title: Dysregulated NK Cell Activation and Myeloid-Lymphoid Imbalance Underpin COPD Progression: Insights from High-Dimensional Immune Profiling and Smoking-Induced Immune Remodeling

city: Beijing

first: Qin

last: Qiao

address: Tsinghua Garden

institute: Tsinghua University

summary: Background: Chronic obstructive pulmonary disease (COPD) is a significant global health concern, marked by persistent inflammation and immune dysregulation. Although it is widespread and has substantial clinical implications, the systemic immune mechanisms driving disease progression are not fully understood. Since blood contains a diverse array of immune cells and offers a non-invasive means of assessing immune homeostasis and overall physiological status, investigating immune dysregulation through blood sampling offers considerable value for both basic research and clinical application. This approach can provide novel insights into the pathogenesis of COPD. Methods: This study employed high-dimensional flow cytometry and RNA sequencing to comprehensively characterize peripheral immune cells from a cohort of 69 COPD patients spanning clinical stages 1 to 4, alongside 41 healthy donors as controls. To capture granulocyte populations typically excluded from peripheral blood mononuclear cell analyses, fresh whole blood samples were analyzed directly. Results: Our study revealed a marked shift in the myeloid-lymphoid balance, characterized by elevated neutrophils, eosinophils, and classical monocytes that correlated with disease severity, alongside reduced CD8⁺ T cells and circulating T follicular helper cells. Transcriptional profiling identified oxidative stress pathways, T cell suppression, and aberrant natural killer (NK) cell activation as hallmarks of advanced COPD. Notably, activated Nkp44⁺ NK cells were significantly enriched in severe stages, implicating their role in perpetuating inflammation. Smoking exacerbated immune perturbations, including upregulated complement activation and B cell pathways, though cessation partially restored transcriptional homeostasis. Conclusions: This study underscores the value of peripheral immune profiling in capturing the heterogeneity of COPD. The results reveal systemic immune dysregulation-especially NK cell hyperactivity and point to potential therapeutic avenues aimed at modulating immune responses to slow disease progression.

overall_design: To elucidate the immune perturbations associated with COPD progression, we recruited 41 healthy donors and 69 patients with COPD spanning a spectrum of clinical severities: 19 with mild (stage 1), 26 with moderate (stage 2), and 24 with severe (stages 3-4) COPD. Flow-cytometric analysis was performed on all peripheral blood samples. To explore the transcriptomic changes underlying COPD pathogenesis, we randomly selected 35 of these samples for bulk RNA-seq: 10 from healthy donors and 25 from patients (stage 1, n = 6; stage 2, n = 12; stages 3–4, n = 7).

first: Yazheng

last: Yang

first: Ke

last: Huang

first: Hongtao

last: Niu

first: Xiaoxia

last: Ren

first: Shiwei

last: Qumu

first: Wei

last: Li

first: Chen

last: Dong

first: Ting

first: Ling

last: Ni


title: Canopy1/Cnpy1 is required for proper V2R processing and transport; its functional loss impairs basal/V2R vomeronasal sensory neurons function and circuit organization.

status: Public on Nov 18 2025

city: Albany

first: Paolo

middle: Emanuele

last: Forni

state: NY

address: 1400 Washington Avenue

department: Biology

country: USA

institute: University at Albany

type: Expression profiling by high throughput sequencing

summary: The Vomeronasal Organ (VNO) of rodents is populated by different types of Vomeronasal Sensory Neurons (VSNs). Apical VSNs, located near the lumen, express the transcription factor(TF) Meis2, V1R family receptors, and the G protein subunit Gao; the VSNs distributed closer to the basal lamina express the TF Tfap2e/AP-2ε, V2R receptors, and the G protein subunit Gai2. In addition sparse cells in the VNO express  the Formyl Peptide Receptors (FPRs). Single-cell mRNA sequencing (scRNA Seq) identified over 300 differentially expressed genes between these cell types, with many linked to the endoplasmic reticulum (ER). Among these ER proteins, Canopy1 (Cnpy1), was found to be among the most enriched genes in V2R+ VSNs. Only previously studied in zebrafish, Cnpy1 was found to affect Fgfr1 signaling and thus is also known as “FGF signaling regulator-1”. In a previous study we discovered that AP-2e upregulates Cnpy1 expression. Although Cnpy1 knockout mice are viable and have normal VNO development at birth, they experience a progressive degeneration and loss of V2R-expressing VSNs. Before symptoms appear, the basal VSNs of KO mice display reduced V2R protein immunoreactivity in the soma and a complete absence of the protein in the lumen of the VNO, rendering the neurons non-functional. Cnpy1 KOs exhibit altered guidance cues and adhesion molecule expression, along with disrupted connectivity to the accessory olfactory bulb.  Our study shows that distinct neuronal types depend on unique ER protein repertoires to maintain proper proteostasis. The loss of Cnpy1 highlights the importance of cell–type–specific ER factors in the differentiation and function of specific neurons, revealing mechanisms that drive neuronal diversity and vulnerability to ER gene disruption.

overall_design: Whole postnatal vomeronasal organs were dissected from P21 (Cnpy1 KO) C57BL/6J-Cnpy1em1cyagen mice, cells were isolated then analyzed using scRNAseq.

first: Nick

middle: A

last: Mathias

first: Nikki

middle: M

last: Dolphin

first: Anthony

last: Ruquet

middle: E


title: Spheroid cultures reveal cancer stem cell enrichment and distinct chemotherapeutic response in triple-negative breast cancer

city: Gothenburg

first: Toshima

middle: Z

last: Parris

address: Sahlgrenska Center for Cancer Research, Medicinaregatan 1G, 6th floor

country: Sweden

institute: University of Gothenburg

summary: Three-dimensional (3D) cancer spheroid models provide physiologically relevant platforms for studying tumor biology and therapeutic response. Using the liquid overlay method, we optimized conditions for reproducible spheroid generation from MDA-MB-231 breast cancer cells and extended this approach to 10 human cancer cell lines. Growth outcomes varied, yielding compact spheroids, loose aggregates, or no spheroids, with compactness strongly associated with high breast cancer stem cell (BCSC) content (≥20%). Transcriptomic profiling revealed distinct gene expression programs between spheroid-forming and non-forming cells, highlighting pathways in extracellular matrix remodeling, differentiation, and developmental lineage specification. Notably, PROM1, HOXB4, BMP5, and TENM4 emerged as key regulators, with TENM4 consistently upregulated across compact spheroid models. Drug response assays demonstrated increased chemoresistance in 3D spheroids compared to 2D cultures, with triple-negative breast cancer (TNBC) spheroids exhibiting reduced sensitivity to bortezomib+nedaplatin despite context-dependent synergy. Collectively, these findings establish optimized experimental parameters for spheroid culture, define molecular features linked to spheroid formation and stemness, and underscore the importance of 3D models for evaluating therapeutic efficacy.

overall_design: RNA-seq profiling of cancer cells grown as 2D monolayers and 3D cancer spheroids (Day 10 and Day 20 after seeding).

first: Sithumini

last: Sarathchandra

first: Chiara

last: Fittipaldi

first: Nanfizat Abiket

last: Alamukii

first: Anikó

last: Kovács

first: Toshima Z


title: Translatome and Translation dynamics analysis of a RiboCancer cell line panel reveals that leukemia-associated Rps15 Mutations Rewire Translation through Codon-Specific tRNA Accommodation defects.

city: Leuven

first: Marino

last: Caruso

address: Herestraat 49

department: Oncology

country: Belgium

institute: KULeuven

summary: Various somatic deletions and point mutations targeting ribosomal proteins (RPs) have been identified in cancer. To assess how these affect ribosomal translation, we applied genomewide translatome analysis (proteome, Ribo-seq and total RNA-seq) on an isogenic cell line library modeling the most recurrent RP defects (Rpl5+/-, Rpl11+/- , Rpl22+/- , Rpl22-/-, Rpl10 R98S, Rps15 P131S and Rps15 H137Y). We detected little translational changes in the RP knock-out models. In contrast, RP point mutations were associated with a significant number of translation efficiency changes, reaching up to 10% of expressed genes in Rps15 mutants associated with Chronic Lymphocytic Leukemia (CLL). Further cryo-electron microscopy and biochemical analyses revealed that these Rps15 mutations alter the stability of the C-terminal Rps15 domain, affecting the translation elongation cycle dynamics of these mutant ribosomes, and deregulating accommodation of aminoacylated tRNAs at the ribosomal A-site. Using Ribo-seq and translation reporter assays, we show that this accommodation defect shows codon specificity, explaining the reduced translation efficiency of genes enriched for these codons in Rps15 mutant cells, such as histones. Finally, the genes presenting reduced translation efficiency in Rps15 mutants are enriched for transcriptional regulators such as transcription factor Runx3, resulting in downregulation of Runx3 target genes involved in immune regulation. Overall, this study compares the translational rewiring imposed by the most frequent somatic RP mutations. We provide unprecedented mechanistic insights in the translation defects induced by CLL-associated Rps15 mutations, and reveal an unappreciated cross-talk between translational and transcriptional dysregulation in these RP mutant cells.

overall_design: RNA-seq and RIBO-seq profiling of a RiboCancer cell line panel including CRISPR-CAS9 clones of ribosomal protein defects (WT, Rpl5+/-, Rpl11+/- , Rpl22+/- , Rpl22-/-, Rpl10 R98S, Rps15 P131S and Rps15 H137Y) generated from Ba/F3 mouse cell line.

first: Anaïs

last: Astier

first: Stijn

last: Vereecke

first: Coralie

last: Capron

first: Steven

last: Verbruggen

first: Gerben

last: Menschaert

first: Pierre

last: Close

first: Francesca

last: Rapino

first: Célia

last: Plisson-Chastang

first: Kim

last: De Keersmaecker


title: Evolutionary remodeling of the ventral retina enables aerial vision in the four-eyed fish Anableps anableps

city: Baton Rouge

first: Patricia

last: Schneider

state: Louisiana

address: LSU 131 Life Sciences Building

department: Biological Sciences

institute: Louisiana State University

summary:
The emergence of evolutionary novelties remains a central challenge in biology, particularly for
complex organs such as the vertebrate eye. While visual systems have repeatedly undergone
degeneration or loss, examples of innovation are rare and poorly understood. Mechanistic insight into
such gains requires tractable systems in which novel visual functions have evolved. Here we leverage the
four-eyed fish Anableps anableps to uncover the cellular and molecular basis of adaptation to
simultaneous aerial and aquatic light. Combining single-nucleus and spatial transcriptomics, we show that
the Anableps ventral retina has been developmentally and functionally reconfigured for aerial vision. In
fully aquatic teleosts such as zebrafish, ventral photoreceptors are tuned to red-shifted, downwelling light.
In contrast, we show that Anableps exhibits an expanded and diversified ventral cone population
encompassing ultraviolet-, blue-, green-, and red-sensitive types, enabling broad-spectrum, high-intensity
vision from above water. To estimate Anableps opsin spectral properties, we employed OPTICS, a
machine-learning model of opsin spectral tuning, which predicts that the ventrally expressed opn1mw2
opsin is red-shifted relative to typical green-sensitive pigments. Analysis of dorsal-ventral patterning
markers reveals an overrepresentation of ventral retinal cell types. Consistent with this, comparative
spatial transcriptomic profiling of developing and adult Anableps and the killifish (Fundulus heteroclitus)
indicates a dorsal displacement of the dorsal-ventral boundary in Anableps, resulting in a proportionally
larger ventral retinal domain. Collectively, our results reveal that Anableps achieved its unique dual visual
capacity by developmentally expanding and functionally reprogramming the ventral retina for aerial light
detection, an evolutionary innovation that redefines how retinal domains can adapt to radically different
visual environments.

overall_design: Single nucleus suspensions were prepared from the adult whole retina of Anableps. Nuclei were fixed and frozen using the Evercode Nuclei Fixation Kit v3 (#NF100, #NF200) from Parse Biosciences following the manufacturer’s protocol. Fixed and frozen nuclei were then processed to generate barcoded libraries aiming for 5,000 nuclei per sample using the Evercode WT Mini v3 Kit (#ECWT3100) as per manufacturer’s instructions. Resulting libraries were sequenced by Novogene at the depth of 20,000 reads per nuclei in an Illumina Novaseq Xplus platform (paired-end, 150bp).

first: Louise

last: N. Perez

first: Josane

last: de Freitas Sousa


title: Integration of Network Pharmacology, Transcriptomics and Single-cell Sequencing to Explore the Effect of Rougan Keli in Alleviating Liver Cirrhosis

city: Nanjing

first: xujing

last: Gu

address: Xianlin Street, Qixia District, Nanjing, Jiangsu Province, China

country: China

institute: Nanjing Unicersity of Chinese Medicine

summary: Background: The progression of liver cirrhosis leads to severe complications, significantly threatening the survival and prognosis of patients. Rou gan ke li (Rgkl), a herbal formula derived from classical prescriptions, has used clinically over two decades and has good efficacy. However, its molecular mechanisms and active components remain undefined. Purpose: Exploring the molecular mechanisms of Rgkl in alleviating liver cirrhosis. Methods: CCl4-induced liver cirrhosis mice models were established. Liver stiffness and intrahepatic blood flow velocity were assessed using imaging. Serum ALT, AST, HA, and histopathology were analyzed. Hepatic stellate cells (HSCs) activation, liver sinusoidal endothelial cells (LSECs) fenestration, and angiogenesis were evaluated using immunohistochemistry and scanning electron microscopy. UPLC-Q-TOF-MS/MS and network pharmacology identified active components. Transcriptomics and single-cell sequencing identified key targets and pathways, validated via WB, immunofluorescence, and molecular docking. Results: Rgkl significantly reduced Liver stiffness and collagen deposition while increasing intrahepatic blood flow velocity in cirrhotic mice. Serum ALT, AST, and HA were markedly decreased. Rgkl inhibited α-SMA expression in HSC and downregulated pathological angiogenesis by reducing VEGF and CD34 expression. Additionally, Rgkl enhanced eNOS expression and preserved sinusoidal fenestration in LSEC. Furthermore, Rgkl ameliorated liver cirrhosis by modulating LSEC metabolic functions via the CD36/PPAR/CPT-1 pathway and suppressing HSC activation through the RhoA/ROCK/YAP and PI3K/AKT/NF-κB pathways. Eighteen active components, such as Levistilide A and Quercetin, were strongly correlated with the amelioration of liver cirrhosis. Conclusions: Rgkl significantly attenuated hepatic injury and fibrosis. Mechanistically, Rgkl modulated LSEC lipid metabolism and phenotypic regulation, and suppressed HSC contraction and activation. Key active components contributing to these effects included Paeonilactone C, Levistilide A, and Quercetin.

overall_design: LSEC and HSC are key effector cells that promote cirrhosis progression. In the first half of this study, although we precisely enriched the core signaling pathway interfered by Rougan Keli through transcriptomics, its effect on LSEC and HSC needs to be further clarified. Therefore, with the help of single-cell sequencing technology, we effectively differentiated various types of cells in the liver and identified 10 types of cells including LSEC and HSC.

first: Chengbo

last: Jin

first: Tianle

last: Ma

first: Yiheng

last: Zhang

first: Xujing

first: Tong

last: Zhu

first: Xinyu

last: Wu

first: Xin

last: Ding

first: Suzhou

last: Huang

first: Yulan

last: Wang

first: Zhipeng

last: Chen

first: Huihua

last: Fang

first: Li


title: LincRNA-EPS regulates iron metabolism and osteoclastogenesis under inflammatory micro-environment

city: Shanghai

first: Jin

address: Yanchang Road

institute: Shanghai Tongji Stomatological Hospital and Dental School

summary: This study aims to investigate the impact of LincRNA-EPS on osteoclast lineage differentiation. We established LincRNA-EPS gene knockout mice on the C57BL/6 background and isolated bone marrow-derived macrophages (BMDMs) from both knockout and wild-type mice. These BMDMs were induced with RANKL in vitro for 2 days to generate pre-osteoclast cells. Transcriptome sequencing was performed on these cells, followed by further analysis of differentially expressed genes to explore the effects of LincRNA-EPS deficiency on osteoclast differentiation in BMDMs. In this study, we found that dysregulated osteoclastogenesis and iron homeostasis without lincRNA-EPS, represented by up-regulated gene Lcn2.

overall_design:
Experimental Grouping: KO group: Bone marrow-derived macrophages (BMDMs) isolated from LincRNA-EPSknockout mice (LincRNA-EPS knockout); WT group: BMDMs isolated from wild-type (C57BL/6) mice. Each group included three biologically independent replicates.
Experimental Procedure: BMDM induction:BMDMs from both groups were cultured in complete α-MEM medium supplemented with 40 ng/mL M-CSF for 24 hours to promote macrophage proliferation and priming.Osteoclast precursor differentiation:After initial M-CSF priming, 40 ng/mL RANKL was added to the culture medium for an additional 48 hours to induce pre-osteoclast formation .
Sample processing:Cells were harvested and total RNA was extracted.
Transcriptome sequencing:RNA sequencing was performed on the Illumina NovaSeq X plus platform.

first: Yabing

first: Zhanwei

first: Jiansheng

last: Su


title: The hereditary spastic paraplegia type 21 (SPG21) protein is a RAB7A effector that promotes non-canonical mTORC1-catalyzed TFEB phosphorylation and cytoplasmic retention

city: Bethesda

first: Ryan

last: Dale

state: MD

address: Rm 10D39, 10 Center Drive

department: National Institute of Child Health and Human Development

institute: National Institutes of Health

summary: Hereditary spastic paraplegia type 21 (SPG21) is an inherited neurological disorder caused by mutations in the SPG21 gene, which encodes a protein named SPG21 or maspardin. Herein, we report that SPG21 localizes to endolysosomes through interaction with the GTP-bound form of RAB7A. Disease-associated SPG21 variants reduce expression of SPG21 and disrupt its endolysosomal localization in both non-neuronal cells and neurons. Consistent with this localization, functional dependency analysis links SPG21 to endolysosomal and mTORC1 signaling pathways. Biochemical studies reveal that SPG21 depletion does not affect phosphorylation of canonical mTORC1 substrates such as ULK1, S6K1, 4E-BP1, but reduces phosphorylation of the non-canonical mTORC1 substrate TFEB. This enhances nuclear localization of TFEB and upregulation of TFEB-target genes. We conclude that SPG21 acts as a RAB7A effector that promotes non-canonical mTORC1-catalyzed phosphorylation of TFEB, thereby suppressing its nuclear localization and transcriptional activity. These findings link SPG21 dysfunction to altered endolysosomal signaling, offering new insights into SPG21 pathogenesis.

overall_design: RNA-seq profiling of wild-type HeLa with or without transient knockdown of the protein SPG21.

first: Jennifer

middle: K

last: Kunselman

first: Chad

middle: D

last: Williamson

first: Adriana

last: Golding

first: Rui

last: Jia

first: Mira

last: Sohn

first: Juan

middle: S

last: Bonifacino


title: Single cell transcriptomics of mouse abdominal aortic aneurysm induced by AngII plus salt [28days]

first: xianxian

address: Building 5, Panjiayuan Nanli, Chaoyang District, Beijing

institute: Institute of Laboratory Animal Science: Beijing, CN

summary: In this study, we constructed mouse abdominal aortic aneurysm (AAA) model by adiministration of AngII plus salt. We conducted single-cell RNA sequencing (scRNA-seq) to profile gene expression on various cell types and explored the endothelial cell heterogeity in the development of AAA.

overall_design: Single cell RNA-Seq of mouse abdominal aorta from control group and AngII and salt group, by pooling aortic tissues from five mice as one sample.

first: Xianxian

first: Zhiwei

last: Yang


title: RNA Sequencing of Brain Tissue from Estuarine Dolphins

city: Miami

first: David

last: Davis

state: Florida

address: 1951 NW 7th Avenue

department: Neurology

institute: University of Miami Miller School of Medicine

summary: Climate warming is one factor increasing the severity of harmful algal blooms (HABs). Innovative exposure models are needed to understand how HABs affect brain health. Here, we examined HAB exposure on the brain transcriptome of dolphins found stranded in Florida’s Indian River Lagoon. We report the neurotoxin 2,4-diaminobutyric acid (2,4-DAB) is 2,900 times more concentrated in dolphin brains during bloom seasons compared to non-bloom seasons. The same dolphins show 536 differentially expressed genes whose enrichment reveal impairment in GABAergic synapses, basement membrane alteration, and Alzheimer’s disease (AD) risk factors that increase with each subsequent season. Dolphins also display concurrent AD-like neuropathological changes and elevated AD gene expression with 2,4-DAB exposure. Our study demonstrates disproportionate seasonal exposure to 2,4-DAB increases AD signatures in the brain transcriptome. As our climate warms, HABs will continue to intensify. Understanding the impact of HAB exposures will help to identify populations at risk for neurological illnesses.

overall_design: RNA Seq of dolphin brain samples collected during bloom seasons vs non-bloom seasons

first: Susanna

middle: P

last: Garamszegi


title: BRD9 CFT8634 ChIP Seq(primary cells)-Highly discriminative globin gene activation by the non-canonical BAF chromatin remodeling complex

city: Philadelphia

first: Eugene

last: Khandros

state: PA

address: 3615 Civic Center Blvd.

department: Hematology

institute: The Children's Hospital of Philadelphia

type: Genome binding/occupancy profiling by high throughput sequencing

summary: The regulation of beta-like globin genes, including the switch from fetal (HBG) to adult (HBB and HBD) genes has served as a paradigm of developmental transcriptional control. While HBG repressors such as BCL11A and LRF have been well characterized, recent discoveries of HIF1a and BGLT3 as HBG-specific activators have raised interest in finding new activators as potential therapeutic targets. The ATP-dependent chromosome remodeler BRG1 has previously been demonstrated as necessary for hemoglobin gene transcription. In loss of function studies in adult-like HUDEP2 cells, we found that BRG1 depletion preferentially decreased fetal HBG and the minor adult HBD. BRG1 forms multisubunit complexes with combinations of 27 subunits in 3 families - canonical BAF, polybromo BAF, and non-canonical BAF (ncBAF) - to regulate chromatin accessibility and gene transcription in a cell and condition-specific manner. Using CRISPR loss of function studies in HUDEP2 cells and primary adult erythroid cells, we identified the non-canonical BAF complex as a positive regulator of HBG and HBD. Loss of the non-canonical BAF complex subunits BRD9 and BAF60A/SMARCD1 preferentially decreased HBG and HBD in HUDEP2 cells and primary adult erythroid cultures, while accelerating erythroid differentiation and hemoglobinization. Pharmacological depletion of BRD9 led to acute decreases in chromatin accessibility at HBD and HBG promoters and reduced their primary transcripts, while longer treatment recapitulated  genetic ablation effects. This surprising demonstration of BAF complex selectivity within a multi-gene cluster extends our understanding of globin gene transcriptional regulation and identifies a potential target for therapeutic manipulation of beta-like globins.

overall_design: We performed ChIP seq analysis in 3 healthy donor erythroid cells derived from CD34+ HSPC after DMSO or CFT8634 treatment. Chromatin immunoprecipitation (ChIP) was performed on Phase 3 Day3 using the following antibodies: BRD9 (Abcam, Anti-BRD9 antibody [EPR23888-5], 5ug/IP), H3K27ac (Abcam, Anti-Histone H3 (acetyl K27) antibody ab4729 - ChIP Grade, 5ug/IP). ChIP-seq libraries were prepared using NEBNext Ultra II DNA Library Prep Kit with NEBNext unique dual index primer pairs.

first: Abhirup

last: Bagchi

first: Sindhu

last: Billakanti

first: Vanessa

last: Shehu

first: Kathryn

last: Farrell

first: Sarah

last: Cotteta

first: Bailey

middle: N

last: Godfrey

first: Micaela

last: Alpert

first: Jacob

middle: F

last: Zeranski

first: Krista

last: Budinich

first: Zhuangzhuang

last: Geng

first: Osheiza

last: Abdulmalik

first: Belinda

last: Giardine

first: Cheryl

last: Keller

first: Ross

middle: C

last: Hardison

first: Junwei

last: Shi


title: BRD9 CFT8634 ATAC Seq (HUDEP-2 clones)-Highly discriminative globin gene activation by the non-canonical BAF chromatin remodeling complex

overall_design: We performed ATAC seq analysis in 4 HUDEP-2 clonal lines generated using single cell cloning of a pool of HUDEP-2 cells. ATAC-seq was conducted following the Omni-ATAC protocol (https://www.nature.com/articles/nmeth.4396). In brief, 100,000 cells underwent lysis using a buffer containing 0.1% NP-40, 0.1% Tween-20, and 0.01% digitonin, followed by incubation with Tagment DNA Enzyme (Illumina cat # 20034197) at 37°C for 30 minutes. DNA was subsequently purified using the QIAGEN MinElute Reaction Cleanup Kit. For library preparation, fragmented DNA was amplified with Phusion High-Fidelity PCR Master Mix supplemented with HF Buffer (Thermo Fisher Scientific, catalog no. F531S) and indexed primers featuring unique dual barcodes. Libraries were purified with AMPure XP beads (Beckman Coulter, catalog no. A63881) at a 1:1 ratio, adhering to the manufacturer’s guidelines.


title: Control and Tgfbr2 cKO Mouse Endometrial Epithelial Organoids +/- E2/MPA Treatment

city: Houston

first: Peixin

last: Jiang

state: TX

address: One Baylor Plaza

institute: Baylor College of Medicine

summary: Evaluate the changes in estrogen and progesterone response within the endometrial epithelium of mice with and without uterine Tgfbr2 expression.

overall_design: Endometrial epithelial organoids from control and Tgfbr2 cKO mice were cultured for two days prior to treatment with either vehicle (6 days) or 10nM E2 (2 days) + 10 nM E2/1 µM MPA (4 days). After six days of treatment, organoids were collected in trizol and RNA was isolated for RNAseq.

first: Sydney

last: Parks

first: Diana

last: Monsivais


title: Highly discriminative globin gene activation by the non-canonical BAF chromatin remodeling complex.

first: Eugene

last: Khandros

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

overall_design: Refer to individual Series

first: Abhirup

last: Bagchi

first: Sindhu

last: Billakanti

first: Michaela


title: Efferocytosis-induced metabolic shift in bone macrophages drives lactate production and modulates inflammation and osteoclastogenesis

city: ANN ARBOR

first: Hernan

last: Roca

state: MI

address: 1011 N. University Ave.

department: Periodontics and Oral Medicine

institute: University of Michigan

summary: During the critical process of homeostatic efferocytosis, macrophages clear apoptotic cells and subsequently transition to reparative functions that promote the resolution of inflammation and support tissue repair. Their inherent plasticity enables rapid changes in macrophage activity suited to specific microenvironments. However, the heterogeneity in their cell states also presents challenges in characterizing subsets of macrophages and analyzing their specific contributions post-efferocytosis. In this study, single-cell RNA sequencing data from bone-marrow derived macrophages engulfing apoptotic osteoblasts (OB) was used to characterize macrophage subpopulations enriched during efferocytosis. Clustering analysis revealed two subpopulations (c3 and c9) that were unique to efferocytic macrophages. These distinct subpopulations displayed a transcriptional profile characterized by enhanced glycolytic energy metabolism, along with an anti-inflammatory gene signature. Notably, HIF-1 signaling, glycolysis/gluconeogenesis, and carbon metabolism were among the top five most significantly enriched pathways in c3 and c9 macrophages. qRT-PCR analysis revealed that macrophages engulfing apoptotic OBs exhibited increased expression of key glycolytic enzymes and solute carriers, including Slc2a1, Pdk1, Ldha, and Slc16a3. Metabolomics analysis revealed a significant increase in intracellular lactate, phosphoenolpyruvic acid, glycerol-3-phosphate, 2-/3-glycerophosphate, and fructose-6-phosphate, indicative of enhanced glycolysis. In addition, efferocytic macrophages showed increased extracellular lactate production compared to control macrophages, as confirmed by lactate ELISA. The effects of lactate (0-20mM) on osteoblast mineralization, osteoclast differentiation and function, and macrophage-derived inflammatory factors were evaluated through various in vitro experiments. While no effect was seen in osteoblast mineralization, high lactate concentrations significantly reduced the number of multinucleated osteoclasts and their resorptive activity. Interestingly, extracellular lactate also significantly upregulated M2-like macrophage markers (Arg1, Il1rn, Klf4). These results support the concept that macrophage efferocytosis of apoptotic osteoblasts alters macrophage energy metabolism, which in turn plays a distinct and pivotal role in modulating the bone microenvironment.

overall_design: single-cell RNA sequencing (scRNA-seq) was performed using the 10X Genomics platform on flow-sorted control macrophages (F4/80-APC+) and efferocytic macrophages engulfing Cell-Trace-stained apoptotic osteoblasts, 1 sample per group

first: Nicholas

middle: J

last: Carruthers

first: Rahasudha

last: Kannan


title: Pan-cyclin-dependent kinase inhibition is a potential treatment for adenoid cystic carcinoma that downregulates the MYB::NFIB fusion and induces tumor regression

first: Mattias

last: Andersson

address: Medicinaregatan 1F

department: Sahlgrenska Cancer Center

summary: Adenoid cystic carcinoma (ACC) is a MYB-driven, aggressive glandular cancer with a poor long-term prognosis that is resistant to systemic therapies. Therefore, there is an urgent need to develop novel therapeutic strategies for these patients. An in vitro small-molecule inhibitor screen identified sensitivity of cultured ACC cells to pan-CDK inhibitors. We here used RNA-seq analysis to investigate the cell biological and molecular effects of treatment with the pan-CDK inhibitor dinaciclib in ACC cells from two different cases. Our results show that dinaciclib treatment led to a downregulation of genes involved in cell cycle progression, DNA replication, and DNA repair. Upregulated genes affected programmed cell death and included genes involved in proteolysis and autophagy. ACC MYB signature genes were significantly downregulated in dinaciclib-treated ACC cells, indicating that dinaciclib perturbs MYB-driven gene expression.

overall_design: Cultured ACCX11 and ACCX67 cells treated with 25 nM dinaciclib or DMSO (vehicle control) for 24 h.

first: Junchi

last: Huang


title: LsCLE12 peptide delays senescence in lettuce and prolongs postharvest life of roses

city: Singapore

first: Sally

last: Koh

address: 1 Research Link

country: Singapore

institute: Temasek Life Sciences Laboratory

summary: The widespread use of chemical inputs in agriculture has raised concern over their long-term environmental and health impacts, driving demand for sustainable, natural product-based alternatives. Peptide-hormone signaling interactions, particularly those regulating senescence, are promising targets for developing such alternatives to advance postharvest technologies. However, most current insights stem from model species, limiting their translation to crops and ornamental plants. This study identifies the CLAVATA3/EMBRYO-SURROUNDING REGION-related (CLE) family CLE12 peptide (CLE12p) in Lactuca sativa as a peptide that shows potential for postharvest management of crops and ornamental flowers. CLE12p interacts with salt (NaCl), abscisic acid (ABA) and ethylene pathways, influencing lettuce development and morphology. Notably, CLE12p induced stomatal closure in a similar manner to that of ABA, suggesting overlapping signalling pathways. Transcriptomic analyses revealed how CLE12p modulates immune and defence responses, fluid transport, and leaf senescence, indicating a broader role in leaf senescence regulation similar to that of ethylene. Functionally, CLE12p delayed senescence in lettuce by preserving leaf greenness and water content following dark treatment and harvest. Additionally, CLE12p extended the longevity of cut roses by maintaining fresh weight and water content. These findings demonstrate the practical potential of CLE peptides in non-model species and support their application as natural tools for extending postharvest shelf life in crops and ornamental plants, paving the way for further exploration of their role in agricultural biotechnology.

overall_design: L. sativa var. Little Gem were grown on MS basal medium under 24 h continuous light at 22°C. Four-day-old seedlings were transferred to the following treatment for 2 h incubation: control (H2O), ethanol control (0.02% ethanol), 10 µM ABA, 10 µM ACC, 10 µM CLE12p, 10 µM ABA + CLE12p and 10 µM ABA + ACC.

first: Daisuke

last: Urano


title: Single-cell mapping of the glycocalyx in the adult human and mouse ocular surface

city: Boston

first: Pablo

last: Argüeso

state: MA

address: 800 Washington St

department: Ophthalmology

institute: Tufts Medical Cener

summary: The glycocalyx is a critical but often underappreciated modulator of cellular behavior. Its diversity across cell types within tissues remains poorly understood, but recent advances in single-cell profiling now enable more precise analysis of cell surface composition. Here, we applied single-cell glycan and RNA sequencing to profile glycocalyx diversity across human and mouse ocular surface cell types. Glycocalyx patterns effectively distinguished epithelial subtypes, with corneal epithelial cells enriched in complex and high-mannose N-glycans, conjunctival cells in fucosylated structures, and goblet cells in O-glycans. We also observed dynamic changes during epithelial maturation, marked by regulated shifts in sialic acid structures. In the mouse ocular surface, glycocalyx patterns distinguished major cell types, but the glycan profiles differed from those in humans, pointing to species-specific features. These findings demonstrate that glycocalyx composition is closely linked to cell identity and maturation and provide a foundation for exploring its roles in tissue organization and disease.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Single-cell RNA sequencing (scRNA-seq) analysis, utilizing nine distinct lectins conjugated to DNA, to characterize the glycocalyx composition of human and mouse ocular surface cells. The experiment consists of three scRNA-seq capture reactions per species, leveraging lectin-based profiling to reveal glycocalyx diversity

first: Rafael

last: Martínez-Carrasco


title: Lyn expression in macrophages promotes TLR activation and restricts proliferation in an isoform-independent manner

city: Minneapolis

first: Anders

middle: John

last: Lindstedt

state: MN

address: 2101 6th Street SE

department: Department of Pharmacology

institute: University of Minnesota

summary: Toll-like receptor (TLR) signaling is vital to macrophage function, and dysregulation is associated with many disease states. The Src-family kinase (SFK) Lyn serves activating and inhibitory roles downstream of TLRs, yet distinct functions of the isoforms LynA and LynB in TLR signaling have not been investigated. We used isoform-specific knockout mice (LynAKO and LynBKO) to interrogate the contribution of each isoform to TLR signaling in bone marrow-derived macrophages (BMDMs). Bulk RNA sequencing and cytokine analyses revealed that complete Lyn deficiency (LynKO) dampens TLR4- and TLR7-induced inflammatory gene expression and TNF production, but enhances extracellular matrix (ECM) gene expression and promotes cell-cycle progression and macrophage proliferation. In contrast, expression of either LynA or LynB was sufficient to preserve wild-type (WT) transcriptional responses and TNF production, despite moderately reduced total Lyn levels. These data suggest that Lyn promotes macrophage activation downstream of TLRs and restrains aberrant proliferation and matrix deposition in a largely dose-dependent rather than isoform-specific manner. Our findings provide new insight into the roles of LynA and LynB in TLR signaling and can inform future studies of macrophage-driven pa-thology in autoimmune disease, fibrosis, and cancer.

overall_design: RNA sequencing of bone marrow-derived macrophages (BMDMs) from wild-type (WT), LynAKO, LynBKO, and total LynKO mice under different cell-culture conditions. BMDMs were differentiated for 7-days in macrophage-colony stimulating factor (M-CSF)-containing media, rested overnight, and then treated for 2h with M-CSF-free media (NT), lipopolysaccharide (LPS), or resiquimod (R848). Samples are biological replicates, with cells derived from 4 mice of each genotype (2 male and 2 female), independently differentiated and treated.

first: Tanya

last: Freedman


title: Single-cell RNA sequencing reveals  ulcerative colitis patients’ response to anti-TNF-α therapy

city: kunming

first: Zhangqin

last: Li

address: No. 295, Xichang Road, Wuhua District, Kunming City, Yunnan Province

institute: The First Affiliated Hospital of Kunming Medical University

summary: We performed single-cell sequencing of intestinal mucosal tissue from eight UC patients at pre- and post- anti-TNF-α therapy prescription and analyzed them with relation to therapy response (response and no-response). We found that responders have different immune cell profiles from non-responders between pre- and post- anti-TNF-α therapy. Specifically, the proportion of cDC3 decreased in all responders after receiving anti-TNF-α therapy, whereas non-responders could not be distinguished from UC patients who had not received anti-TNF-α therapy. When further compared the difference of type 3 conventional dendritic cells (cDC3) from pre- and post-therapy of anti-TNF-α, distinct transcriptomes were identified in responders, while no significantly different genes were found in cDC3 from non-responders. The interactions analysis of ligands and receptors showed that the interactions between HLA-DPA1/DPB1 in fibroblast and TNFSF9 in cDC3, CD44 in fibroblast and TYROBP in cDC3 were significantly weakened in Post-R group compared with Pre-R group. Cell trajectory analysis demonstrated that the cells differentiated to cDC3 significantly decreased in the Post-R group, and the genes related MAPK signaling pathway obviously increased in these cells.

overall_design: The patients received IFX treatment for fourteen weeks. Inflammation and lesions were assessed using endoscopy and intestinal mucosa in lesions were taken before IFX treatment (Pre-) and after 14 weeks ago (Post-).

first: Ruijie

last: Ma

first: Jie

last: Jia

first: Jiarong

last: Miao

first: Yinglei


title: Spatially resolved single-cell analysis uncovers protein kinase Cδ-expressing microglia with anti-tumor activity in glioblastoma [scRNA-seq]

city: Charlestown

first: Reid

last: Mcneil

address: 149 13th Street

department: Cancer Research

institute: Mass General Hospital

summary: The glioblastoma (GBM) microenvironment contains resident immune cells with intrinsic anti-tumor potential, particularly microglia. Understanding the single-cell spatial heterogeneity and interactions between immune cells and brain tumor–initiating cells (BTICs) is essential for identifying therapeutic targets to reprogram immune responses and suppress BTIC growth. Using single-cell and spatial transcriptomics, we mapped immune cell populations in the GBM microenvironment and identified signaling networks mediating immune–cancer cell interactions. We discovered a previously unrecognized subset of microglia expressing protein kinase Cδ (PKCδ) with anti-tumor activity against BTICs. The presence of PKCδ-expressing microglia was validated in human GBM specimens. Enhancing PKCδ in microglia via adeno-associated virus or niacin increased phagocytosis of patient-derived BTICs in vitro and improved survival in GBM-bearing mice. Data from The Cancer Genome Atlas (TCGA) revealed that high PKCδ expression correlates with increased apoptosis, phagocytosis, and immune signaling pathways. These findings offer highlight PKCδ+ microglia as a promising therapeutic target in GBM.

overall_design: Using GBM mouse models, mice were either treated with niacin or untreated controls, the tumors were resected from the mice and analyzed using single-cell RNAseq.

first: Reza

last: Mirzaei

last: McNeil

first: Charlotte

last: D’Mello

first: Britney

last: Wong

first: Susobhan

last: Sarkar

first: Frank

last: Visser

first: Candice

last: Poon

first: Pinaki

last: Bose

first: Wee

last: Yong


title: Spatially resolved single-cell analysis uncovers protein kinase Cδ-expressing microglia with anti-tumor activity in glioblastoma [Spatial Transcriptomics]

overall_design: Using GBM mouse modelts, mice were either treated with niacin or untreated, the tumors were resected from the mice and analyzed using spatial transcriptomics.


title: In vivo screening for essential metabolic mediators in mouse model of lung inflammation using AAV9-ME CRISPR library

city: Beijing

first: Shuo

last: Liu

address: 5 Dong Dan San Tiao

department: Department of Immunology

institute: Chinese Academy of Medical Sciences

summary: Timely termination of innate immune response and resolution of inflammation is critical for preventing tissue damage and keeping homeostasis. Immunometabolism is verified to be important in innate immunity and inflammation. However, metabolic processes and related enzymes involved in inflammatory resolution remain largely unknown. To identify the key mediators in the metabolic regulation of inflammation resolution, we generated an AAV9-ME CRISPR library containing 17090 sgRNAs targeting 2682 mouse metabolic genes and performed an in vivo CRISPR screen in influenza virus (IAV) infection-induced lung inflammation model in Sftpc-Cas9 mice. We found that a very long chain fatty acid elongase, ELOVL5, could promote the resolution of lung inflammation. These results provide evidence for the metabolic gene-mediated regulation of immune inflammation and also insights for treating inflammatory diseases through manipulating metabolism.

overall_design: Next-generation sequencing of IAV_screen and Input sample, RNA-seq of type 2 AEC from mice infected with IAV for different times: Day0: C1,C2; Day7: I1,I2; Day12: R1,R2.

first: Xuetao

last: Cao


title: Evolutionary remodeling of the ventral retina enables aerial vision in the four-eyed fish Anableps anableps [Spatial Transcriptomics]

first: Patricia

last: Schneider

summary:
The emergence of evolutionary novelties remains a central challenge in biology, particularly for
complex organs such as the vertebrate eye. While visual systems have repeatedly undergone
degeneration or loss, examples of innovation are rare and poorly understood. Mechanistic insight into
such gains requires tractable systems in which novel visual functions have evolved. Here we leverage the
four-eyed fish Anableps anableps to uncover the cellular and molecular basis of adaptation to
simultaneous aerial and aquatic light. Combining single-nucleus and spatial transcriptomics, we show that
the Anableps ventral retina has been developmentally and functionally reconfigured for aerial vision. In
fully aquatic teleosts such as zebrafish, ventral photoreceptors are tuned to red-shifted, downwelling light.
In contrast, we show that Anableps exhibits an expanded and diversified ventral cone population
encompassing ultraviolet-, blue-, green-, and red-sensitive types, enabling broad-spectrum, high-intensity
vision from above water. To estimate Anableps opsin spectral properties, we employed OPTICS, a
machine-learning model of opsin spectral tuning, which predicts that the ventrally expressed opn1mw2
opsin is red-shifted relative to typical green-sensitive pigments. Analysis of dorsal-ventral patterning
markers reveals an overrepresentation of ventral retinal cell types. Consistent with this, comparative
spatial transcriptomic profiling of developing and adult Anableps and the killifish (Fundulus heteroclitus)
indicates a dorsal displacement of the dorsal-ventral boundary in Anableps, resulting in a proportionally
larger ventral retinal domain. Collectively, our results reveal that Anableps achieved its unique dual visual
capacity by developmentally expanding and functionally reprogramming the ventral retina for aerial light
detection, an evolutionary innovation that redefines how retinal domains can adapt to radically different
visual environments.

overall_design: Spatial transcriptomics of the Anableps anableps eye during development (stages 3, 4, and 5) and adult.

first: Louise

last: N. Perez

first: Josane

last: de Freitas Sousa


title: Pulmonary Epithelial Injury Impairs Innate Defense and Reprograms Macrophage Function, Reducing Early Mycobacterium tuberculosis Clearance

city: Tianjin

first: Xuan

address: No. 890 Jingu Road, Shuanggang Town, Tianjin, China

institute: Haihe Clinical School, Tianjin Medical University, Tianjin, China

summary: Tuberculosis (TB), caused by Mycobacterium tuberculosis (Mtb), remains a significant global health burden, characterized by complex host–pathogen interactions that drive heterogeneous clinical outcomes. While pulmonary epithelial cells are increasingly recognized as active participants in innate immunity during Mtb infection, how host defences are altered when the epithelial barrier is compromised remains unclear . In this study, we developed a murine model combining naphthalene-induced airway epithelial injury with Mtb infection, and found a pronounced impairment in pulmonary bacterial clearance. Through single-cell RNA sequencing (scRNA-seq), we mapped the pulmonary cells landscape and identified widespread suppression of epithelial immune functions. Notably, we observed macrophages transition from an antimicrobial to an antigen-presenting phenotype, indicating waning pulmonary innate defenses and heightened adaptive immune activation. These findings highlight the pivotal role of pulmonary epithelial integrity in shaping host immunity against Mtb and offer new insights into potential therapeutic strategies targeting barrier–immune crosstalk.

overall_design: This study analyzed lung tissues from four groups of C57BL/6 mice to investigate the impact of pulmonary epithelial injury on immune responses during Mycobacterium tuberculosis (Mtb) infection. The groups included: (1) untreated healthy controls (Control), (2) mice infected with Mtb H37Rv (H37Rv), (3) mice with naphthalene-induced epithelial injury alone (Naph), and (4) mice subjected to both epithelial injury and subsequent Mtb infection (Naph+H37Rv). One mouse per group was used for single-cell RNA sequencing. Lung cells were enzymatically dissociated from each mouse and processed individually. The design incorporates both infection and injury conditions to enable comparison between baseline, injury alone, infection alone, and combined injury-infection states. No technical replicates were included, and all samples were analyzed separately without pooling.

first: Xue

last: Li

first: Zuokuan

last: He

first: Guiying

last: Xu

first: Yu

first: Qi

last: Wu

first: Huaiyong

last: Chen


title: Endogenous retroviruses promote aberrant T cell differentiation in systemic lupus erythematosus via the RIG-I pathway [SLE]

first: Xiaoli

last: Min

address: No.4 Jiang Wang Temple Street

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

summary: The dysregulated differentiation of T lymphocytes play an important role in systemic lupus erythematosus (SLE). However, the underlying mechanism remains unclear. Here, we screened and identified many overexpressed HERVs in CD4+ T cells from SLE patients by RNA-seq and found that some HERVs formed dsRNAs to induce the activation of the RIG-I signaling pathway. And our findings reveal the roles and mechanism of HERVs and RIG-I in regulating the aberrant differentiation of T cells in SLE patients.

overall_design: mRNA profiles of CD4+T cells of SLE and HCs

first: Yaqian

last: Yu

first: Zhi

last: Hu


title: Epigenetic Remodeling of Sheep Oocytes and Embryos Induced by Maternal Methionine Supplementation

city: Madison

first: Hasan

last: Khatib

state: Wisconsin

address: 1675 Observatory Drive

department: Animal and Dairy Sciences

institute: University of Wisconsin Madison

type: Methylation profiling by high throughput sequencing

summary: Environmental factors, including diet, can influence gene expression and developmental outcomes through epigenetic modifications. While maternal diet is known to affect offspring DNA methylation and phenotypes, its impact on the oocyte and subsequent embryo epigenome remains less understood. Here, we investigated how maternal methionine supplementation affects DNA methylation patterns in oocytes and embryos of Polypay ewes. Whole genome bisulfite sequencing (WGBS) was performed on oocytes collected from 16 twin ewe pairs (8 methionine-treated and 8 control). These ewes were later bred to control rams, and embryos were flushed for WGBS as well. In oocytes, 2,056 differentially methylated cytosines (DMCs) were identified, with 2,034 hypomethylated and 22 hypermethylated. Additionally, 17 mitochondrial DMCs were identified, with 12 hypermethylated and 5 hypomethylated. In embryos, 113 DMCs were identified, with 112 hypermethylated and only one hypomethylated. Mitochondrial DNA analysis revealed 22 hypermethylated DMCs. To assess the inheritance of methyl marks, we compared DMCs between oocytes and embryos. While no direct overlaps were found in nuclear DNA, three CpGs exhibited opposite methylation trends—hypomethylated in oocytes but hypermethylated in embryos. In contrast, five mitochondrial DMCs overlapped between oocytes and embryos. To functionally assess the role of differentially methylated genes, we performed siRNA-mediated knockdown of two embryo DMC-associated genes: SCRIB and CERS3. Knockdown of SCRIB led to a 16.4% average decrease in blastocyst formation rate (p = 0.001), while CERS3 knockdown resulted in a 9.5% decrease (p = 0.005). These results demonstrate that maternal methionine supplementation alters both nuclear and mitochondrial DNA methylation in oocytes and embryos and that affected genes may play critical roles in early embryonic

overall_design: Whole genome bisulfite sequencing (WGBS) was performed on oocytes collected from 16 twin Polypay ewe pairs (8 methionine-treated and 8 control). These ewes were later bred to control rams, and embryos were flushed for WGBS as well.

first: Jessica

last: Townsend

first: Mehmet

last: Kizilaslan

first: Zeynep

first: Todd

last: Taylor


title: Epigenetic changes by EZH2 inhibition increase translocations in B cells with high AID activity or DNA repair deficiency [HTGTS in MEC-1]

first: Roberto

last: Chiarle

state: Ma

address: Longwood 333

institute: Boston Childrens Hospital

summary: The EZH2 histone methyltransferase inhibitors tazemetostat and valemetostat have recently received approval for clinical use in follicular lymphoma and adult T-cell leukemia/lymphoma, respectively. In follicular lymphoma, the expression of AID is responsible for increased mutational signatures and genomic instability. Because EZH2 inhibitors induce epigenetic and transcriptional changes in normal and lymphoma B cells, we studied whether these inhibitors could alter the pattern of AID-mediated chromosomal translocations. Here we show that treatment with EZH2 inhibitors did not significantly change AID expression or AID-dependent chromosomal translocation frequency when used as monotherapy in either CH12F3 mouse B cells or MEC-1 human B cells. In contrast, when combined with PI3Kδ inhibition, which enhances AID expression, EZH2 inhibition significantly increased the frequency of chromosomal translocations compared to either EZH2 or PI3Kδ inhibition alone both in mouse CH12F3 cells and human MEC-1 cells. EZH2 inhibition also further enhanced translocation formation in mouse B cells that were Ligase4 deficient. Mechanistically, EZH2 inhibition in B cells depletes the repressive histone modification H3K27me3 while concurrently enhancing the active histone modification H3K27ac, thereby selectively increasing transcriptional activity and facilitating chromosomal translocation formation in the presence of high AID activity or Ligase4 deficiency. These findings highlight the impact of drugs that induce epigenetic changes to influence chromosomal translocations and demonstrate the genetic safety of EZH2 inhibitors as monotherapy while highlighting the increased risk of genomic instability when used in cells prone to translocations, such as B cells with high AID levels or DNA-repair deficiency.

overall_design: We performed LAM-HTGTS to assess chromosomal translocation alterations following treatment with tazemetostat or valemetostat in MEC-1 cells.

first: Tao

last: Jianli

first: Luca

last: Alessandri


title: Endogenous retroviruses promote aberrant T cell differentiation in systemic lupus erythematosus via the RIG-I pathway [mouse_CD4]

summary: The dysregulated differentiation of T lymphocytes play an important role in systemic lupus erythematosus (SLE). However, the underlying mechanism remains unclear. Here, we screened for differentially expressed genes in CD4+T cells derived from WT and DDX58 KO mice by RNA-seq. And our findings reveal the roles and mechanism of RIG-I in regulating the aberrant differentiation of T cells in SLE patients.

overall_design: mRNA prifules of CD4+T cells of WT and Ddx58 KO mice


title: Distinct spatial organization governs oral mucosal immunity

first: Niki

last: Moutsopoulos

address: 9000 Rockville Pike

department: National Institute of Allergy and Infectious Diseases

summary: Immune responsiveness at barrier surfaces is tailored to the exposures of each tissue. In the oral mucosa, mechanisms by which a permeable epithelium coexists with diverse microbiota and maintains integrity during inflammatory pathology remain poorly understood. We compile a multi-omics spatial map of this exposed mucosal microenvironment and uncover remarkable immune zonation with organization that is preserved even during inflammatory disease. At steady state, we identify a dynamic epithelium at the tooth interface marked by a tonic inflammatory signature which is underlined by a layer of neutrophils and a zone of APC-lymphocyte aggregates. During disease, inflammatory zones expand and organize into immature tertiary lymphoid structures, suggesting local antibody production.  This spatial immune organization meets the demands for continuous protection in health and expands in function and complexity during disease while preserving key elements of zonation. Our work provides insights into tissue-specific wiring of immunity at a vulnerable human mucosal barrier.

overall_design: Periodontitis cells from four samples (P5, P8, P9, P16) were profiled by CITE-seq using TotalSeq-A Human Universal Cocktail panel of 154 antibodies.


title: Low Input Asay for Transposase-Accessible Chromatin Identifies Epigenetic Signatures of Liver Group 1 Innate Lymphoid Cells

city: Munich

first: Kevin

last: Schmid

state: Bavaria

address: Ismaninger Str. 22

country: Germany

institute: TU Munich

summary: Epigenetic regulation is fundamental to the development, activation, and memory formation of Group 1 innate lymphoid cells (ILCs), including NK cells and ILC1s. Studying chromatin accessibility in these cells, especially from tissues, is limited by low cell numbers. Here, we present a robust, low-input bulk ATAC-seq protocol optimized for primary NK cells and ILC1s. Our improved protocol achieves high-quality libraries with as few as 5,000 cells. Applying this protocol to liver-derived NK cells and ILC1s, we identify over 30,000 differentially accessible regions, reflecting distinct epigenetic regulation. Our findings reveal key transcription factor motifs and confirm known gene accessibility patterns, highlighting functional divergence between these cell types. This accessible protocol enables reliable epigenetic profiling of rare ILC populations, improving the study of tissue-resident immunity.

overall_design: ATAC-seq profiling of primary Group 1 ILCs from wild-type C57Bl/6J mouse liver.

first: Robin

last: Schenk

first: Gabriela

last: Wiedemann


title: Mouse bone marrow-derived mucosal mast cell model characterization

city: Toulouse

first: Eric

last: Espinosa

state: France

address: IRSD  INSERM U1220 105 Avenue de Casselardit

department: IMMUNOLOGY

country: France

institute: INSERM

summary: Mast cells (MCs) fall into two subpopulations with well-defined roles and characteristics: connective tissue mast cells (CTMCs) and mucosal mast cells (MMCs). While in vitro models of CTMCs exist in humans and mice, a reliable and relevant model of MMCs is still lacking. Contrary to the pioneer studies' one-step protocol, our two-step differentiation protocol replicates the two major stages of mast cell development to generate mouse bone marrow-derived mucosal mast cells (BM-MMCs) in vitro. Bone marrow cells were cultured in a complete Opti-MEM medium supplemented with IL-3 and SCF for 4 weeks to induce MC commitment before adding for one additional week both IL-9 and TFG-β1 to promote MMC proliferation and maturation  BM-MMCs were phenotypically and functionally compared to the previously established CTMC model (PCMCs). RNAseq analysis showed that BM-MMC exibit MMC typical characteristics and recapitulate gene expression observed in isolated MMCs. BM-MMCs, which have transcriptomic characteristics similar to MMCs, can be an effective tool for studying a significant number of viable MMCs during a 15-day period in vitro

overall_design: RNAseq profiling of  BM-MMCs (n=3)   as compared with  PCMCs (n=3)

first: Louise

last: Battut

first: Jasper

last: Kamphuis

first: Nadine

last: Serhan

first: Laurent

last: Reber

first: Nicolas

last: Cenac

first: Gilles

last: Dietrich


title: Dipeptidyl peptidase 4 restoration facilitates anti-tumor immunity in KRAS–LKB1–mutant lung cancer

city: Asahikawa

first: Hiraku

last: Yanada

country: Japan

institute: Asahikawa Medical University

summary: KRAS proto-oncogene, GTPase (KRAS)-liver kinase B1 (LKB1)-mutant (KL) non-small cell lung cancer (NSCLC), characterized by a profoundly immunosuppressive tumor microenvironment (TME), is highly resistant to immune checkpoint inhibitors (ICIs). Despite their high tumor mutation burden, KL tumors exhibit low programmed cell death ligand 1 (PD-L1) expression, reduced immune cell infiltration, and suppressed immune signaling pathways. Through systematic genomic analyses, we found that LKB1 loss suppresses dipeptidyl peptidase 4 (DPP4) expression and activity in KRAS-mutant lung cancer. Therefore, we evaluated the therapeutic potential of restoring DPP4 function to improve the immune response in KL lung cancer using patient-derived tumor samples and syngeneic mouse models. Restoration of DPP4 expression reprogrammed the TME and significantly increased immune-related gene signatures, including those involved in T cell migration and natural killer (NK) cell activation. Indeed, DPP4 restoration in three-dimensional microfluidic models enhanced NK cell chemotaxis and activity in targeting tumor spheroids. Furthermore, DPP4 restoration in syngeneic KL murine models synergized with anti-PD-1 therapy to achieve significant tumor regression. These findings suggest that LKB1 loss suppresses DPP4 expression, contributing to the immunosuppressive characteristics of the TME in KL-NSCLC cells, whereas, restoring DPP4 expression promotes NK cell recruitment, facilitates immune activation, and enhances the effects of anti-PD-1 therapy. These results highlight DPP4 as a key immune modulator and a promising therapeutic target, offering a novel strategy to overcome immune resistance and enhance immunotherapy outcomes in this challenging subset of lung cancer.

overall_design: Examination of gene expression using H2122 cells with two biological replicates. H2122-LUC cells were used as controls, and expression levels were compared with H2122-DPP4 cells.

first: Toshiyuki

last: Tenma

first: Ryohei

last: Yoshida

first: Kiichi

last: Nitanai

first: Toshihiro

last: Nagato

first: Kyohei

last: Oyama

first: Nobunari

last: Sasaki

first: Mizuki

last: Homme

first: Chie

last: Mori

first: Takayuki

last: Ohkuri

first: Yusuke

last: Ono

first: Shoichiro

last: Tange

first: Yoshinori

last: Minami

first: Hiroya

last: Kobayashi

last: Mizukami

first: David

last: Barbie

first: Shunsuke

last: Kitajima

first: Takaaki


title: Connecting cilium, stress response and proteostasis abnormalities inform variant and therapy assessment in RPGRIP1 retinal organoid [scRNA-seq]

city: Westmead

first: Nader

last: Aryamanesh

state: NSW

address: 214 Hawkesbury Rd

department: Bioinformatics

country: Australia

institute: CMRI

summary: RPGRIP1 encodes a connecting cilium (CC) protein essential for normal photoreceptor cell development and maintenance. Damaging variants in RPGRIP1 cause severe inherited retinal degeneration (IRD) and currently incurable vision loss, with mice studies showing promising preclinical gene augmentation therapy results. Almost one-half of variants in RPGRIP1 in the ClinVar database are variants of uncertain significance (VUS), hindering genetic diagnosis for affected individuals, and hence access to clinical trials of novel therapies and other management options. Here, we use human induced pluripotent stem cell (iPSC)-derived retinal organoids to model RPGRIP1-associated IRD, detecting biomarkers of disease including CC interactome dysfunction, stress response and proteostasis abnormalities. In parallel, utilising these novel disease biomarkers we demonstrate the pathogenicity of a missense VUS, RPGRIP1 c.2108 T>C p.(Ile703Thr). Finally, RPGRIP1 gene augmentation therapy rescued disease phenotypes, further supporting the utility of these biomarkers of RPGRIP1-LCA disease for reclassifying VUS and testing response to therapy.

overall_design: scRNA-seq profiling of wild type (Control-1) and RPGRIP1 variant (LCA-1 and MS-VUS) retinal organoids performed with replicates.

first: To Ha

last: Loi

first: Anson

last: Cheng

first: Hani

last: Kim

first: Milan

last: Fernando

first: Benjamin

last: Nash

first: John

middle: R

last: Grigg

first: Pengyi

last: Yang

first: Anai

last: Gonzalez Cordero

first: Robyn

middle: V

last: Jamieson


title: Connecting cilium, stress response and proteostasis abnormalities inform variant and therapy assessment in RPGRIP1 retinal organoids [RNA-Seq]

overall_design: RNA seq profiling of wild type (Control-1) and RPGRIP1 variant (LCA-1 and MS-VUS)  retinal organoids performed in triplicate.


title: Direct recruitment of TONSOKU by the N-terminal tails of histone variants H2A.X and  H2A.W coordinates DNA repair

city: hangzhou

first: li

last: qi

address: xihu

institute: zhejiang university

summary: Histone variants play crucial roles in DNA replication and DNA damage repair. Plant TONSOKU (TSK) specifically recognizes unmethylated histone H3.1 to bind post-replicative chromatin for DNA repair, but the precise mechanisms of TSK recruitment to double-stranded break (DSB) sites remain unclear. Here, we discovered that the leucine-rich repeats (LRR) domain of TSK recognizes the N-terminal tail of the histone variant H2A.X, the phosphorylated form of which (also known as γ-H2A.X) is a critical marker of DSBs at both replication forks and euchromatin. Additionally, the LRR domain of TSK recognizes the N-terminal tail of the plant-specific histone H2A variant H2A.W, which is essential for DSB repair in heterochromatin, but not H2A.Z, which primarily regulates gene expression. A motif containing multiple basic and acidic residues (BAR motif), unique to H2A.Z, prevents TSK from binding. Notably, mutations that abolish TSK-H2A.X/W binding interfere with TSK-mediated DNA repair and root development. Genetic analyses indicate that H2A.X and H2A.W are redundantly required for DNA damage repair in vivo, with the N-terminus of H2A.X being particularly indispensable. This study reveals a novel H2A.X/W-dependent mechanism for recruiting the DNA repair protein TSK, highlighting the critical role of their previously uncharacterized N-terminal tails in plants.

overall_design: RNA-seq profiling of Col, h2a.x-1, h2a.w-2, tsk and h2a.x/w-2 in 7-day-old seedling

first: Qi

last: Li

first: Jie

last: Dong


title: High throughput transcriptomics analysis of ovine mammary epithelial cells stimulated with Staphylococcus aureus in vitro

city: Talas

first: Ghulam Asghar

last: Sajid

state: Kayseri

address: Yenidoğan, Turhan Baytop Sokak No:1

department: Animal Science Department

country: Turkey

institute: Erciyes University

summary: In sheep, the innate immune response of mammary epithelial cells (MECs) plays a central role in combating mastitis, yet our understanding of their resistance mechanisms remains limited. This study aimed to elucidate the gene expression profiles of ovine MECs following in vitro stimulation with Staphylococcus aureus (S. aureus) using RNA-Seq technology. Bioinformatics analysis identified a total of 175 differentially expressed genes (DEGs), including 172 up-regulated and 3 down-regulated genes in the stimulated group compared to the non-stimulated control group. Gene ontology annotation and functional pathway analysis indicated that these DEGs are primarily involved in ribosomal functions, which are essential for protein synthesis and first target of pathogens, as well as in immune response dysregulations, infection, phagocytosis, and bacterial invasion of epithelial cells. Validation via quantitative real-time PCR (qRT-PCR) confirmed the RNA-Seq results. These findings significantly contribute to the understanding of how ovine MECs respond to S. aureus stimulation, providing a foundation for further research, particularly regarding the immune defense mechanisms, strategies and implications in dairy industry.

overall_design: Mammary mammary parenchyma tissue samples were collected from healthy ewes. Mammary epithelial cell culture was prepared and stimulation model was develop as 3 tissue plates were treated, keep 3 plate as non-stimulated control. Total RNA was isolated and cDNA libraries were prepared for RNA sequencing. A standard bioinformatics pipeline was developed to extract the results.

first: Saif Adil Abbood

last: Al-Janabi

first: Sidra

last: Zeb

first: Muhammad Jasim

last: Uddin

first: Mehmet Ulas

last: Cinar


title: HERV-K10 as a mediator of immune modulation in hepatitis infections

city: Neuherberg

first: Salih

last: Özer

address: Ingolstädter Landstraße 1

department: Institute of Virology

institute: Helmholtz Munich

summary: The human genome contains ~8% of endogenous retroviruses (HERVs), which are implicated in diseases such as cancer and autoimmune disorders. Among these, HERV-K10 has attracted attention for its potential role in immune modulation and viral infections. This study investigates HERV-K10 expression in hepatitis virus infections, focusing on its impact on host gene expression and immune responses. We analysed HERV-K10 in PBMCs from patients chronically infected with hepatitis C virus (HCV) and in HBV-infected liver cell models. Our results show a significant upregulation of HERV-K10 in HBV-infected HepG2-NTCP cells, HCV-infected PBMCs and a trend in HBV-infected primary hepatocytes. HERV-K10 activation was specific to hepatitis infection, as no effect was seen with HBV entry inhibitors or adenovirus 5. RNA sequencing of HBV-infected HepG2-NTCP cells revealed distinct clustering based on HERV expression profiles, including HERV-K10 encoding the MAG1 domain, an immune response target. To investigate the potential immunomodulatory role of HERV-K10 MAG1, we vaccinated mice with the MAG1 peptide, which resulted in activation of CD4+ and CD8+ T-cell responses and higher levels of MAG1-specific antibodies. Furthermore, chronic HBV patients exhibited an immune response to MAG1 characterized by elevated levels of IL-6 and IL-1β cytokines. Taken together, our data suggest that HERV-K10 plays an important role in immune modulation during viral hepatitis infection and may contribute to the pathogenesis of autoimmune diseases.

overall_design: RNA-seq profiling of HBV-wt infected HepG2-NTCP cells two days post infection

first: Romano

last: Strobelt

first: Anna

last: Kosinska

first: Goar

last: Frishman

first: Lisa

last: Pleninger

first: Wen

last: Liang

first: Andreas

last: Ruepp

first: Jochen

last: Wettengel

first: Nina

last: Körber

first: Tanja

last: Bauer

first: Michelle

last: Vincendeau

first: Ulrike

last: Protzer


title: Ribo-lite, a method defining paired transcriptome (RNA-seq) and translatome (Ribo-seq) at low input on AML cells from mice treated with 5+3 or vehicle and fed standard amino acids (AA) or branched chain amino acid (BCAA) free diets. [II]

first: Christina

last: Mayerhofer

address: 185 Cambridge St.

department: Center for Regenerative Medicine

institute: MGH, Simches building

summary: Acute Myeloid Leukemia (AML) commonly relapses after initial chemotherapy response. We assessed metabolic adaptations in chemoresistant cells in vivo before overt relapse, identifying altered branched-chain amino acid (BCAA) levels in patient-derived xenografts (PDX) and immunophenotypically identified leukemia stem cells from AML patients. Notably, this was associated with increased BCAA transporter expression with low BCAA catabolism. Restricting of BCAAs further reduced chemoresistant AML cells but relapse still occurred. Among the persisting cells we found an unexpected increase in protein production. This was accompanied by elevated translation of 2-oxoglutarate-and-iron-dependent oxygenase 1 (OGFOD1), a known ribosomal dioxygenase that adjusts the fidelity of tRNA anticodon pairing with coding mRNA1–3 and upregulates protein synthesis in AML driving disease aggressiveness. Inhibiting OGFOD1 impaired translation processing, decreased protein synthesis and improved animal survival even with chemoresistant AML through regulation of protein synthesis. Leukemic cells can therefore persist despite the stress of chemotherapy and nutrient deprivation through adaptive control of translation while sparing normal hematopoiesis. Targeting OGFOD1 may offer a distinctive, translation modifying means of reducing the chemopersisting cells that drive relapse.

overall_design: Leukemic mice with either control or Ogfod1 KO were fed branched chain amino acid (BCAA) free or standard amino acid (AA) diet to determine effects in gene expression and translated mRNAs using Riboseq, following 5+3 (cytarabine and doxorubicin) chemotherapy

first: Ernst


title: BRD9 KO RNA seq (primary cells)-Highly discriminative globin gene activation by the non-canonical BAF chromatin remodeling complex

city: Philadelphia

first: Eugene

last: Khandros

address: 3615 Civic Center Blvd.

institute: The Children's Hospital of Philadelphia

summary: The regulation of the switch from fetal (HBG) to adult (HBB and HBD) beta-globin gene expression has served as a paradigm for clinically relevant developmental transcriptional control. Mechanistic studies of this switch have focused overwhelmingly on HBG repressors with comparatively little attention paid to potential HBG activators. We found that in adult type HUDEP2 erythroid cells, the ATP-dependent chromatin remodeler BRG1 preferentially activates the HBG genes as well as the minor adult HBD gene. BRG1 is a core catalytic subunit of three BAF complexes, canonical BAF, polybromo BAF, and non-canonical BAF (ncBAF) that regulate chromatin accessibility in distinct gene- and cell-type contexts. Using comprehensive CRISPR mediated dissection of these complexes in HUDEP2 and primary adult erythroid cells, we pinpointed the HBG and HBD selective activation function of BRG1 to the ncBAF complex. Loss of the ncBAF complex subunits BRD9 and BAF60A preferentially decreased HBG and HBD transcription while accelerating erythroid differentiation and hemoglobinization. Pharmacologic depletion of BRD9 acutely decreased chromatin accessibility at the HBD and HBG promoters and reduced their transcriptional activities. This surprising demonstration of BAF complex selectivity within a multi-gene cluster extends our understanding of BAF complex functions and identifies a potential target for therapeutic manipulation of beta-like globin genes.

overall_design: We performed RNA seq analysis in 3 healthy donor erythroid cells derived from CD34+ HSPC after BRD9 knockout. Total RNA was isolated form erythoid cells after 10 days of erythroid differentiation targetted with CRISPR control sgRNA (AAVS1 targetting) or BRD9 targetting. Transcriptome changes were analysed by bulk RNA-seq after ribosomal RNA removal.

first: Abhirup

last: Bagchi

first: Sindhu

last: Billakanti

first: Vanessa

last: Shehu

first: Kathryn

last: Farrell

first: Sarah

last: Cotteta

first: Bailey

last: Godfrey

first: Micaela

last: Alpert

first: Jacob

last: Zeranski

first: Krista

last: Budinich

first: Zhuangzhuang

last: Geng

first: Osheiza

last: Abdulmalik

first: Belinda

last: Giardine

first: Cheryl

last: Keller

first: Ross

last: Hardison

first: Junwei

last: Shi


title: Polysome based Translatome profiling in AML cell lines with OGFOD1 overexpression

first: Christina

last: Mayerhofer

summary: Acute Myeloid Leukemia (AML) commonly relapses after initial chemotherapy response. We assessed metabolic adaptations in a minimal-residual disease setting of chemoresistant cells in vivo before overt relapse, identifying elevated branched-chain amino acid (BCAA) levels in patient-derived xenografts (PDX) and immunophenotypically identified leukemia stem cells from AML patients. Notably, this was associated with increased BCAA transporter expression without indications for higher BCAA catabolism. Restricting of BCAAs further reduced chemoresistant AML cells but relapse still occurred. Among the persisting cells we found an unexpected Acute Myeloid Leukemia (AML) commonly relapses after initial chemotherapy response. We assessed metabolic adaptations in chemoresistant cells in vivo before overt relapse, identifying altered branched-chain amino acid (BCAA) levels in patient-derived xenografts (PDX) and immunophenotypically identified leukemia stem cells from AML patients. Notably, this was associated with increased BCAA transporter expression with low BCAA catabolism. Restricting of BCAAs further reduced chemoresistant AML cells but relapse still occurred. Among the persisting cells we found an unexpected increase in protein production. This was accompanied by elevated translation of 2-oxoglutarate-and-iron-dependent oxygenase 1 (OGFOD1), a known ribosomal dioxygenase that adjusts the fidelity of tRNA anticodon pairing with coding mRNA1–3 and upregulates protein synthesis in AML driving disease aggressiveness. Inhibiting OGFOD1 impaired translation processing, decreased protein synthesis and improved animal survival even with chemoresistant AML through regulation of protein synthesis. Leukemic cells can therefore persist despite the stress of chemotherapy and nutrient deprivation through adaptive control of translation while sparing normal hematopoiesis. Targeting OGFOD1 may offer a distinctive, translation modifying means of reducing the chemopersisting cells that drive relapse.

overall_design: Human AML cell lines with OGFOD1 overexpression or control underwent polysome profiling and RNA sequencing.

first: Ernst


title: BRD9 CFT8634 ATAC Seq (primary cells)-Highly discriminative globin gene activation by the non-canonical BAF chromatin remodeling complex

first: Eugene

last: Khandros

overall_design: We performed ATAC-seq analysis in 3 healthy donor erythroid cells derived from CD34+ HSPC after acute CFT8634 treatment.

first: Abhirup


title: Acute Nutritional Ketosis During Early Recovery from Exercise Does Not Affect Skeletal Muscle Transcriptomic Response

city: Liverpool

first: Daniel

last: Owens

address: Byrom Street

department: Research Institute for Sport and Exercise Science

country: United Kingdom

institute: Liverpool John Moores University

summary: Purpose: Recent research suggests that Ketone monoester (KME) supplementation may enhance skeletal muscle adaptation to exercise, possibly modulated be an increased secretion of erythropoietin (EPO) and increased muscle glycogen resynthesis, but the precise molecular pathways driving the response remain uncertain. Therefore, we aimed at characterising the modulation of skeletal muscle by KME through a genome-wide characterisation of the transcriptome early post-exercise. Methods: Following a randomized, double-blind, crossover design, recreationally active men [n= 9; age: 26 ± 5 (means ± SD); height: 1.80 ± 0.07 m; body mass: 80 ± 9 kg; V̇O2max: 47 ± 4 mL·kg-1·min-1]) completed two experimental trials where they ingested either 1.25 g/kg of KME (>96% (R)-3-hydroxybutyl (R)-3-hydroxybutyrate) or a taste-matched placebo (PLA) drink around exercise (90-min cycling  at 60% of VO2max) in standardised conditions. . Venous blood samples were taken throughout baseline, exercise and recovery, and skeletal muscle biopsies were taken at baseline and 3-h post exercise. Results: KME intake elevated serum ßHB concentrations post-exercise to 3.3 ± 0.5 mM and remained elevated levels during recovery (range: ~4.0–4.7 mM). Remarkably, there was no marked upregulation or downregulation of differentially expressed genes at 3-h post-exercise with KME supplementation compared to PLA. Although, baseline serum EPO levels were higher in PLA (12.8 ± 3.9 IU/L) compared to KME (10.1 ± 5.7 IU/L) and remained slightly higher in PLA throughout recovery, there was not significant difference between conditions. Pre- and post- exercise muscle glycogen concentrate were similar between conditions (KME PRE: 501 ± 118 mmol/kg, KME POST, 284 ± 117 mmol/kg, PLA PRE: 492 ± 102 mmol/kg, PLA POST: 282 ± 184 mmol/kg). Conclusions: Our findings indicate that despite a sustained post-exercise increase in serum ßHB concentrations, KME supplementation did not induce transcriptional changes in skeletal muscle. Additionally, in contrast to previous studies, KME did not affect glycogen concentrations or serum EPO levels during the first 3-hours post-exercise.

overall_design: Our aims were to perform an exploratory assessment of post-exercise skeletal muscle transcriptome response, to replicate findings on the effect of KME on EPO, and skeletal muscle glycogen during recovery from exercise. We hypothesized that KME would have a marked modulatory effect on skeletal muscle transcriptome and enhance EPO concentrations along with increased muscle glycogen resynthesis. This randomized, double-blind, placebo-controlled, crossover design study involved two experimental sessions involving two experimental visits, separated by 7-14 days. Each experimental sessions compromised a 90 min cycling session at 60% V̇O2max. Subjects received either 1.25 g·kg-1 (range: ~85 – 110 g) of KME [>96% (R)-3-hydroxybutyl (R)-3-hydroxybutyrate] or a volume and taste-matched placebo (PLA) drink. Both drinks were spread in four dosages (0.5 g·kg-1, 0.25 g·kg-1, 0.25 g·kg-1, 0.25 g·kg-1) to be ingested i) at min 45 of exercise ii) immediately after exercise completion iii) 1-h post-exercise iv) 2-h post exercise. On trial days, participants arrived at the laboratory on the morning (0730 to 0830 h) and venous blood samples were drawn at i) ~30 min pre-exercise ii) immediately after exercise completion iii) 60-min post-exercise iv) 120-min post exercise v) 180-min post exercise. Skeletal muscle biopsies were obtained from the vastus lateralis of the quadriceps ~60-min pre-exercise and 180-min post-exercise. Subjects received a standardized breakfast containing 1 g·kg-1 carbohydrate, 0.1 g·kg-1 protein, 0.03 g·kg-1 fat exactly 30 min pre-exercise. After exercise completion, participants consumed at 30-min post-exercise a recovery drink containing 1 g·kg-1 carbohydrate, 0.4 g·kg-1 protein. During exercise finger-prick blood samples were collected to measure lactate concentrations (Biosen C-Line, EKF Diagnostics, Cardiff, UK), respiratory gas exchange (VyntusTM CPX metabolic cart, Vyaire, Mettawa, Illinois, USA), heart rate monitor (H10 Polar, Polar Electro Oy, Kempele, Finland), and rating of perceived exertion (RPE, 6-20 scale, ((Borg, 1982)) were also recorded on the last 3-min of every 15-min.

first: Erick

middle: L

last: Mosquera

first: Julien

last: Louis

first: Jason

last: Edwards

first: Jamie

last: Pugh

first: Mark

last: Viggars

first: Jose

last: Areta


title: MYC-FACT Interactions Drive Z-DNA Formation to Modulate Transcriptional Dynamics [RNA-seq CC & CL]

city: Hangzhou

first: Dong

last: Fang

state: Zhejiang

address: 866 Yuhangtang Road

department: Life Sciences Institute

institute: Zhejiang University

summary: DNA topology is critical for regulating transcription and maintaining cellular homeostasis. Z-DNA is a left-handed DNA helix in regions with high transcriptional activity. Its physiological function remains poorly understood. Here, we demonstrate that oncoprotein MYC induces the formation of Z-DNA by recruiting the chromatin remodeler FACT, independent of RNA Polymerase II activity. FACT facilitates Z-DNA formation by remodeling H2A/H2B dimers within intact nucleosomes. Additionally, the phosphorylation of FACT regulates its liquid-liquid phase separation, promoting its efficient recruitment to chromatin by MYC. Through a genome-wide analysis and characterization of engineered Z-DNA promoters, we found that Z-DNA directly facilitates the loading of RNA Polymerase II, thereby promoting transcriptional activity. This study elucidates the molecular mechanisms of Z-DNA dynamics and emphasizes its functional importance in transcriptional regulation, providing insights into the role of non-B-DNA structures in chromatin biology and MYC-driven cancer.

overall_design: RNAseq in U2OS cells (#1 CC, #1 CL, #2 CC, #2 CL, #3 CC, #3 CL, #4 CC, #4 CL)


title: MYC-FACT Interactions Drive Z-DNA Formation to Modulate Transcriptional Dynamics [ChIP-seq]

overall_design: ChIP-seq for Z-DNA in U2OS cells（WT，Zαβ，MYC，MYC + Zαβ）


title: MYC-FACT Interactions Drive Z-DNA Formation to Modulate Transcriptional Dynamics

overall_design: TTseq in U2OS cells（WT，Zαβ，MYC，MYC + Zαβ）


title: MYC-FACT Interactions Drive Z-DNA Formation to Modulate Transcriptional Dynamics [CUT&TAG]

overall_design: CUT&Tag for protein MYC, RNAP II and SSRP1 in U2OS cells（WT，Zαβ，MYC，MYC + Zαβ）


title: MYC-FACT Interactions Drive Z-DNA Formation to Modulate Transcriptional Dynamics [MNase-seq]

overall_design: Mnse-seq in U2OS cells（WT，Zαβ，MYC，MYC + Zαβ）


title: Early life starvation and Hedgehog-related signaling activate innate immunity downstream of daf-18/PTEN and lin-35/Rb causing developmental pathology in adult C. elegans

city: Durham

first: Ivan

last: Falsztyn

state: NC

address: French Science Center

department: Biology

institute: Duke University

summary: Early life experiences such as malnutrition can affect development and adult disease risk, but the molecular basis of such protracted effects is poorly understood. In the nematode C. elegans, extended starvation during the first larval stage causes the development of germline tumors and other abnormalities in the adult gonad, limiting reproductive success. Insulin/IGF signaling (IIS) acts through WNT signaling and lipid metabolism to promote starvation-induced gonad abnormalities, but IIS-independent modifiers have not been identified. We show that the tumor suppressors daf-18/PTEN and lin-35/Rb act independently of IIS to suppress starvation-induced abnormalities. We found that lin-35/Rb antagonizes activity of the Hedgehog (Hh) signaling homologs ptr-23/PTCH-related, wrt-1/Hh-like, wrt-10/Hh-like, and tra-1/GLI, which promote starvation-induced abnormalities. These Hh-related genes transcriptionally activate several genes associated with innate immunity in adults, which also promote starvation-induced gonad abnormalities. Surprisingly, we found that in addition to causing developmental abnormalities, early-life starvation induces an innate immune response later in life, leading to increased resistance to multiple bacterial pathogens. This work identifies a critical tumor-suppressor function of daf-18/PTEN independent of IIS, and it defines a regulatory network, including lin-35/Rb, Hh-related signaling, and the innate immunity pathway, that affects development of tumors and other developmental abnormalities resulting from early life starvation. By revealing that early-life starvation increases immunity later in life, this work suggests a fitness tradeoff between pathogen resistance and developmental robustness.

overall_design: Bulk RNA-seq profiling of early adult wild type (N2) Caenorhabditis elegans. Individuals experienced 8 days of L1 arrest then were recovered on the indicated food source (RNAi treatment)

middle: B

first: Jingxian

last: Chen

first: Rojin

last: Chitrakar

first: James

last: Jordan

first: Ryan

last: Baugh


title: RNA-seq of macrophages treated with or without MLN4924

first: Jiao

last: Fan

address: Fuxing road

institute: 301 hospital

summary: MLN4924, a neddylation inhibitor with potential in cancer treatment, has anti - inflammatory effects. As atherosclerosis is an inflammatory disease related to autophagy and cell senescence, this study explored MLN4924's role in it. Focusing on foam cell formation, a key step in atherosclerosis, results showed MLN4924 downregulated scavenger receptors like CD36, and upregulated cholesterol efflux gene ABCA1. It also inhibited macrophage senescence and SASP - related factors, possibly via enhanced autophagy. Analyses revealed its impact on macrophage transcriptome and metabolism, with genes enriched in the PI3K - AKT pathway. MLN4924 inhibited AKT phosphorylation, regulated gene expression such as ABCA1, and modulated metabolite production. These suggest its potential as an anti - atherosclerotic agent for further research.

overall_design: RNA sequencing analysis of macrophages treated with ox-LDL alone and macrophages treated with ox-LDL+MLN4924 at the 48th hour of transformation into foam cells

first: Lei

last: Xing

first: Yu

last: Bu

first: Jingqi

first: Jing

last: Liu

first: Minghua

last: Zhang


title: MYC-FACT Interactions Drive Z-DNA Formation to Modulate Transcriptional Dynamics [RNA-seq]

overall_design: RNAseq in U2OS cells (WT，Zαβ，MYC，MYC + Zαβ)


title: Transcriptomic and Epigenetic Signatures of Tomato Fruit After Postharvest UV-C Irradiation Are Associated with the Maintenance of Fruit Quality

first: Lili

last: Ma

address: 50 Changhua Road, Haidian District

institute: Beijing Academy of Agriculture and Forestry Science

summary: Exposure to UV-C irradiance effectively maintains fruit quality, however the mechanisms by which it ensures tomato postharvest quality remain unclear. We conducted a comprehensive analysis of the transcriptome, volatilome, DNA methylome, and RNA methylome of tomato fruit in order to construct a UV-C regulatory network. UV-C was found to induce the expression of hormones-related, flavonoid, and resistance genes, to delay senescence, and to promote the expression of sugar and acid metabolism genes, leading to the synthesis of soluble sugars and degradation of citric acid in a manner that maintains fruit taste characteristics. Volatilome results demonstrated that UV-C enhanced the accumulation of 3-methyl-2-butenal, β-damascenone, and 6-methyl-5-heptene-2-one by regulating genes associated with the metabolism of these volatile organic compounds. Additionally, UV-C contributed to the retention of accessible chromatin regions (ACRs), with genes associated with plant hormones showing variation in expression corresponding to chromatin accessibility. UV-C also inhibited the decrease in DNA methylation during ripening, with dynamic changes in DNA methylation and RNA methylation of genes related to hormones, flavonoids, texture, and resistance, ultimately delaying fruit senescence. This study is the first to comprehensively elucidate the regulatory role of UV-C on tomato postharvest ripening and as such provides valuable insights into epigenetic modifications which could represent important tools for future quality-based breeding programs.

overall_design: ‘Original No. 1’ fresh tomatoes at the firm red ripe maturity stage were harvested at the “Hehe Yuan” farm (Miyun District, Beijing, China) and transported to the laboratory within 1 h, and intact fruit were randomly divided into two groups of 200. Tomatoes were put into polyethylene boxes (crisper). One group was treated with UV-C light (TUV 30 W, Philips, Netherlands) at a dose of 4 kJ/m2, and the irradiation dose was measured by ultraviolet irradiation meter (LS125, Linshang, Shenzhen), while the other group was not treated in order to serve as a control. After irradiation, it is sealed with plastic wrap and stored at 20 ℃ in darkness. On day 8, the control group fruit had lost commercial value, so the fruit of 0 d (CK-0d) and 8 d (CK-8d and UV-C-8d) were selected for omics analysis (Transcriptome, volatile metabolome, DNA methylation, and RNA methylation). Each group randomly selected 10-12 fruit. The middle pericarp tissue tomatoes (containing epicarps, without seeds) were cut into pieces and mixed well, then immediately frozen with liquid nitrogen and stored in a refrigerator at -80°C. All experiments were parallel three times.

first: Jiejie

last: Tao

first: Yanyan

last: Zheng

first: Jinhua

last: Zuo

first: Yunxiang

last: Wang

first: Alisdair

last: R. Fernie

first: Chunmei

last: Bai


title: Noncanonical PRC2 chromatin association reveals a key role of EZHIP in intergenerational epigenetic memory transition

first: Feng

last: Kong

address: Qinghuayuan Street

institute: Tsinghua University

summary: Epigenetic memory undergoes drastic reprogramming in intergenerational gamete-to-embryo transition. H3K27me3, a histone mark catalyzed by PRC2 (Polycomb Repressive Complex 2), is an evolutionally conserved histone mark that plays essential roles in stably repressing developmental genes to maintain cell-type-specific programs.  In mice, H3K27me3-mediated epigenetic memory, however, undergoes both resetting at classic Polycomb targets and inheritance of broad H3K27me3 domains that mediate noncanonical imprinting (noncanonical H3K27me3 domains) during early embryogenesis. How such erasure and inheritance are achieved simultaneously remains elusive. Here, through temporal profiling of chromatin occupancy, we show that PRC2 associates with H3K27me3 at both Polycomb targets and noncanonical domains in growing oocytes. PRC2 then pre-disassembles and dissociates from chromatin in mature oocytes, before H3K27me3 erasure at Polycomb targets after fertilization. PRC2 reappears at Polycomb targets in blastocysts when H3K27me3 initiates restoration. By contrast, noncanonical H3K27me3 domains persist to blastocysts, where PRC2 rapidly rebinds by the end of 1-cell embryos until blastocysts. Mechanistically, we found that such noncanonical PRC2 action is regulated by EZHIP, a PRC2 inhibitory factor. By doing so, EZHIP plays a key role in both H3K27me3 erasure and intergenerational inheritance, as well as H3K27me3 re-establishment. Ezhip is highly translated around fertilization, which strongly restricts PRC2 activity, leading to gradual decrease of H3K27me3. Upon knockout of maternal Ezhip, PRC2 becomes hyperactive, binds promiscuously in the genome and deposits widespread H3K27me3. Consequently, H3K27me3 allelic bias and noncanonical imprinting, including imprinted X chromosome inactivation, are impaired. Unexpectedly, the widespread ectopic H3K27me3 paradoxically causes de-repression of H3K27me3 targets, possibly due to dilution of H3K27me3 effectors. Loss of Ezhip further attenuates H3K27me3 restoration at Polycomb targets during peri-implantation, and leads to gene dysregulation and sub-lethality after implantation. Taken together, we uncover PRC2 noncanonical chromatin association during the parental-to-embryonic transition and identify Ezhip as a crucial regulator for both erasure and inheritance of intergenerational epigenetic memories.

overall_design: To investigate PRC2 regulation during mouse oogenesis, Ribo-lite was used to examine the translatome in wild-type D14 oocytes.

first: Yitian

last: Zeng

first: Wei

last: Xie


title: Noncanonical PRC2 chromatin association reveals a key role of EZHIP in intergenerational epigenetic memory transition [ChIP-Seq, Cut & Run]

overall_design: To investigate PRC2 regulation on chromatin during mouse oogenesis and early embryogenesis, we generated temporal EZH2/EED chromatin association datasets across 7 developmental stages with CUT&RUN. To explore the function of EZHIP in H3K27me3 intergenerational transition, we deleted Ezhip in mouse oocytes or embryos. CUT&RUN was used to examine H3K27me3, H2Aub, EZH2, and EED in wild-type and Ezhip KO oocytes or mKO embryos. At least two replicates were involved for each sample.


title: Age deceleration and reversal gene patterns in dauer diapause

city: Cologne

first: Walter

last: Sandt

address: Joseph-Stelzmann-Str. 26

institute: CECAD

summary: The aging process is characterized by a general decrease in physical functionality and poses the biggest risk factor for a variety of diseases such as cancer, cardiovascular diseases, and neurodegenerative disorders among others. Understanding the naturally evolved mechanisms that could slow aging and rejuvenate an animal could provide important concepts as to how aging could be slowed and functionality restored. The C. elegans dauer state is a robust and long-lived alternative developmental state that upon dauer exit has a normal adult lifespan with fully retained fecundity. In order to understand how dauer longevity and rejuvenation following dauer exit is mediated, we here characterized the gene expression changes during dauer and upon exit. We assessed how biological age, as determined via BiT Age, a transcriptional aging clock, is affected during dauer and upon dauer exit. During the dauer stage, we measured a decelerated increase in age compared to the chronological age and an age reversal following dauer exit. We characterized the underlying gene expression patterns that indicate major metabolic shifts as well as enhanced biomolecular degradation. Our data provide new insights into the underlying mechanisms of naturally occurring age deceleration and rejuvenation.

overall_design: RNA-sequencing of C.elegans for 1, 4, 15 and 30 days in dauer diapuse and 6h and 24h post-exit on each of the 4 days, plus L3 as a reference.

first: Khrystyna

last: Totska

first: João C. V. V.

last: Barata

first: David

last: Meyer

first: Bjoern

last: Schumacher


title: Endogenous retroviruses promote aberrant T cell differentiation in systemic lupus erythematosus via the RIG-I pathway [whole transcriptomics]

city: Nanjing

first: Xiaoli

last: Min

address: No.4 Jiang Wang Temple Street

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

summary: The dysregulated differentiation of T lymphocytes play an important role in systemic lupus erythematosus (SLE). However, the underlying mechanism remains unclear. Here, we screened and identified many overexpressed HERVs in CD4+ T cells from SLE patients by whole transcriptome resequencing and found that some HERVs formed dsRNAs to induce the activation of the RIG-I signaling pathway. And our findings reveal the roles and mechanism of HERVs and RIG-I in regulating the aberrant differentiation of T cells in SLE patients.

overall_design: mRNA prifules of CD4+T cells of SLE and HCs

first: Yaqian

last: Yu

first: Zhi

last: Hu


title: Endogenous retroviruses promote aberrant T cell differentiation in systemic lupus erythematosus via the RIG-I pathway [naïve]

summary: The dysregulated differentiation of T lymphocytes play an important role in systemic lupus erythematosus (SLE). However, the underlying mechanism remains unclear. Our findings reveal many overexpressed HERVs in CD4+ T cells from SLE patients. To investigate the role of dsRNA/RIG-I pathway in CD4+ T cell differentiation, RNA-seq was performed in CD3 and CD28 antibodies activated naive CD4+ T cells transfected with poly(I:C). The expression profiles of genes related to the effector T cell differentiation reveal that dsRNAs induced the activation of the RIG-I signaling pathway plays important roles in aberrant differentiation of T cells.

overall_design: mRNA prifules of naive CD4+T cells of HCs with poly(I:C) treatment