Mitigates Axonal and Molecular Pathology

status: Public on Feb 12 2026

city: Bethesda

first: Regina

last: Armstrong

state: MD

address: 4301 Jones Bridge Rd

department: Anatomy, Physiology and Genetics

institute: Uniformed Services University of the Health Sciences

summary: Damage to long myelinated axons of white matter tracts is a hallmark pathology resulting from traumatic brain injury (TBI) forces and secondary injury processes. 4-aminopyridine (4-AP) is an FDA-approved Kv1 potassium channel inhibitor designed to mitigate axon dysfunction.  We examined repurposing 4-AP as an acute TBI treatment using clinically-oriented neuropathology of axon damage combined with unbiased genome-wide spatial transcriptomics for comprehensive analysis of secondary injury processes.  Adult male and female mice received a non-penetrating impact TBI with 4-AP (i.p., b.i.d) on days 1-7 post-injury. Along corpus callosum (CC) axons, TBI disrupted node of Ranvier domains, exposing the putative 4-AP target of mislocalized Kv1 channels (p < 0.005). Clinically reasonable 4-AP dosing (0.5 mg/kg) reduced nodal Nav1.6 channel loss (p < 0.05) and Caspr heminode formation (p < 0.005) after injury. Quantification of β-amyloid precursor protein immunolabeling showed significantly reduced CC axon damage at 4-AP doses of 0.5 mg/kg and 5 mg/kg (each p < 0.005). 4-AP safety, based on potential seizure risk after TBI, was unaltered with vehicle or 0.5 mg/kg 4-AP, while the 5 mg/kg dose induced seizure behavior in sham and TBI groups (p < 0.0001). Spatial transcriptomics mapped molecular signatures to tissue pathology. TBI increased axonal injury response genes in the CC and in motor and somatosensory cortex sites of CC projection neurons. TBI induced disease-associated glial phenotypes that mapped predominantly within the CC. TBI increased pathway expression for immune and vascular functions, neuron and glial cell signaling, and cellular dyshomeostasis, while reducing expression in myelination-related pathways. Gene expression analysis of 4-AP treatment (0.5 mg/kg) indicated potassium channel target engagement and increased neuroaxonal activity, along with dampened secondary injury responses. Collectively, these findings reveal underlying molecular pathology of the secondary injury response and advance 4-AP translation to reduce axon damage and stimulate activity-dependent repair after acute TBI.

overall_design: At 8 weeks of age, 24 male C57BL/6J mice received TBI (non-penetrating closed-skull impact, centered at bregma) or sham procedures, followed by 4-AP (0.5 mg/kg) or saline vehicle injections (i.p., b.i.d.) until analyses at 7 days post-injury. To identify comprehensive molecular changes resulting from acute 4-AP treatment in relation to TBI-induced pathology, genome-wide spatial transcriptomic (Visium CytAssist) and complementary adjacent immunohistochemical analyses of 7 µm thick FFPE coronal brain tissue sections were conducted under the impact site in cortical gray and corpus callosum white matter regions of interest.

first: Genevieve

last: Sullivan

first: Kryslaine

last: Radomski

first: Shaoqiu

last: He

first: Matthew

last: Wilkerson

first: Clifton

last: Dalgard

first: Camille

last: Alba

first: Xiaomei

last: Zi

first: Martin

last: Doughty


title: Translation in Proximity to Forming Autophagosomes During Sustained Autophagy

city: Ottawa

first: Theodore

last: Perkins

state: ON

address: 501 Smyth Rd.

department: Cellular and Molecular Medicine

institute: Ottawa Hospital Research Institute

summary: Autophagy is an evolutionarily conserved catabolic process. In a process requiring a cascade of over 35 autophagy-related genes (Atg), a cupped phagophore membrane expands to surround cytoplasmic material, and seals itself to form an autophagosome, which finally fuses with lysosomes. Large numbers of autophagosomes form during stress responses, while simultaneously cells drastically reduce translation to conserve energy. Here, using proximity-labeling and Fluorescence in situ Hybridization we demonstrate that multiple mRNAs encoding proteins required for autophagy preferentially localize in proximity to forming autophagosomes. Polysome fractionation and proteomics of nascent proteins in proximity to forming autophagosomes provides evidence for the local translation of these mRNAs. Translation and the ribosome-binding protein RACK1 were required for the localization of these mRNAs to forming autophagosomes. Inhibition of translation or knockdown of RACK1 caused depletion of several proteins required for autophagy and a reduction in the number of autophagosomes. Local translation may enable a rapid, energy-efficient supply of proteins for autophagy to enable cells to massively induce autophagy while conserving energy during cell stress.

overall_design: Human cells expressing APEX2 fused to defined subcellular markers were used to identify RNAs proximal to forming autophagosomes. Samples include APEX2-DFCP1 (phagophore), APEX2-LC3B (autophagosome), and control APEX2 constructs (diffuse cytosolic V5 tag or Perilipin). Autophagy was induced by mTOR inhibition (Torin1). Untreated cells serve as non-autophagy controls. Proximity labeling was activated with biotin-phenol and H₂O₂, with -H₂O₂ samples as labeling controls. Biotinylated RNAs were affinity-purified and sequenced. Primary comparisons are DFCP1-APEX2 or LC3B-APEX2 (+H₂O₂) versus matched  -H₂O₂ controls to define proximal RNAs, and DFCP1-APEX2 versus cytosolic or Perilipin APEX2 to identify phagophore-specific enrichment.

first: Yunping

last: Xue

first: Kaela

last: O'Connor

first: Karsten

last: Nalback

first: Alexandre

last: Savard

first: Karen E.

last: King

first: Ryan C.

last: Russel

first: Christain

last: Bahrends

first: Derrick

last: Gibbings


title: Nuclear receptor ESRRA promotes ERa-positive breast cancer through dual action on super enhancers and promoters to regulate gene transcriptional programs

city: Xiamen

first: Guosheng

last: Hu

state: Fujian

address: Xiang'an South Road, Xiang'an District

department: School of Pharmaceutical Sciences

institute: xiamen University

summary: ESRRA (Estrogen-Related Receptor Alpha) is an orphan nuclear receptor that plays a pivotal role in regulating cellular metabolism, mitochondrial biogenesis, and energy homeostasis through gene transcriptional control. It has been implicated in various physiological processes and diseases, including cancer. However, the transcriptional programs it regulates and the underlying molecular mechanisms are yet to be comprehensively understood. Here, we demonstrated that ESRRA serves as a central regulator in orchestrating the transcriptional activation of ERa-occupied super enhancers (ERSEs) and cognate ERa-target genes. Meanwhile, it downregulates the expression of cytosolic RNA sensors RIG1 and MDA5 to inhibit type I interferon (IFN) and type I IFN-stimulated genes (ISGs). In terms of mechanism, ESRRA is recruited to ERSEs together with ERain response to estrogen, and their binding with ESREs are regulated in a mutual-dependent manner. However, ESRRA directly binds to the promoter regions of RIG1 and MDA5 to suppress their expression. Consistent with these findings, pharmacological inhibition of ESRRA with its inverse agonist XCT790 inhibits estrogen/ERa-induced gene transcription, and enhances the type I IFN pathway and antitumor immunity to suppress breast cancer cell growth both in vitro and in vivo. Co-treatment with XCT790 and endocrine therapy drugs including ERa antagonist Tamoxifen or degrader Fulvestrant exhibited synergistic effects. Furthermore, combination treatment with XCT790 and tamoxifen re-sensitized Tamoxifen-resistant (TamR) ERa-positive breast cancer cells to Tamoxifen treatment. In summary, our findings unveiled that ESRRA is a pivotal regulator of ERSEs and estrogen/ERa-target gene activation, as well as the type I IFN signaling pathway, offering a promising therapeutic avenue for treating ERa-positive breast cancer in the clinic.

overall_design: MCF7 cells were transfected with control siRNA (siCTL) or siRNA specifically targeting ESRRA (siESRRA) in stripping medium for three days, and then treated with either 10-7 M 17-β-estrodial (E2) or ethanol for 6 h followed by RNA-seq analysis.

first: Zao-zao

last: Zheng

first: Lu

last: Chen

first: Guo-sheng

first: Qi-xuan

last: Huang

first: An

last: Wang

first: Chuan

first: Wen

last: Liu


overall_design: MCF7 cells were maintained in stripping medium for three days,  and then treatment of either 10-7 M 17-β-estrodial (E2) or ethanol for 1 h followed by ChIP- seq.


title: Trained immunity of alveolar macrophages promotes injury resolution by enhancing KLF4-MERTK-mediated efferocytosis [2]

first: Jalees

last: Rehman

address: 900 S. Ashland Ave. MBRB 2072, Mailcode 669

department: Biochemistry and Molecular Genetics

institute: UIC

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

overall_design: Refer to individual Series


title: Does three copies of App contribute to amplified basal interferon signaling in the brain of the Dp1Tyb mouse model of Down syndrome

city: London

first: Frances

last: Wiseman

address: Queen Square House

country: United Kingdom

institute: University College London

summary: We have previously found that the Dp1Tyb mouse model of Down syndrome exhibits amplified basal interferon signaling in the brain but that the Dp2Tyb mouse does not unless challenged with interferon. The Dp1Tyb model carries an additional copy of the App gene, whereas the Dp2Tyb model does not. The additional copy of App has been suggested to contribute to altered neuroimmune biology in the brains of a genetically similar DS mouse model. Moreover, raised amyloid-β triggers interferon signaling in the brain of amyloid accumulation mouse models. Here we test if an additional copy of App contributes to altered interferon signaling in the Dp1Tyb mouse model of Down syndrome in the hippocampus.

overall_design: To determine if three-copies of App contribute to amplified basal interferon signaling in the brain of the Dp1Tyb mouse model, we crossed the Dp1Tyb model with heterozygous App knock-out mice, to generate Dp1Tyb mice with three or two-copies of the App gene. We aged these mice to 6-months of age alongside WT controls. Animals were euthanized by perfusion with PBS and hippocampus, cortex and rest of brain isolated. For this study the hippocampus was used for bulk RNA-Seq.

first: Cliona

last: Farrell


title: NAD+ controls circadian rhythmicity during cardiac aging

city: Bad Nauheim

first: Pieterjan

last: Dierickx

state: Hessen

address: Parkstrasse 1

department: Circadian Regulation of Cardiometabolism

institute: Max Planck Institute for Heart and Lung Research

summary: Disruption of the circadian clock as well as reduced NAD+ levels are both hallmarks of aging. While circadian rhythms and NAD+ metabolism have been linked in heart disease, their relationship during cardiac aging is less clear. Here, we show that aging leads to disruption of diurnal gene expression in the heart. Long-term supplementation with the NAD+ precursor nicotinamide riboside (NR) boosts NAD+ levels, reprograms the diurnal transcriptome, and reverses naturally occurring cardiac enlargement in aged female mice. In addition, drastic reduction of NAD+ levels in CMs impairs PER2::luc oscillations, which is rescued by NR supplementation. Finally, we demonstrate that changes to the cardiac transcriptome due to NR treatment partially depend on the activity of SIRT1. These findings reveal an essential role for NAD+ in regulation of the cardiac circadian clock upon aging, which opens up new avenues to counteract age-related cardiac disorders.

overall_design:
mRNA sequencing was performed on the hearts of 12-month-old female mice collected at ZT10 or ZT22, either under normal housing conditions (MQ) or with nicotinamide riboside supplemented in the drinking water (NR) from the age of 2 months. For comparison, hearts collected at ZT10 or ZT22 from 2-month-old female mice with no treatment were also sequenced.
HL-1 cells (commercial cell-line dervied from atrial tumours of a female C57BL6/J mouse) were cultured under normal (Ctrl) conditions, in the presence of the NAMPT (NAD+ synthesis) inhibitor FK866 to induce a low NAD state, with FK866 and nicotinamide riboside (NAD precursor) to rescue NAD levels, or with FK866+nicotinamide riboside+Ex527 (sirtuin inhibitor to block deacetylation activity during high NAD) to investigate reliance on sirtuins for the rescue experiment.

first: Bryce

last: Carpenter

first: Margaux

last: Lecacheur

first: Yannick

last: Mangold

first: Kai

last: Cui

first: Stefan

last: Günther

first: Marit

middle: W

last: Vermunt


title: Single cell RNA-seq of intrahepatic biliary epithelial cells following bile duct ligation.

city: Tuscaloosa

first: Sergei

last: Bombin

address: 739 University Blvd

institute: The University of Alabama

summary: To identify biomarkers of subpopulations of intrahepatic biliary epithelial cells, we performed single cell RNA-seq on adult biliary epithelial cells (BECs) and hybrid hepatocytes (HybHeps) from uninjured or cholestatic injured Sox9EGFP+ mice. After 7 days, the livers were dissociated to single cells and stained for CD31, CD45, Annexin V, and 7ADD. BECs were isolated by first excluding positively stained cells and then sorting EGFP+ cells. Gallbladder and extrahepatic tissues were excluded by dissection. Sorted cells were fixed according to manufacturer instructions using Parse Whole Cell Fixation kit. Fixed cells were frozen and stored at -70C for less than 6mo, prior to being used in split-pool combinatorial barcoding according to manufacturer instructions. Barcoding and library construction was carried out with the Parse Evercode Whole Transcriptome v2 kit.

overall_design: Clustering and differential expression analysis identified 9 BEC and 6 non-BEC clusters.

first: Kendall

last: Kanakanui

first: Adam

last: Gracz


title: Diversity of Cortical Progenitors Directs Neuronal Layer Formation and Regional Glial Patterning

first: Ana Cristina

last: Ojalvo Sanz

state: Madrid

address: Avenida Doctor Arce 37, Laboratorio MB02

institute: Cajal Neuroscience Centre

summary: How distinct cortical progenitor types and their temporal differences influence neuronal and glial development remains a key question in neurobiology. While Radial Glial Cells (RGCs) are well-established as Neural Progenitors (NPCs), NG2+NPCs have emerged as contributors with unique characteristics. Here, we combined in utero electroporation, lineage tracing and transcriptomic profiling to compare lineage outputs of NG2+ and GFAP+NPCs in the developing mouse cortex. Both generate neurons and glia, but with partially distinct patterns in lineage output, spatial distribution, and fate bias. NG2+NPCs undergo an early neurogenic phase followed by glial differentiation, producing glia with laminar distribution biases. In contrast, GFAP+NPCs sustain neurogenesis longer, producing a broader range of neurons and more constant glia during development. Our findings highlight the contribution of progenitor identity and temporal differences to distinct, partially overlapping lineage trajectories, supporting a model in which fate potential and timing co-regulate the spatial and laminar organization of cortical cell types.

overall_design: Bulk RNA-seq of NG2+ and GFAP+FACS-isolated cells at E14, after targeting at E12 by IUE using the StarTrack method.

last: Ojalvo-Sanz

first: María

last: Figueres-Oñate

first: Sonsoles

last: Barriola

first: Carolina

last: Pernia-Solanilla

first: Rebeca

last: Sanchez-Gonzalez

first: Lina

last: Delgado García

first: Laura

last: López-Mascaraque


title: The Age-Dependent Resident Myonuclear Multi-Omic Response to an Acute Skeletal Muscle Hypertrophic Stimulus in Mice

city: Fayetteville

first: Pieter

middle: Jan

last: Koopmans

state: AR

address: 155 N Stadium Drive

department: Cell and Molecular Biology

institute: University of Arkansas - Fayetteville

type: Methylation profiling by high throughput sequencing

summary: A detailed analysis of how muscle fiber nuclei (myonuclei) respond to a hypertrophic stimulus would provide a critical step toward understanding compromised skeletal muscle plasticity with age. We used recombination-independent doxycycline-inducible myonucleus-specific fluorescent labelling, tissue RNA-sequencing, myonuclear DNA methylation analysis, multi-omic integration, and single myonucleus RNA-sequencing to define the molecular characteristics of adult (6-8 month) and aged (24 month) murine skeletal muscle after acute mechanical overload (MOV). In adult and aged MOV muscles, we found that: 1) similarities in the transcriptional response to loading – specifically in metabolism genes – were partly explained by a post-transcriptional microRNA-mediated mechanism, which we corroborated using an inducible muscle fiber-specific miR-1 knockout model, 2) differences in age-dependent transcriptional responses were linked to the magnitude and location of differential DNA methylation in resident myonuclei, specifically around hypertrophy-associated genes such as Myc, Runx1, Mybph, Ankrd1, collagen genes, and minichromosome maintenance genes, 3) adult and aged resident myonuclear transcriptomes had differing enrichment for innervation-related transcripts as well as unique transcriptional profiles in an Atf3+ “sarcomere assembly” population after MOV, and 4) cellular deconvolution analysis supports a role for neuromuscular junction regulation in age-specific hypertrophic adaptation. These data are a roadmap for uncovering molecular targets to enhance aged muscle adaptability.

overall_design: Bulk RNA-Seq (n=6-7/group), Myonuclear RRBS (n=3-4/group), and single myonucleus RNA-Seq (n=1 replicate/group) profiling of adult and aged HSA-GFP mouse plantaris muscle that underwent 72 hours of synergist ablation mechanical overload (MOV) or sham (control) surgery.

first: Ronald

last: Jones III

first: Ana Regina

last: Cabrera

first: Kevin

middle: A

last: Murach


title: A genome-wide DNA methylation survey reveals salicylic acid-induced distinct hypomethylation linked to defense responses against biotrophic pathogens

city: Kifissia

first: Theoni

last: Margaritopoulou

state: Attiki

address: Stefanou Delta 8

department: Phytopathology

country: Greece

institute: Benaki Phytopathological Institute

summary: DNA methylation is a conserved regulatory mechanism of gene expression, genome stability, and development, and highly associated with effective induction of defense responses for plant priming. In the Green Deal era, the use of plant defense inducers (PDIs), compounds that activate defense and prime plants against imminent pathogen attacks, is a safe and environmentally sustainable approach to support plants against pathogens. Even though efforts have deciphered part of PDIs mode of action, more information is needed to understand the underlying pathways of their effectiveness. Here, salicylic acid (SA) loaded in chitosan nanoparticles, increased hypomethylation more than 25% on 56 genomic regions that corresponded to defense-related genes, such as pectin lyases, defensins and leucine-rich repeat transmembrane protein kinases against the biotrophic fungal pathogen Podosphaera xanthii. A genomic region of the promoter of SKP1A, a core member of the SCF E3 ubiquitin ligase complex, was found to be a Differentially Methylated region (DMR) with 60% hypomethylation, both after PDI application and pathogen inoculation, possibly indicating a similar activation mechanism. Examination of this DMR revealed the presence of SA-, auxin-, and defense-related cis-elements. Investigation of proteins associated with the above cis-elements showed significant upregulation in expression after PDI. Moreover, association of the identified DMR with transcriptomics showed enrichment of the SA pathway. Overall, these findings shed light on the epigenetic mechanisms that underlie SA- related defense priming in plants

overall_design: All sequencing experiments were performed at Biomedical Research Foundation Academy of Athens on an Illumina NovaSeq 6000 sequencing system. Libraries were sequenced with 2 × 150 bp paired end reads at 30× coverage. According to NIH Roadmap Epigenomics Project the use of two biological replicates with a combined total coverage of 30× in sufficient for unbiased genome-wide DNA methylation profiling.

first: Spyros

last: Foutadakis

first: Giannis

last: Vatsellas

first: Martina

last: Samiotaki

first: Emilia

last: Markellou