title: Sexually Dimorphic Sail Feathers in the Mandarin Duck: A Model for Lifelong Developmental Modulation [RNA-seq]

first: Ya-Chen

last: Liang

overall_design: We performed bulk RNA-seq to profile gene expression in regenerating Mandarin duck (Aix galericulata) sail feather follicles during early growth in the breeding season (“in season”). Feathers were dissected to separate pulp (mesenchymal) and epithelium (epidermal) compartments from the lateral blue and medial brown vanes of male sail feathers. In total, eight libraries were generated: blue sail feather pulp (Chuong-152, Rep 1; Chuong-251, Rep 2), brown sail feather pulp (Chuong-151, Rep 1; Chuong-248, Rep 2), blue sail feather epithelium (Chuong-150, Rep 1; Chuong-250, Rep 2), and brown sail feather epithelium (Chuong-149, Rep 1; Chuong-247, Rep 2). Each sample represents an independent biological replicate, enabling differential expression analysis between tissue compartments (pulp vs. epithelium) and feather vane identities (blue vs. brown). This design allowed us to identify gene expression programs associated with asymmetry and pigmentation in sexually dimorphic sail feather morphogenesis.

first: Pin-Chi

last: Tang

first: Hsu-Chen

last: Cheng

first: Gee-Way

last: Lin

first: Yung-Chih

last: Lai

first: Ping

last: Wu

first: Tzu-Chiao

first: Chen Siang

last: Ng

first: Isheng

last: Tsai

first: Ang

last: Li

first: Wen Hsiung

first: Chih-Feng

last: Chen

first: Cheng-Ming

last: Chuong

first: Randall

last: Widelitz


title: The regulatory code of injury responsive enhancers enables precision cell state targeting in the CNS [multiome]

status: Public on Oct 15 2025

city: Stockholm

first: Margherita

last: Zamboni

address: Solnavägen 9

department: Cell and Molecular Biology

country: Sweden

institute: Karolinska Institutet

type: Expression profiling by high throughput sequencing

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Enhancer elements in the genome direct cell type-specific gene expression programs. In response to injury, cells undergo dynamic changes in gene expression that drive adaptation and initiate tissue repair. Here, we investigate how injury-induced gene expression programs are encoded within enhancer elements in the mammalian CNS. Leveraging single-nucleus transcriptomics and chromatin accessibility profiling, we identify thousands of injury-induced, cell type-specific enhancers in the mouse spinal cord. These enhancers are more abundant in glial cells and retain cell type specificity, even when regulating shared wound response genes. By modeling glial injury-responsive enhancers using deep learning, we reveal that their architecture encodes cell type specificity by integrating generic stimulus response elements with cell identity programs. Finally, through in vivo enhancer screening, we demonstrate that injury-responsive enhancers can selectively target reactive astrocytes bordering lesion sites across the CNS using therapeutically relevant gene delivery vectors. Our decoding of the principles of injury-responsive enhancers enables the design of sequences that can be programmed to therapeutically target disease related cell states.

overall_design: Single nuclei from the mouse spinal cord were processed with a multiomic approach, sampling transcriptomics and chromatin accessibility information. We collected tissue from uninjured mice, as well as mice subjected to a spinal cord injury (ie., contusion, at 1, 3, 7, and 28 dpi). Nuclei were either sampled unbiased or subjected to FAC sorting to enrich for specific populations (e.g., astrocytes, ependymal cells, immune cells and perivascular cells). Each experimental condition encompassed 1-2 animals per sex.

first: Enric

last: Llorens Bobadilla


title: The regulatory code of injury responsive enhancers enables precision cell state targeting in the CNS [CUT&Tag]

overall_design: CUT&Tag for Sox9 was performed on spinal cord samples from animals subjected to contusion injury (1dpi) or uninjured controls.


title: Dosing interval is a major factor determining the quality of T cells induced by SARS-CoV-2 mRNA and adenoviral vaccines

city: Oxford

first: Sam

middle: Michael

last: Murray

address: Nuffield Department of Medicine, Roosevelt Dr, Headington

country: United Kingdom

institute: University of Oxford

summary: Functional T cell responses are crucial for protective immunity induced by COVID-19 vaccination, but factors influencing the quality of these responses are incompletely understood. We employ an activation induced marker (AIM) assay and single-cell transcriptomic sequencing to analyze SARS-CoV-2 spike-responsive T cells following mild SARS-CoV-2 infection or following one or two doses of mRNA-LNP or adenoviral vectored COVID-19 vaccines. Our findings reveal broad functional and clonal heterogeneity in T cells generated by vaccination or infection, including multiple distinct effector populations. T cell function was largely conserved between COVID-19 vaccine platforms but was distinct compared to SARS-CoV-2 infection. Notably, the dosing interval greatly influenced the quality of T cells after two vaccine doses, particularly after mRNA-LNP vaccination, where a longer interval led to reduced inflammatory signaling and increased secondary proliferation. These insights enhance our understanding of SARS-CoV-2 specific T cells and inform the optimization of mRNA vaccination remens.

overall_design: CITE-seq: Peripheral blood mononuclear cells (PBMC) were sampled from human donors before or after one or two doses of BNT162b2 or ChAdOx1 nCoV-19 vaccination (COVID-19 vaccines), or at two timepoints post mild COVID-19 infection. Within vaccine groups, individuals were vaccinated at 3-4week (short) or 8-12week (long) intervals - leading to a total of 5 study groups with six participants in each group: ChAd-short, ChAd-long, BNT-short, BNT-long, COVID-19 infected. For samples taken at timepoints after COVID-19 vaccination or infection, PBMC were incubated for 24hours with overlapping peptide pools covering the entire SARS-CoV-2 spike protein (S1 and S2).  Fluorescence-assisted cytometric sorting (FACS) was used to sort CD69+ and OX-40+ and/or 4-1BB+ CD3+ lymphocytes. 10X Genomics 5' Next-GEM single-cell multi-omic sequencing was then performed on the sorted T cells, assessing gene expression, TCR and a custom panel of 8 CITE-seq markers.  For PBMC samples taken pre-vaccination, a CellTrace Violet (CTV) proliferation assay was performed, whereby cells were incubated for 7-days with overlapping peptide pools covering the entire SARS-CoV-2 spike protein. CTV low CD3+ cells were sorted using FACS and sequenced as above.

middle: M

first: Nicholas

last: Provine


title: Rationalistic Modulation of Immune Mechanisms Synergizes the Anti-Tumor Effects of Targeted Radiation Therapy in Pre-clinical Models

city: San Diego

first: Tao

last: Xie

state: CA

address: 10770 Science Center Dr

country: USA

institute: Pfizer

summary: In this study, we employed a diverse set of pre-clinical syngeneic murine tumor models with varying immune profiles to investigate the immunological impact of radiotherapy. We observed that immunologically 'hot' tumors demonstrated stronger tumor growth inhibition (TGI) following RT compared to 'cold' tumors. Additionally, RT induced both pro- and anti-inflammatory shifts within the tumor immune microenvironment, with significant changes noted in the spleen and tumor-draining lymph nodes. Importantly, radiotherapy led to an intratumoral increase in proliferating CD8 T cells, while the population of proliferating macrophages was notably reduced. Our findings also reveal that STING deficiency compromises TGI, particularly in tumors with a high baseline population of macrophages expressing an interferon response signature. Moreover, we identified a synergistic tumor growth control effect when combining HPK1 deficiency with RT. Collectively, these results provide insights into the complex interplay between RT and the immune response, offering potential avenues for novel therapeutic combination strategies.

overall_design: Syngeneic cancer cell lines were implanted subcutaneously in the flanks of mice, and established tumors were treated with 12 Gy radiotherapy. Six days post-treatment, tumors were harvested, immune cells were isolated by magnetic-assisted cell sorting, and analyzed by single-cell RNA sequencing.

first: Tristan

last: Wirtz

first: Catherine

last: Lee

first: Sripad

last: Ram

first: Sepideh

last: Mojtahedzadeh

first: Nicole

last: Streiner

first: Kavon

last: Noorbehesht

first: Lisa

last: Manzuk

first: Allison

last: Rohner

first: Edward

last: Cabral

first: Vinicius

last: Bonata

first: Timothy

last: Affolter

first: Manfred

last: Kraus

first: Christopher

last: Dillon

first: Anand

last: Giddabasappa


title: Effects of Bach2 on pancreatic gene expression in mice

city: Rostock

first: Robert

last: Jaster

address: Ernst-Heydemann-Str. 6

department: Dept. of Medicine, Div. of Gastroenterology

country: Germany

institute: Rostock University Medical Center

summary: The role of Bach2, a transcriptional repressor and key regulator of adaptive immunity, in the pathogenesis of autoimmune pancreatitis (AIP) is incompletely understood. Here, we used C57BL/6N mice with a targeted deletion of the Bach2 gene (Bach2 knockout mice) to study their susceptibility to AIP. The mice were kept for 18 weeks, before the pancreas and further organs were assessed histopathologically and by molecular methods. Results: Consistent with previous studies, Bach2 knockout mice showed reduced growth and developed a fatal lung disease necessitating humane euthanasia at the age of 18 weeks. Pancreatic infiltrates with lymphocytes frequently occurred in Bach2 knockout mice but not in wild-type animals. In more severe cases, this was associated with the focal destruction of exocrine pancreatic tissue and mild fibrosis. In the principal component analysis of pancreatic RNA-seq data, Bach2 knockout mice clustered separately from wild-type comtrols. Further data analysis revealed the activation of processes linked to adaptive immunity in pancreatic tissue of Bach2 knockout mice. We conclude that Bach2-deficient C57BL/6N mice are prone to the spontaneous development of AIP. This could be due to a disturbed immune homeostasis with dysregulated B and T cell activation. We thank Helmholtz Zentrum Muenchen - German Research Center for Environmental Health  (GmbH) for providing the mutant mouse line (Allele:  Bach2tm1b(EUCOMM)Wtsi), INFRAFRONTIER/EMMA (www.infrafrontier.eu, PMID: 25414328), and EMMA node at Helmholtz Zentrum Muenchen - German Research Center for Environmental Health (GmbH) from which the mouse line was distributed (RRID:IMSR_EM:09104). Associated primary phenotypic information may be found at www.mousephenotype.org.

overall_design: Mice with a constitutive whole-body knockout of the Bach2 gene and wild-type controls were kept until they reached 18 weeks of age. RNA was isolated from pancreatic tissue of 2 males and 2 females per group and subjected to RNA seq analysis.


title: An early-stage 3D fibroblast-featured tumor model mimics the gene expression of the naïve tumor microenvironment, including cancer progression and drug resistance phenotype.

city: Philadelphia

first: Francesca

last: Costabile

address: 3400 Civic Center BLVD

institute: University of Pennsylvania

summary: The tumor microenvironment (TME) plays a crucial role in cancer progression, yet the interactions between tumor cells and stromal components, such as fibroblasts, remain poorly understood. Traditional two-dimensional (2D) culture models fail to accurately replicate the complexities of the TME, hindering progress in cancer research and drug development. This study presents a novel 3D spheroid model, generated using the hanging drop system, that incorporates both tumor cells (B16F10 mouse melanoma) and fibroblasts (NIH/3T3), and aimed at simulating the early-stage TME. We demonstrate that fibroblasts are essential for ECM deposition, a key feature absent in tumor cells-only spheroids. Co-cultured spheroids exhibited a more organized structure, enhanced ECM deposition (type-VI collagen), and more closely resembled the morphology of native tumors compared to monocultures. RNA sequencing analysis revealed that the gene expression profile of B16F10–NIH/3T3 spheroids closely matched that of in vivo tumors, with 693 genes involved in critical pathways such as “pathways in cancer” and those linked to drug resistance. These findings highlight the importance of fibroblast inclusion in 3D models to replicate the genetic and structural features of the TME. Our spheroid system provides a more accurate representation of early tumor stages and offers a promising platform for drug screening, reducing the need for in vivo models by allowing the selection of the most effective compounds for further testing. This work underscores the potential of 3D culture systems in advancing our understanding of tumor biology and improving the precision of cancer therapeutics.

overall_design: RNA-seq profiling of B16F10 cells cultured in the different condition: sorted from B16F10–NIH/3T3 spheroids, sorted from B16F10 generated - in vivo tumor, sorted from standard two dimentional B16F10–NIH/3T3 coculture, B16F10 standard two dimentinally cultured.

first: Subin

last: George

first: Andrea

last: Facciabene

first: Gilberto

last: Filaci

first: Maddalena

last: Mastrogiacomo


title: Carnosic acid in topical rosemary extract promotes TRPA1-mediated scarless wound healing

city: philadelphia

first: Emmanuel

last: Rapp Reyes

address: 421 curie boulevard brb 1039

summary: Mammalian skin wounds typically heal with a scar, characterized by fibrotic tissue that disrupts original tissue architecture and function. Therapies to reduce fibrosis and scar formation remain a large unmet clinical need. Rosemary extract, particularly in the form of topical oils and creams, has gained massive popularity within the general public as a potential treatment to reduce scar formation. However, its efficacy and potential mechanism of action remain unknown. We show in adult wound healing mouse models that an ethanol-based rosemary extract reduces fibrosis and promotes scarless wound healing. More specifically, carnosic acid within rosemary leaves acts as an active ingredient and stimulates the TRPA1 nociceptor on cutaneous sensory neurons to promote scarless wound healing. Mice lacking TRPA1 on sensory neurons do not exhibit rosemary-induced tissue regeneration. Taken together, topical rosemary extract may be an effective and cost-efficient therapeutic to reduce scar formation in skin wounds.

overall_design: Inured ears of WT mice treated with control cream or rosemary cream collected at day 7 post injury

last: Rapp

first: Thomas

last: Leung


title: The role of Endothelial cell-derived Extracellular Vesicles in modulating Fibroblast Function in Skin Wound Healing

city: Chicago

first: Heidi

last: Yuan

address: 801 S Paulina St

institute: University of Illinois Chicago

summary: An adequate wound healing response requires the coordination of intercellular signals between multiple cell types. Following skin injury, endothelial cellsand fibroblasts provisionally replace the site of injury with newlyvascularized granulation tissue. Given the spatial and temporal overlap ofthese two cell types in the healing response, we hypothesize that endothelial cell-derived extracellular vesicles (ECEVs) could provide a communication pathway by which ECs influence fibroblast behavior. In this study we investigated the effects of ECEVs on fibroblast function both in vitro and in vivo through a murine skin wound healing model. Transcriptomic analysis showed that the uptake of ECEVs by fibroblasts altered functions and pathways relating to cel ldivision, ECM organization, and fibrosis. This transcriptomic analysis was functionally corroborated through in vitro assays in which fibroblasts demonstrated enhanced proliferation, migration, cell cycle progression,collagen contraction, and altered ECM deposition following ECEV treatment. Administration of ECEVs to healing mouse wounds led to greater collagen density and an upregulated fibroblast quantity in the wound bed of scar tissue. Our study highlights a novel role for ECEVs in affecting fibroblast function in the context of wound repair. Future studies can extrapolate this to other contexts where vascularization may affect tissue fibrosis.

overall_design: RNA-sequencing (RNA-seq) profiling of primary human dermal fibroblasts treated with control media or control media containing endothelial cell-derived extracellular vesicles (ECEVs) for 72h.

first: Anna

last: Salapatas

first: Trevor

middle: R

last: Leonardo

first: Chen

last: Han

first: Bhavya

last: Vegesna

first: Koushik

last: Debnath

first: Lin

last: Chen

first: Sriram

last: Ravindran

first: Luisa

middle: A

last: DiPietro


title: WAGO-4 Germ Granule Localization Ensures Fidelity in Small RNA Loading

city: Wien

first: Stela

last: Jelenic

address: Dr. Bohr-Gasse 3

country: Austria

institute: Institute of Molecular Biotechnology

type: Non-coding RNA profiling by high throughput sequencing

summary: Germ granules are liquid-like condensates that regulate small RNA pathways and gene expression, ensuring genome stability and fertility in animals. In C. elegans, several Argonaute proteins, central players of small RNA pathways, localize to germ granules, yet the functional significance of this spatial enrichment remains unclear. Here, we disrupted the localization of WAGO-4 to germ granules by introducing targeted mutations in the FG repeats of Vasa-like GLH proteins. These mutations did not disrupt overall germ granule architecture but significantly reduced WAGO-4 partitioning, leading to its predominant localization in the cytoplasm. Functional analyses revealed that this mislocalization partially compromised WAGO-4 activity, resulting in reduced WAGO-4 binding of small RNAs targeting specific genes, particularly those not co-regulated by CSR-1. This selective effect highlights the importance of WAGO-4’s spatial localization for efficient small RNA loading and gene regulation. Our findings demonstrate that germ granules serve as specialized compartments that fine-tune Argonaute function, emphasizing the role of phase-separated condensates in modulating RNA pathways and gene regulatory networks.

overall_design: Comparative rRNA depleted RNA-seq of C. elegans mutants 3xFG, WAGO-4 KO (200a) and WAGO-4(Y611E)(272a) compared to the WT

first: Mathias

middle: S

last: Renaud

first: Samantha

last: Del Borrello

first: Joseph

last: Gokcezade

first: Janos

last: Bindics

first: Lisa

last: Frasz

first: Philipp

last: Czermak

first: Peter

last: Duchek

first: Julie

last: Claycomb


title: Enhancer rewiring orchestrates inflammation and loss of cell identity during muscle stem cell aging

city: Baltimore

first: Supriyo

last: De

state: Maryland

address: 251 Bayview Blvd

department: Laboratory of Genetics and Genomics

institute: NIA-IRP, NIH

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

overall_design: Refer to individual Series


title: Genome-wide DNA methylation profiling of human CD4+ T helper cells define epigenetic signatures for Th1, Th17 and non-classic Th1 cells

city: Braunschweig

first: Stefan

last: Floess

address: Inhoffenstr. 7

department: Experimental Immunology

institute: Helmholtz-Centre for Infection Research

type: Methylation profiling by high throughput sequencing

summary: Naive CD4+ T cells are highly plastic cells that - upon activation - can differentiate into various T helper (Th) cell fates characterized by the expression of specific transcription factors and effector cytokines. Their expression can be stabilized by epigenetic mechanisms including DNA methylation. So far, our knowledge about the link between DNA methylation and T helper cell differentiation processes is fragmentary. In this study, we generated methylomes of CD4+ T helper cells by performing a whole-genome bisulfite sequencing analysis of ex vivo isolated human naïve T cells and selected clones of Th1, non-classic Th1 and Th17 cells. The pairwise comparison of the methylomes identified differentially methylated regions (DMRs) mainly located within introns, followed by promoter regions and exons. In accordance with literature, several DMRs were identified within lineage-specific loci like IFNG, TBX21, IL-17A and RORC. Thus, the identified DMRs are suitable to further characterize the T helper lineages and might be helpful to define lineage-specific regulatory modules involved in human T helper cell differentiation.

overall_design: Whole-genome bisulfite sequencing (WGBS) of human ex vivo isolated naive T cells and Th1, Th17 and non-classic Th1 cell clones.

first: Michael

last: Beckstette

first: Anna

last: Ntalli

first: Jochen

last: Huehn


title: Gene expression profiling of skin and blood in human acute cutaneous Lyme disease

city: Vienna

first: Georg

last: Stary

address: Waehringer Guertel 18-20, 1090 Wien

department: Department of Dermatology

institute: Medical University Of Vienna

summary: We investigated T cell polarization and Langerhans cell gene expression in acute Lyme disease compared to autoloagous healthy control skin.

overall_design:
Single cell suspensions were obtained from skin and blood of LD patients, FACS-sorted and sequenced using the 10XGenomics platform as previously described (Saluzzo et al. STAR Protoc. 2022 Jun 17;3(3):101470. doi: 10.1016/j.xpro.2022.101470. PMID: 35769924; PMCID: PMC9234085.

***************************************************************
Raw files for human/patient samples were not submitted to GEO due to concerns about submitting personally identifiable sequence data for open access.
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first: Johanna

last: Strobl

first: Florian

last: Winkler

first: Ruth

last: Dingelmaier-Hovorka


title: Single cell multi-omic sequencing of human peripheral mononuclear cells (PBMCs) post heart transplantation

city: La Jolla

first: Yuko

last: Tada

address: 9500 Gilman Dr

institute: UC San Diego

summary: PBMCs collected from patients with or without graft dysfunction post heart transplantation  were analyzed using the BD Rhapsody CITE-seq and VDJ-seq. The study groups include patients with cardiac allograft dysfunction (CAV), non-specific graft dysfunction (NGD), and normal graft function (Normal HTx).

overall_design:
Cryopreserved PBMCs were thawed and labeled with antibody derived tags followed by single cell CITE-seq and VDJ-seq.

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Submitter states that missing raw files are due to file loss.
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first: Paul

last: Kim

first: Klaus

last: Ley


title: Alternative splicing and differential gene expression during changes in endometrial receptivity in patients with recurrent implantation failure

city: Hangzhou

first: Jiucheng

last: Chen

state: Zhejiang

address: No.866 Yuhangtang Road

country: China

institute: Zhejiang University

summary:
Study question  : Are there changes in transcript levels as endometrial receptivity changes in patients with recurrent implantation failure (RIF), particularly in alternative splicing events (ASEs), differential expression genes (DEGs) and  immune cells   levels?
Summary answer: Changes in endometrial receptivity affect alternative splicing (AS), differential gene expression and immune cells, and the RNA-binding protein (RBP) KHDRBS3 likelymaybe？ Likely？ is involved in the dysregulation of alternative splicing.
What is known already: Endometrial receptivity is critical for RIF patients. Researchers have developed transcript level-based models to predict personalized endometrial assessment at WOI. Investigating alternative splicing dysregulation, immune cells and other transcriptional biomarkers in RIF patients may further help to predict endometrial receptivity and improve embryo implantation success.
Study design, size, duration: 90 endometrial samples were collected from fertile healthy controls (HC) during the menstrual cycle, including pre-receptive (PR) (luteinising hormone (LH)+3 days/LH+5 days), receptive (R) (LH+7 days) and post-receptive (PS) (LH+9 days) phases. 73 endometrial samples were taken from RIF patients on hormone replacement therapy (HRT) cycles, including PR (progesterone (P)+3 days), R (P+5 days) and PS (P+7 days) phases.
Participants/materials, setting, methods: Total RNA was extracted and sequenced for all 163 endometrial samples  . Bioinformatic analysis was used to comprehensively investigate the endometrial transcriptome at different stages of receptivity. We elucidated the biological significance of the universal alteration of alternative splicing in endometrial receptivity and further analyzed the role of immune cells and gene expression in this process. In addition, an ASEs-RBPs correlation network was constructed in the endometrial receptive period to evaluate the dysregulation of AS by RBPs.
Main results and the role of chance: Global splicing profiling of the four subgroups (HC: PR vs. R; HC: PS vs. R; RIF: PR vs. R; RIF: PS vs. R) consistently showed that skipped exons (SE) and mutually exclusive exons (MXE) were the most common changes in splicing events. These different ASEs occurred at high frequency and some of them could affect the coding potential of a transcript. A greater number of genes with different alternative splicing events (ASEGs) were identified between PR and R phases than between PS and R phases, and the same trend was observed for differentially expressed genes (DEGs). Both ASEGs and DEGs showed enrichment for pathways associated with cell function and adhesion, while DEGs were also enriched for cytokine and immune-related pathways. Immune cell prediction shows that changes in endometrial receptivity phases result in differences in the levels of CD8 T cells, resting NK cells and other immune cells. During the R phase, there was a significant decrease in CD8 T cells and resting NK cells, whereas monocytes and M0 macrophages were significantly increased compared to the endometrial PR phase in RIF patients. Similarly, analysis of immune cells showed a significant increase in CD8 T cells, M1 and M2 macrophages and a significant decrease in activated NK cells in the R phase compared to the PS phase of the endometrium in RIF patients. Some of these immune cells, such as NK cells and monocytes, are highly correlated with certain ASEs. Investigation of ASEs-RBPs networks suggested that KHDRBS3 may be a key regulator of splicing events.
Limitations, reasons for caution: The sample size needs to be increased and cover a variety of situations, such as different age, pathology, dietary habits, etc., and alsothe molecular experimental validation needs to be considered. The lack of direct comparisons between natural and HRT cycles in the same sample and molecular experimental validation need to be considered.
Wider implications of the findings: Comprehensive transcriptional analysis of the endometrium provide valuable insight into potential biomarkers related to endometrial receptivity and potential treatments for RIF patients.
Study funding/competing interest(s): This study was supported by Supported by the Program of National Natural Science of China (Grant No. 81701409). All authors declared that they have no conflicts of interest.

overall_design: Additionally, 73 samples were collected from RIF patients undergoing hormone replacement therapy (HRT) cycles, including PR (progesterone (P) +3 days), R (P +5 days), and PS (P +7 days).

first: Mao-li

last: Wang

first: Bing-jie

last: Lu

first: Han

last: Wu

first: Wen-bi

last: Zhang


title: African spiny mice show resistance to DMBA/TPA-induced squamous carcinogenesis with distinct benign tumor profile [acomys_dt]

city: Rochester

first: Francesco

last: Morandini

state: NY

address: 402 Hutchison Hall

institute: University of Rochester

summary: African spiny mice, Acomys cahirinus (Acomys), is an emerging mammalian model for regeneration studies because of its ability to morphologically and functionally regenerate several tissues. However, as limited regenerative abilities in most mammals are viewed as an antitumor strategy, it is intriguing how Acomys balance regeneration and tumor suppression. To date, no spontaneous tumors have been reported in Acomys, suggesting it may be resistant to cancer. In this study, we use in vitro and in vivo approaches to investigate if A. cahirinus is susceptible to carcinogenesis. We report that, like mice, two oncogenic hits - activation of oncogenic RasG12V and inactivation of p53 or pRb - are sufficient to transform skin fibroblasts from A. cahirinus. However, surprisingly, active p53 dramatically reduces the tumor forming ability of the transformed fibroblasts.  In the DMBA/TPA treatment model of in vivo skin carcinogenesis with animals aged 1-1.5 years, C57BL/6 laboratory mice formed pre-malignant skin tumors in 22 weeks, while A. cahirinus did not show any effects. A longer TPA treatment period of 30 weeks induced benign sebaceous adenomas in A. cahirinus. This showed that A. cahirinus was resistant to squamous skin carcinogenesis typically induced by DMBA/TPA treatment. A. cahirinus showed significantly higher upregulation of immune response and differentially expressed tumor suppressors and oncogenes in DMBA/TPA-treated skin when compared to mice. Wnt/β-catenin signaling, a major mediator of squamous carcinogenesis associated with DMBA/TPA treatment, was inhibited in A. cahirinus. Several Wnt/β-catenin targets and effector transcription factors, especially Lef1, were transcriptionally downregulated in A. cahirinus. Overall, our study shows that A. cahirinus may be resistant to DMBA/TPA-induced squamous carcinogenesis, and this resistance may be mediated by a strong immune response and the inhibition of Wnt/β-catenin signaling.

sample_organism:
Acomys cahirinus
Mus musculus

platform_organism:
Acomys cahirinus
Mus musculus

overall_design: RNA-seq profiling of A. cahirinus, C56BL6 mouse, wild mouse fibroblasts 24h and 12d after 0 gy / 10 gy / 20 gy gamma irradiation

first: Fathima

last: Athar

first: Iqra

last: Fatima

first: Isabella

last: Silvestri

first: Sei Joong

first: Minseon

last: Lee

first: Xiaoyan

last: Liao

first: Andrei

last: Sharov

first: Vladimir

last: Botchkarev

last: Seluanov

first: Vera

last: Gorbunova


title: Targeting PLK1 with Onvansertib and Radiation in Head and Neck Cancer

city: Cincinnati

first: Julianna

last: Korns

state: OH

address: 3100 Eden Avenue

institute: University of Cincinnati

summary: Radiotherapy is a key treatment for head and neck squamous cell carcinoma (HNSCC) especially in patients with human papilloma virus-negative (HPV-) HNSCC. Our studies identify polo-like kinase 1 (PLK1) as a promising target for HNSCC. PLK1 inhibition with onvansertib reduces cell viability and spheroid growth of HPV- HNSCC cells. PLK1 inhibition combined with radiation increases G2/M arrest, decreases colony formation of HPV- HNSCC cells, and reduces tumor growth in vivo. RNA-sequencing demonstrates radiation increases MMP10 expression but PLK1 inhibition reverses this effect in HPV- HNSCC cells. PLK1 inhibition also reduces MMP10 activity in HPV- HNSCC cells. These findings suggest that combining PLK1 inhibition with radiation may improve therapeutic response for HPV- HNSCC via MMP10, offering a novel approach to overcome radiation resistance.

overall_design: RNA-seq profiling of human head and neck squamous cell carcinoma (HNSCC) cell lines (HN5, UMSCC47) treated with the PLK1 inhibitor onvansertib in combination with radition. HNSCC cells were treated with vehicle, onvansertib alone, radiation alone, and onvasertib combined with radiation (n = 3). HNSCC cells were treated with onvansertib for 24 hours then irradiated (4 Gy), and RNA was collected 24 hours later.

first: Mario

last: Medvedovic

first: Vinita

last: Takiar


title: Mechanisms of assembly and function of the Hsp70-Hsp40 chaperone machinery

city: Heidelberg

first: Santiago

last: Maya

address: Im Neuenheimer Feld 329

institute: Zentrum für Molekulare Biologie der Universität Heidelberg (ZMBH)

summary: The orchestration of protein folding and maintenance by molecular chaperones is crucial for cellular homeostasis. Among them, Hsp70 and its co-chaperone Hsp40 form a central machinery that remodels client proteins involved in numerous biological processes. Despite their essential role, how Hsp70 and Hsp40 assemble into a functional complex has remained elusive. Here, we integrated cryo-electron microscopy and nuclear magnetic resonance spectroscopy to determine the architecture of the full-length Hsp70-Hsp40 machinery. The structure of the complex in a physiological inactive state reveals distinct inhibitory mechanisms. In the active state, a new set of interactions is formed wherein the Hsp40 G/F-rich region acts as a pseudo-substrate and binds to the Hsp70, thereby directly modulating the refolding process. The G/F region maintains an Hsp40 autoinhibited state; upon binding to Hsp70, this state is disrupted, exposing a cryptic client-binding site and enabling interaction with protein clients to facilitate their refolding. The transition between these states is central to regulating the machinery’s refolding efficiency. Disrupting the Hsp40 autoinhibited state or the G/F–Hsp70 interaction leads to functional impairments that trigger a pronounced heat shock response in cells as a compensatory mechanism. Our findings provide crucial insights into the regulatory dynamics of this essential chaperone machinery, with broad implications for understanding protein homeostasis and the cellular response to stress.

sample_organism: Escherichia coli str. K-12 substr. MG1655

overall_design: Ribosome profiling analalysis of wt E. coli MG1655 and mutant (DnaJF98A - F102A, CbpAF98A - F102A) cells, at optical density (OD600) of 0.5 and 2.0.

first: Yajun

last: Jiang

first: Ziad

last: Ibrahim

first: Mary

last: Clay

first: Youlin

last: Xia

first: Alexandre

last: Myasnikov

first: Kaylan

last: Immadisetty

first: Zhilian

first: Liang

last: Tang

first: Paolo

last: Rossi;

first: Pritha

last: Ganguly

first: Jiangshu

last: Liu

first: Darcie

last: Miller

first: Meixia

last: Che

last: palacios

first: Gunther

last: Kramer

first: Bernd

last: Bukau

first: Charalampos

middle: B

last: Kalodimos


title: African spiny mice show resistance to DMBA/TPA-induced squamous carcinogenesis with distinct benign tumor profile [acomys_irrad]

first: Francesco

last: Morandini

first: Fathima

last: Athar

first: Iqra

last: Fatima

first: Isabella

last: Silvestri

first: Sei Joong

last: Kim

first: Minseon

last: Lee

first: Xiaoyan

last: Liao

first: Andrei

last: Sharov

first: Vladimir

last: Botchkarev

last: Seluanov

first: Vera

last: Gorbunova


title: Chaperone-mediated autophagy sustains muscle stem cell regenerative function

city: Barcelona

first: Ignacio

last: Ramirez Pardo

state: Barcelona

address: Carrer del Doctor Aiguader, 88, Ciutat Vella

department: Department of Medicine and Life Sciences

country: Spain

institute: Universitat Pompeu Fabra

summary: Proteostasis supports stemness and its loss correlates with the functional decline of diverse stem cell types. Here we identify that chaperone-mediated autophagy (CMA), a selective autophagy pathway, is necessary for the regenerative capacity of muscle stem cells (MuSCs) throughout life. We show that CMA is active in MuSCs, while genetic loss of CMA in MuSCs or failure of CMA in normally aged cells causes a proliferative impairment, resulting in defective skeletal muscle regeneration. Reactivation of CMA in old MuSCs boosts their proliferative capacity and improves their regenerative ability. Using comparative proteomics to identify CMA substrates, we uncovered actin cytoskeleton and glycolytic metabolism as key processes altered in aged mice and human MuSCs. Our findings reveal CMA to be a decisive stem-cell-fate regulator, with implications for fostering muscle regeneration in old age.

overall_design: bulk RNA-seq of isolated muscle stem cells (MuSCs) at 0 (uninjured), 1 and 4 dpi (days post injury) from Lamp2a-WT and Lamp2A-Pax7CreER mice.

last: Ramírez-Pardo

first: Silvia

last: Campanario-Sanz

first: Eusebio

last: Perdiguero

first: Joan

last: Isern

first: Pura

last: Muñoz-Cánoves

first: Ana Maria

last: Cuervo


title: Mesenchymal Colorectal Cancers Secrete Vesicles With Unique Cargo That Can Be Used For Liquid Biopsy Based Diagnostics

city: Amsterdam

first: Sander

middle: Roeland

last: van Hooff

address: Meibergdreef 9

department: Center for Experimental and Molecular Medicine (CEMM),

country: Netherlands

institute: AUMC


title: African spiny mice show resistance to DMBA/TPA-induced squamous carcinogenesis with distinct benign tumor profile

first: Francesco

last: Morandini


title: Thymosin α1 Inhibiting Ferroptosis of Dental Pulp Cells in Pulpitis

city: Guangzhou

first: Zhongchun

last: Tong

address: Lingyuan West Road 56#

department: Hosiptal of Stomatology

institute: Sun Yat- Sen University

summary: Tooth pulpitis is a common oral ailment. Understanding the underlying mechanisms of pulpitis and developing effective treatment strategies are of significant importance.In this study, single-cell RNA sequencing (scRNA-seq) was used to identify cellular heterogeneity between 3 pulpitis tissue and 3 healthy pulp tissue. Raw data of ScRNA-seq of 3 healthy pulp tissue have been uploaded (GEO accession: GSE274562; Control1:  GSM8452929; Control2: GSM8452930; Control3: GSM8452931).

overall_design: To dissect cellular heterogeneity and investigate ferroptosis in each type of cells in inflammatory dental pulp, we harvested 3 healthy dental pulp and 3 inflammatory dental pulp. The 6 samples were dissociated into single cells suspension and then performed scRNA-seq. After quality control, the transcriptomes of 40,231 cells (Pulpitis: 17,814; Healthy pulp: 22,417) were obtained. These cells were subsequently visualized into 12 distinct cell clusters including CD4+ T cell, CD8+ T cell, B cell, Macrophage, Neutrophil, Mast cell, Plasma cell, Fibroblast, Endothelial cell, Mesenchymal stem cell (MSC), Schwann cell (ScC), nonmyelinating ScC (nmScC), by generating uniform manifold approximation and projection (UMAP) plots, based on hierarchical clustering and established lineage-specific marker genes

first: Jie

last: Wu

first: Qimei

last: Gong

first: Wenxuan

last: Liu

first: Aijia

last: Chen

first: Zekai

first: Yihua

last: Huang

first: Wenkai

last: Jiang


title: Mesenchymal Colorectal Cancers Secrete Vesicles With Unique Cargo That Can Be Used For Liquid Biopsy Based Diagnostics [RNA-seq plasma]

summary: Tumor-derived extracellular vesicles (TEVs) play a crucial role in cancer progression, metastasis and therapy resistance but their distinct profiles across different cancer stages and molecular subtypes remain underexplored. This study analyzed TEVs from epithelial (CMS2) and mesenchymal (CMS4) subtypes of colorectal cancer (CRC) using six cell lines and clinical samples. Investigation of the cargo of vesicles secreted by the two subtypes revealed significant differences in mRNA, miRNA, and protein profiles between the two subtypes. Notably, CMS2 predominantly secreted smaller, Tetraspanin-8 (TSPAN8) enriched EVs, while CMS4 produced both larger and smaller EVs, enriched in TSPAN4. This underscores the complexity of vesicle heterogeneity between these subtypes. Additionally, we assessed miRNA profiles from plasma-derived bulk TEVs in CRC patients. Our integrative analysis identified a distinct miRNA signature specific to each subtype, indicating that TEVs from CMS2 and CMS4 cells can be detected in circulation and may serve as potential diagnostic tool for CRC.

overall_design:
To investigate the mRNA profiles from plasma-derived TEVs in CMS2 and CMS4 subtyped CRC patients.

***************************************************************
Raw files for human/patient samples were not submitted to GEO due to concerns about submitting personally identifiable sequence data for open access.
***************************************************************

first: Paris

middle: J

last: Asif

first: Lauri

middle: H

last: Borghuis

first: Anita

middle: E

last: Grootemaat

first: Monique

last: van Eijndhoven

first: Johan

last: de Rooij

first: Nils

last: Groenewegen

first: Nicole

middle: N

last: van der Wel

first: Arezo

last: Torang

first: Inne

last: Borel Rinkes

first: Cristina

last: Gómez-Martín

first: Hans

middle: F

last: Pruijt

first: Jennifer

last: Perez Boza

first: Onno

last: Kranenburg

first: Tineke

last: Buffart

first: Michiel

middle: D

last: Pegtel

first: Jan Paul

last: Medema


title: The regulatory code of injury responsive enhancers enables precision cell state targeting in the CNS [enhancer_aav]

city: Stockholm

first: Margherita

last: Zamboni

address: Solnavägen 9

institute: Karolinska Institutet

summary: Enhancer elements in the genome direct cell type-specific gene expression programs. In response to injury, cells undergo dynamic changes in gene expression that drive adaptation and initiate tissue repair. Here, we investigate how injury-induced gene expression programs are encoded within enhancer elements in the mammalian CNS. Leveraging single-nucleus transcriptomics and chromatin accessibility profiling, we identify thousands of injury-induced, cell type-specific enhancers in the mouse spinal cord. These enhancers are more abundant in glial cells and retain cell type specificity, even when regulating shared wound response genes. By modeling glial injury-responsive enhancers using deep learning, we reveal that their architecture encodes cell type specificity by integrating generic stimulus response elements with cell identity programs. Finally, through in vivo enhancer screening, we demonstrate that injury-responsive enhancers can selectively target reactive astrocytes bordering lesion sites across the CNS using therapeutically relevant gene delivery vectors. Our decoding of the principles of injury-responsive enhancers enables the design of sequences that can be programmed to therapeutically target disease related cell states.

overall_design: Animals from a Cx30-Tom transgenic line were injected with an enhancer AAV library to test the activity of injury-responsive enhancers identified in our analysis. Mice were then injured with a spinal cord contusion model (14dpi) and samples were collected after 3 days (or from uninjured controls). Single cells were obtained through tissue digestion and sorting and were processed using a transcriptomics experiment that included enrichment for barcodes associated with the enhancer viruses. Each condition encompassed 1-2 animals per sex.

first: Enric

last: Llorens Bobadilla


title: Differentially manufactured Arnica montana L. extracts mediate anti-inflammatory effects on human T cells via different signaling pathways

first: Yvonne

last: Samstag

address: INF 305

department: Institute of Immunology

institute: Heidelberg University Hospital

summary: In this study, differentially prepared Arnica extracts and the lead compounds Thymol and Helenalin were directly compared for their immunomodulatory potential using primary human T cells. Extracts from different plant parts (root or whole plant) and prepared by different manufacturing processes inhibited T cell activation and proliferation. nCounter gene expression analysis and subsequent gene set enrichment analysis showed that Arnica preparations intervene in T cell receptor (TCR) signaling. Interestingly, the three tested extracts were, however, found to mediate their effects via interfering with different TCR signaling pathways. The hydroethanolic root extract selectively inhibited the binding of NFκB to DNA, while the aqueous fermented whole plant extract preferentially inhibited NFAT-dependent gene expression. The hydroethanolic whole plant extract (by trend) diminished both, NFκB DNA-binding and NFAT-dependent gene expression.

overall_design: Using the nCounter Human Immunology V2 Panel covering 579 immune-related genes, gene expression in PBTs treated with different Arnica extracts was compared to the respective solvent control sample (H2O or EtOH, respectively). 5 healthy donors were taken for each condition, treated or untreated.

first: Verena

last: Beier


title: Mesenchymal Colorectal Cancers Secrete Vesicles With Unique Cargo That Can Be Used For Liquid Biopsy Based Diagnostics [miRNA-seq plasma]

first: Sander

last: van Hooff

overall_design:
To investigate the miRNA profiles from plasma-derived TEVs in CMS2 and CMS4 subtyped CRC patients.

***************************************************************
Raw files for human/patient samples were not submitted to GEO due to concerns about submitting personally identifiable sequence data for open access.
***************************************************************

first: Paris

last: Asif

first: Lauri

last: Borghuis

first: Anita

last: Grootemaat

first: Monique

last: van Eijndhoven

first: D


title: CD48 affects type-2 innate lymphoid cell function and airway hyperreactivity.

city: Los Angeles

first: Kei

last: Sakano

state: CALIFORNIA

address: 1450 Biggy Street NRT5509

department: Molecular Microbiology and Immunology

institute: University of Southern California

summary: The importance of type-2 innate lymphoid cell (ILC2), as well as its function in allergic airway inflammatory diseases, have attracted growing attention over recent years. It is anticipated that further elucidation of ILC2 function and regulatory mechanisms will provide a new perspective on  potential therapeutic avenues. Here, we describe CD48 as an ILC2 function regulator. The deletion of CD48 has been demonstrated to impair ILC2 activation and proliferation. Moreover, the deletion of CD48 resulted in a reduction in allergen-induced airway hyperresponsiveness in mice. The diminished function of ILC2s resulting from CD48 deletion was offset by the administration of a PKC be-ta stimulator. Taken together, these results suggest that CD48 signaling activates ILC2s via the PKC be-ta pathway and that CD48 inhibition would be a potential new option for asthma therapy.

overall_design: mRNA profiles of mouse lung ILC2s from WT and CD48 KO mice were generated by deep sequencing, in duplicates, using a NextSeq 500.


title: Mesenchymal Colorectal Cancers Secrete Vesicles With Unique Cargo That Can Be Used For Liquid Biopsy Based Diagnostics [RNA-seq]

overall_design: To investigate the mRNA profiles of TEVs derived from CMS2 and CMS4 subtyped cell lines, we used two CMS2 and two CMS4 cell lines.

first: Johan

last: de Rooij

first: Nils

last: Groenewegen

first: Nicole

last: van der Wel

first: Arezo

last: Torang


title: The effect of post-hatching non-feeding on gene expression in the NIES-R and Hd-rR strains of medaka (Oryzias latipes).

city: SHizuoka-shi

first: Taijun

last: Myosho

state: Shizuoka-ken

country: Japan

institute: University of Shizuoka

summary: The aim of this study is to estimate the eleutheroembryonic stage based on gene expression changes following post-hatching starvation in the NIES-R and Hd-rR strains of medaka (Oryzias latipes). We analyzed the changes in gene expression patterns in post-hatching individuals under both feeding and starvation conditions. This analysis revealed how starvation affects development and growth, and identified key genes that play significant roles under nutrient-deficient conditions.

overall_design: To investigate the effect of starvation on gene expression, we obtained gene expression data using Lasy-seq from whole-body RNA samples collected from the NIES-R and Hd-rR strains on the day of hatching, with 4 feeding groups and 4 non-feeding groups (at 2, 4, 6, and 8 days post-hatching). Subsequently, PCA and GO analysis were conducted to identify genes whose expression increased or decreased depending on the presence or absence of feeding.

first: Makoto

last: Kashima


title: Time course analysis of the effect of reseeding on laminin-411 vs fibronectin on gene expression in in vitro differentiated pancreatic progenitor cells.

city: København N

first: Adam Kappel

last: Rump

address: Blegdamsvej 3

department: DanStem

country: Denmark

institute: University of Copenhagen

summary: The extracellular matrix has recently emerged as a key regulator of endocrine fate choice in the pancreas via integrins and YAP signaling. To elucidate the intermediate mechanism between exposure to specific ECM components and induction of a proendocrine transcriptional program, we performed bulk RNA sequencing of in vitro differentiated HUES4-derived pancreatic progenitor cells. Cells were reseeded on either fibronectin (FN, non-endocrinogenic) or laminin-411 (LN411, highly endocrinogenic) and RNA was collected after 3, 6, or 24 hours. The dataset revealed a progressive induction of an endocrinogenic program, but with significant upregulation of endocrine genes only at the 24 hour time point. Interestingly, other YAP target genes were strongly and significantly downregulated already at 6 hours. Additionally, the dataset revealed novel potential intermediate regulators between LN411-exposure, YAP inhibition and endocrine specification.

overall_design: To investigate the intermediate effects between exposure to laminin-411 and endocrine fate choice, ESC-derived pancreatic progenitor cells were reseeded on either laminin-411 or fibronectin and harvested after 3, 6, or 24 hours. Triplicates were collected and sequenced for each condition.

first: Christine

last: Ebeid

first: Adam


title: Single Nucleus Profiling Highlights the All-Brain Echinoderm Nervous System

city: Napoli

first: Maria Ina

last: Arnone

address: Villa Comunale 1

department: Biology and Evolution of Marine Organisms

country: Italy

institute: Stazione Zoologica Anton Dohrn

summary: By combining single nucleus transcriptomics with spatial gene expression analysis and high-resolution electron microscopy, we reconstructed the cell type atlas of post-metamorphic Paracentrotus lividus juveniles.

overall_design: Generation and analysis of Paracentrotus lividus juvenile snRNA-seq data to use along with imaging techniques to generate P. lividus juvenile cell atlas.

first: Paganos

last: Periklis

first: Ullrich-Lüter

last: Jack

first: Almazán

last: Alba

first: Voronov

last: Danila

first: Carl

last: Jil

first: Zakrzewski

last: Anne-C.

first: Zemann

last: Berit

first: Rusciano

last: Maria Lorenza

first: Sancerni

last: Tiphaine

first: Schauer

last: Maria

first: Akar

last: Oğuz

first: Caccavale

last: Filomena

first: Cocurullo

first: Benvenuto

last: Giovanna

first: Croce

last: Jenifer Carol

first: Lüter

last: Carsten

first: Arnone

last: Maria Ina


title: Moderate levels of folic acid benefit outcomes for cilia based neural tube defects

city: Boulder

first: David

last: Engelhardt

state: CO

address: 347 UCB

department: MCDB

institute: CU Boulder

summary: Neural tube defects (NTDs) are a common and devastating birth defect. The current NTD rate in the U.S. reflects an approximately 30% decrease following folic acid (FA) supplementation in the 1990’s as a public health measure. Yet, questions remain including which gene mutations are sensitive to folic acid levels and how folic acid acts to alter NTD risk. We studied the relationship between FA dosage and NTD rates in NTD mouse models and propose that genes involved in cilia formation and function represent an FA sensitive ontology and a possible avenue for further reducing NTD rates. We demonstrate that in cilia mutant mice, a more moderate level of FA is beneficial in reducing NTD rates and promoting primary and motile cilia formation and function. We present a mechanism through which fortified FA levels reduce basal levels of reactive oxygen species (ROS) which in turn reduces ROS-sensitive GTPase activity required for ciliogenesis.

overall_design: This dataset represents an investigation into differential gene expression between wild type and Rsg1L3P/L3P mouse embryonic fibroblasts isolated from F3 moderate and fortified folic acid diet mice.

first: Cara

last: Martyr

first: Lee

last: Niswander


title: Total RNA-seq analysis of WT and Gag chimera HIV-1 virions

city: Saint Louis

first: Sebla

middle: Bulent

last: Kutluay

state: MO

address: 660 S. Euclid Avenue, Campus Box 8230

institute: Washington University School of Medicine

summary: To investigate how the major HIV-1 structural protein Gag packages viral genomes efficiently, we replaced the NC domain of Gag with RNA binding domains from heterologous cellular RNA binding proteins to generate Gag chimeras.  We hypothesize that the Gag chimeras that preferentially bind to purine-rich RNA sequences will package viral genomes efficiently. We performed total RNA-seq to identify the RNAs that are packaged by WT Gag and Gag chimeras in HIV-1 virions.

overall_design: 293T cells in 6-well dishes were transfected with PR- NL4-3-derived proviral plasmids expressing the indicated Gag. Virions were harvested from cell culture supernatants 2 days post transfection and pelleted on a sucrose cushion. Total RNA was extracted from virions using TRIzol following manufacturer’s protocol. RNA was subsequently prepared for sequencing using Illumina Truseq stranded mRNA kit and Nextseq500 platform.


title: An intracellular magnetic torque-regulator drives mechano-sequential antitumor immunity

city: Shanghai

first: Rui

last: Gao

address: Zhennan rode

department: School of Medicine

institute: Tongji University

summary: Nature creates specific structures to control mechanical plasticity for cell fate regulation. Although immune cells can sense and adapt to extracellular mechanical cues, how to activate immune responses via intracellular mechanical stimulation remains a mystery. Here, we present an intelligent intracellular magnetic torque (IIMT) paradigm that is coupled with programmed rotating magnetic fields to generate the selective mechano-stimulations in lysosomes and induce sequential antitumor immune responses. A magnetic rod-shaped micromotor (MagRM) with the a unique geometric aspect ratio is designed to exert tunable torques in antigen-presenting cells (APCs) and tumor cells. Low-intensity torque induces the mild lysosomal membrane damage to trigger antigen cross-presentation and canonical NLRP3 inflammasome activation in APCs, whereas high-intensity torque causes lysosomal membrane rupture for immunogenic cell death. In vivo studies unveil that the optimal antitumor activity is achieved by generating sequential torques from high to low intensities, which is illuminating to explore mechano-immunotherapy with selective and spatiotemporal control.

overall_design: The underlying mechanism of mechano-regulation for tumor cell death post high-frequency megneic field (HMF) were explored for the analysis. The RNA sequencing was performed on OVA-expression B16F10 (B16-OVA) melanoma cells collected at 6 h after Control, magnetic rod-shaped micromotors (MagRMs) with HMF treatments.

first: Chang

last: Xu

first: Zhenyan

last: Zhu

first: Chen

last: Liu

first: Min

last: Wang

first: Xueyan

last: Wei

first: Peng

last: Ning

first: Wei

last: Zhang

first: Yingze

last: Li

first: Junwu

last: Shen

first: Yajuan

first: Yifan

last: Tong

first: Feng

last: Tao

first: Xiaolei

last: Chen

first: Xiaoyi

last: Jiang

first: Huan

last: Ren

first: Cheng

last: Lv

first: Gustavo

last: Plaza

first: Jason

last: Miska

first: Haiming

last: Fan

first: Yichao

last: Tang

first: Yalei

last: Dai

first: Weiwei

last: Yang

first: Bin

last: He

first: Yu

last: Cheng


first: Rui

last: Gao

overall_design: To fully explore the effects of IIMT-activated immune response in tumor microenvironment, we performed single-cell RNA sequencing (scRNA-seq) on all cells in tumors isolated at 24 h after IIMT (HL-2), MagRMs (M-1), and PBS (P-2) treatments for 10 days from the B16-OVA (BO) mice on day 11.

first: Chang

last: Xu

first: Zhenyan

last: Zhu

first: Chen

last: Liu

first: Min

last: Wang

first: Xueyan

last: Wei

first: Peng

last: Ning

first: Wei

last: Zhang

first: Yingze


title: RNA-Seq analysis of mouse neural stem cells with or wihtout DMTF1 knockdown.

city: Singapore

first: Oleg

last: Grinchuk

address: 61 Biopolis Dr, Proteos

country: Singapore

institute: IMCB

summary: Neural stem cells (NSCs) contribute to embryonic brain development and adult brain homeostasis via their unique properties of self-renewal and differentiation. The Cyclin D binding Myb‑like transcription factor 1 (DMTF1) is reported to be a tumor supressor gene in cancer cells. However,the role of DMTF1 in NSCs is poorly understood. The aim of this study is to determine the role of Dmtf1 in NSCs and elucidate the underlying mechanism.

overall_design: RNA-seq analysis of neural stem cells that are derived from wild type control mice without and with treatment  shRNAs against mouse Dmtf1.

first: Yajing

last: Liang

middle: V

first: Derrick

last: Ong


title: UBA2A is a regulator of seed dormancy and DOG1 mRNA chromatin retention in Arabidopsis

city: Warsaw

first: Michal

last: Krzyszton

address: Pawinskiego 5a

country: Poland

institute: Institute of Biochemistry and Biophysics, Polish Academy of Sciences

summary: Multiple factors control primary seed dormancy established during seed maturation and secondary seed dormancy established when a non-dormant seed is exposed to adverse conditions during imbibition. A key player in the control of both primary and secondary dormancy in Arabidopsis thaliana is the DOG1 gene, whose expression is extensively regulated at the transcriptional level. Despite its importance, the influence of posttranscriptional RNA processing and mRNA storage of DOG1 on the determination of dormancy depth remains elusive. We show that the UBA2A protein, a member of the hnRNP family, suppresses primary and secondary seed dormancy through the regulation of the DOG1 gene expres-sion at the posttranscriptional level. Surprisingly, single-molecule FISH, chromatin-attached RNA analysis and Pol II ChIP demonstrated that UBA2A do not control DOG1 gene transcription but rather DOG1 mRNA chromatin retention. Our study highlights chromatin retention as an important step in DOG1 gene expression regulation during dormancy establishment and shows that UBA2A like its human homolog hnRNPAB is implicated in RNA transport in the cell.

overall_design: Comparison of Col-0 and uba2a mutants (uba2a-1 and uba2a-3) dry seed transcriptomes (four biological replicas).

first: Ce

first: Szymon

last: Swiezewski


title: MMPs and NETs are detrimental in CNS-tuberculosis with MMP inhibitor doxycycline improving murine CNS-tuberculosis survival (human)

first: Catherine Wei Min

state: Singapore

address: 14 Medical Drive, MD6, Centre for Translational Medicine, #15-02

institute: National University of Singapore

summary: Despite effective anti-tuberculous treatment (ATT), up to 40% of patients with central nervous system tuberculosis (CNS-TB) suffer permanent neurological deficits and death. The immunopathological mechanism of CNS-TB remains undefined, hindering the development of new CNS-TB therapies. We here report the spatial transcriptomics profiling of human CNS-TB and demyelination CNS tissues using laser capture microdissection RNA sequencing (LCM-RNA Seq). Human CNS-TB and demyelination samples exhibited distinct gene expression profiles based on cellular phenotypes. Genes involved in biological processes relates to cytokine responses, cell-substrate adhesion, extracellular matrix organization, inflammatory and immune responses are highly expressed in both inflammatory and granuloma tissues in contrast to demyelination and normal tissues. Down-regulated genes are associated with CNS-related processes such as synapse and cell junction organisation, myelination, neuron development and differentiation.

overall_design: Clinical formalin-fixed paraffin-embedded (FFPE) samples were retrieved from human CNS-TB (n=6) and demyelination control (n=4). All tissue sections with 7 µm thickness each were stained with hematoxylin prior to LCM-RNA Seq. Target regions including granuloma, inflammatory region and adjacent normal regions were selected and microdissected for library construction. The libraries were sequenced and analysed.

first: Catherine

first: Joshua

middle: X

last: Tay

first: Andres

last: Vallejo

first: Fei

middle: K

last: Loh

last: Teo


title: MMPs and NETs are detrimental in CNS-tuberculosis with MMP inhibitor doxycycline improving murine CNS-tuberculosis survival (mouse)

summary: Despite effective anti-tuberculous treatment (ATT), up to 40% of patients with central nervous system tuberculosis (CNS-TB) suffer permanent neurological deficits and death. Immunopathology in CNS-TB, characterized by extensive brain inflammation and tissue destruction, is driven by a matrix-degrading phenotype, which results from an increase in matrix metalloproteinases (MMPs) relative to their tissue-specific inhibitors (tissue inhibitor of metalloproteinases, TIMPs). The immunopathological mechanism of CNS-TB remains undefined, hindering the development of new CNS-TB therapies. By performing laser capture microdissection RNA sequencing (LCM-RNA Seq), we profiled the spatial transcriptome of murine CNS-TB (M.tb H37Rv-infected C57BL6 Nos2-/- mice) and healthy murine (C57BL6 Nos2-/- mice)  as control. The granulomas from murine CNS-TB treated with ATT alone,  ATT with SB-3CT (MMP-2/9-specific MMP inhibitor), and ATT with doxycycline (broad spectrium MMP-inhibitor) were also profiled. Genes implicated in defense, immune, inflammatory and cytokine responses, neutrophil degranulation, antigen presentation and Toll-Like Receptor cascades, present predominantly in CNS-TB granulomas relative to other tissue types. On the other hand, genes relate to trans-synaptic signaling, synapse and cell junction organisation, neuron development and differentiation had decreased expression in murine CNS-TB inflammatory and granuloma tissues. We also found an increased expression of genes involve in response to oxidative stress, apoptosis and programmed cell death in murine CNS-TB treated with ATT plus adjunctive MMP inhibitors.

overall_design: Formalin-fixed paraffin-embedded (FFPE) brain tissues were processed from murine CNS-TB (n=4), healthy murine (n=4), murine CNS-TB treated with ATT (n=8), murine CNS-TB treated with ATT plus SB-3CT (n=8),  and murine CNS-TB treated with ATT plus doxycycline (n=8). All tissue sections with 7 µm thickness each were stained with hematoxylin prior to LCM-RNA Seq. Target regions including granuloma, inflammatory region and adjacent normal regions were selected and microdissected for library construction. The libraries were sequenced and analysed.


title: Heterogeneity in the metastatic microenvironment: JunB-expressing microglia cells as potential drivers of melanoma brain metastasis progression

city: Tel Aviv

first: Metsada

last: Pasmanik-Chor

address: Ramat Aviv

department: Biology

country: Israel

institute: Tel Aviv University

summary: Reciprocal signaling between melanoma brain metastatic (MBM) cells and microglia reprograms the phenotype of both interaction partners, including upregulation of the transcription factor JunB in microglia. Here, we aimed to elucidate the impact of microglial JunB upregulation on MBM progression. For molecular profiling, we employed RNAseq and RPPA. To test microglial JunB functions we generated microglia variants stably overexpressing JunB (JunBhi) or with downregulated levels of JunB (JunBlo). Melanoma-derived factors, namely Leukemia Inhibitory Factor (LIF), controlled JunB upregulation through JAK/STAT3 signaling. The expression levels of JunB in melanoma-associated microglia were heterogeneous. Flow cytometry analysis revealed the existence of basal-level JunB expressing microglia alongside JunB-highly expressing microglia. Proteomic profiling revealed a differential protein expression in JunBhi and JunBlo cells, including in the expression of microglia activation markers Iba-1 and CD150, and in immunosuppressive molecules SOCS3 and PD-L1. Functionally, JunBhi microglia displayed decreased migratory capacity and phagocytic activity. JunBlo microglia reduced melanoma proliferation and migration, while JunBhi microglia supported the ability of melanoma to proliferate in 3-dimensional co-cultures, that was abrogated by targeting LIFRβ in melanoma-microglia spheroids. Altogether, these data highlight a melanoma-mediated heterogeneous effect on microglial JunB expression, which turns these cells into functionally heterogeneous microglia. Targeting JunB highly-expressing microglia may potentially be utilized for MBM theranostics.

overall_design: Protein lysates from microglia cells were extracted. RPPA was performed, and 492 proteins were identified. Expression data analyzed using Partek Genomics Suite (PGS7.20.0831). Differentially expressed proteins were obtained with cutoff p<0.05 and fold-change difference = 1.25.

first: Orit

last: Adir

last: Sagi-Assif

first: Tsipi

last: Meshel

first: Shlomit

last: Ben-Menachem

first: Dave

last: Hoon

first: Isaac

middle: P

last: Witz

first: Sivan

last: Izraely


title: Effect of removal the skin macrophage on gene expression during aging

city: Guang zhou

first: chen

last: jian

address: No. 1838, North Guangzhou Avenue

institute: Southern Medical University，Nanfang Hospital

summary: To explore the mechanism of macrophage in the regulation of hair aging

overall_design: Comparative gene expression profiling analysis of RNA-seq data for the dermis after macrophage deletion(Three samples in each group are mixed to test and measure)

first: Chen

last: Jian

first: Qu

last: Qian


title: MMPs and NETs are detrimental in CNS-tuberculosis with MMP inhibitor doxycycline improving murine CNS-tuberculosis survival

last: Ong

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

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Refer to individual Series


title: Genome-wide profiling of HPV integration in cervical intraepithelial neoplasia identifies enrichment of genetic elements associated with carcinogenesis

city: Wuhan

first: Jinfeng

last: Xiong

address: 1095 JiefangAnv. Wuhan, Hubei 430030, China

institute: Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China

summary: To investigate differences in the transcriptome between precancerous lesions with HPV integration and precancerous lesions without HPV integration, 15 CIN3 tissue samples (11 integration-positive and 4 integration-negative samples as identified by high-throughput viral integration detection (HIVID)) were collected for RNA sequencing.

overall_design: fifteen CIN3 tissue samples (11 integration-positive and 4 integration-negative samples

first: Hui

last: Shen

first: Da

last: Zhu

last: Wang


title: Mesenchymal Colorectal Cancers Secrete Vesicles With Unique Cargo That Can Be Used For Liquid Biopsy Based Diagnostics [miRNA-seq]

first: Sander

last: van Hooff

institute: AUMC

summary: Tumor-derived extracellular vesicles (TEVs) play a crucial role in cancer progression, metastasis and therapy resistance but their distinct profiles across different cancer stages and molecular subtypes remain underexplored. This study analyzed TEVs from epithelial (CMS2) and mesenchymal (CMS4) subtypes of colorectal cancer (CRC) using six cell lines and clinical samples. Investigation of the cargo of vesicles secreted by the two subtypes revealed significant differences in mRNA, miRNA, and protein profiles between the two subtypes. Notably, CMS2 predominantly secreted smaller, Tetraspanin-8 (TSPAN8) enriched EVs, while CMS4 produced both larger and smaller EVs, enriched in TSPAN4. This underscores the complexity of vesicle heterogeneity between these subtypes. Additionally, we assessed miRNA profiles from plasma-derived bulk TEVs in CRC patients. Our integrative analysis identified a distinct miRNA signature specific to each subtype, indicating that TEVs from CMS2 and CMS4 cells can be detected in circulation and may serve as potential diagnostic tool for CRC.

overall_design: To investigate the miRNA profiles of TEVs derived from CMS2 and CMS4 subtyped cell lines. We used three CMS2 and three CMS4 cell lines.

first: Paris

last: Asif

first: Lauri

last: Borghuis

first: Anita

last: Grootemaat

first: Monique

last: van Eijndhoven

first: Johan

last: de Rooij

first: Nils

last: Groenewegen

first: Nicole

last: van der Wel

first: Arezo

last: Torang

first: Tineke

last: Buffart

first: Michiel

last: Pegtel

first: Jan Paul

last: Medema


title: Disrupted early molar patterning in syngnathia mouse model

status: Public on Oct 14 2025

city: Fuzhou

first: Chensheng

last: Lin

state: Fujian

address: 18 Wulongjiangzhong Avenue, Shangjie Town

department: College of Life Sciences

institute: Fujian Normal University

summary: It is well established that tooth morphogenesis initiates at embryonic day 10.5 (E10.5), characterized by the specification of a single molar germ in each jaw quadrant. However, the developmental mechanisms underlying the establishment of spatially distinct maxillary and mandibular identities along the jaw axis prior to this stage remain elusive. In Wnt1-Cre;pMes-Bmp4 transgenic mice, a pathological model recapitulating human syngnathia (resulting in bilateral jaw fusion), we demonstrate here that BMP4 overexpression in cranial neural crest cells (CNCCs) disrupts early molar patterning with 100% penetrance, manifesting as the formation of only one molar germ within the unilateral fused jaw structure, establishing this model as a valuable tool for investigating the patterning of maxillary and mandibular molar germs during jaw development. Systematic analysis of wild-type embryos revealed the emergence of a single molar primordium in the unilateral first pharyngeal arch (PA1) by E9.0. This primordium undergoes segregation to form the maxillary and mandibular molar between E9.5-E9.75, a process synchronized with the anteroposterior patterning of PA1. Comparative studies with the mutant model indicated that this developmental segregation depends on coordinated migration of CNCC-derived molar primordium and a BMP4/FGF8-dependent gene regulatory network (GRN) at the maxillomandibular junction. Pathologically elevated BMP4 signaling upregulates the p38 intracellular pathway in the mutant jaws, perturbing the finely tuned GRN and disrupting post-migratory CNCC migration. These perturbations collectively lead to defective molar segregation in syngnathia mouse model. Our findings elucidate fundamental mechanisms governing early molar patterning and may provide molecular explanations for syngnathia-associated molar oligodontia in humans.

overall_design: RNA-seq profiles of PA1 tissues from E9.5-E9.75 (25 to 27-somite stage) wild-type and Wnt1-Cre;pMes-Bmp4 embryos

first: Chen

last: Tang

first: Shunting

last: Chen

first: Yan

last: Ji

first: Xinyi

last: Guo

first: YiPing

first: Yanding

last: Zhang

first: Jeremy

last: Green

first: Xuefeng

last: Hu


title: Gestational Low-Protein Diet Impairs Mitochondrial Function and Skeletal Muscle Development by Inducing Immune Responses in Male Offspring

city: Liverpool

first: Atilla

last: ALTINPINAR

address: 6 West Derby street

institute: University of Liverpool

summary: To elucidate the potential underlying mechanisms of protein restriction during gestation, we compared the transcriptomic profiles of GAS muscles from 21-day-old offspring of mothers on a low-protein diet during gestation with those of a control group using RNA-seq analysis.

overall_design: Two weeks prior to mating, nulliparous 8-weeks old female mice were fed either a normal-protein diet (N, 20% crude protein; code 824226, Special Diet Services, UK) or a low-protein diet (L, 8% crude protein; code 824248, Special Diet Services, UK) ad-libitum. Male mice of the same age, which had been fed on a normal-protein diet, were used for mating. The male pups born from both groups were used for this study. New-born offspring were cross-fostered to dams fed normal-protein diet within 24 hours of birth, creating Normal-Normal (NN) and Low-Normal (LN) groups. NN and LN mice were kept on a normal-protein diet after weaning (day 21), generating Normal-Normal-Normal (NNN) and Low-Normal-Normal (LNN) groups of mice respectively.

last: Altinpinar

first: Moussira

last: Alameddine

first: Ufuk

last: Ersoy

first: Ioannis

last: Kanakis

first: Vanja

last: Pekovic-Vaughan

first: Susan

last: Ozanne

first: Katarzyna

last: Goljanek-Whysall

first: Aphrodite

last: Vasilaki


title: Unlocking single cell chromatin accessibility in formalin-fixed paraffin-embedded tissues by scFFPE-ATAC [mouse]

city: Uppsala

first: Xingqi

address: Husargata

department: Immunology, Genetics and Pathology

institute: uppsala university

summary: Formalin-fixed paraffin-embedded (FFPE) samples are the gold standard for clinical tissue preservation. However, single-cell chromatin accessibility profiling technology, a powerful tool to understand epigenetic regulation in human disease, is not available for FFPE samples due to extensive DNA damage. To solve this technical challenge, we introduce scFFPE-ATAC by integrating uniquely designed DNA oligos on Tn5 transposase, split-and-pool ligation DNA barcoding, T7 promoter-mediated DNA damage rescue, and in vitro transcription—the first high-throughput single-cell chromatin accessibility assay for FFPE samples. scFFPE-ATAC enables the simultaneous profiling of up to 64 FFPE samples or spatial regions, with 884,736 cell barcodes in a single run. We successfully applied scFFPE-ATAC to clinical FFPE human lymph nodes stored for 8–15 years and identified distinct epigenetic regulators between tumor centers and invasive edge epithelial cells in human lung cancer. Additionally, we analyzed paired primary and relapsed follicular lymphoma (FL), as well as FL transformed into diffuse large B-cell lymphoma, from clinical samples archived for 2–7 years, revealing key epigenetic drivers of tumor progression. Overall, scFFPE-ATAC enables high-throughput, high-sensitivity chromatin accessibility profiling in long-term archived clinical specimens, paving the way for retrospective epigenetic studies and spatial epigenetic analysis. This technology provides a powerful tool for understanding tumor relapse and metastasis, with broad applications in basic research and personalized medicine.

overall_design: Tagmentation and combinatorial indexing via ligation Before tagmentation, 50,000 purified FFPE single-cell nuclei were centrifuged at 2,000 g for 5 minutes at 4°C and resuspended in 0.1% lysis buffer (10 mM Tris-Cl, pH 7.4, 10 mM NaCl, 3 mM MgCl₂, 0.1% IGEPAL CA-630). The nuclei were then centrifuged again at 2,000 g for 5–10 minutes at 4°C. The nuclei pellet was resuspended in 95 μl of 1× tagmentation buffer (10 mM Tris-HCl, pH 7.5, 5 mM MgCl₂, 10% dimethylformamide, and dH₂O), and 5 μl of barcoded FFPE-Tn5 transposase (2 mM) was added per sample. The reaction mix was gently mixed and incubated at 37°C for 30 minutes at 400 rpm. The reaction was stopped by directly adding EDTA to a final concentration of 30 mM to each sample. All samples were then pooled and centrifuged to obtain a nuclei pellet. Next, three rounds of ligation were performed using the split-and-pool technique in a 96-well plate, with each well containing a ramped specific linker and ligation barcode. Detailed DNA sequences are provided in Supplementary Table 1. In brief, the ligation process was performed as follows: the pooled nuclei were resuspended in 1 mL of 1× NEBuffer 3.1 and mixed with the ligation mix (100 μL of 10× NEBuffer 3.1, 22 μL of 50 mg/mL BSA, 500 μL of 10× T4 DNA ligase buffer, 2,278 μL of ultrapure water, and 100 μL of T4 DNA ligase). A total of 40 μL of this mix was transferred per well, containing 10 μL of the ramped Ligation1 mixture, and incubated at 37°C for 30 minutes at 400 rpm. Then, 10 μL of Blocker-Ligation1 solution (2.64 μL of 100 μM Blocker-Ligation1, 2.50 μL of 10× T4 ligation buffer, and 4.86 μL of ultrapure water) was added per well, and the reactions were incubated at 37°C for 20 minutes at 400 rpm. The samples were then pooled into a 15 mL tube pre-coated with 0.5% BSA, followed by centrifugation at 1,500 g for 10 minutes at 4°C. The second and third ligation reactions were performed identically to the first. After the second and third ligations, blocking reactions were carried out using: 10 μL of Blocker-Ligation2 solution per well (2.64 μL of 100 μM Blocker-Ligation2, 2.50 μL of 10× T4 ligation buffer, and 4.86 μL of ultrapure water); 7.5 μL of Terminator_Ligation3 solution per well (2.64 μL of 100 μM Terminator_Ligation3, 2.50 μL of 0.5 M EDTA, and 2.36 μL of ultrapure water) for reaction termination. After gently mixing, the samples were pooled again into a 15 mL tube pre-coated with 0.5% BSA, followed by centrifugation at 1,500 g for 10 minutes at 4°C. After the third ligation, each nucleus acquired a unique combination of barcodes. Finally, the pellet was resuspended in PBS, nuclei were counted, and the sample was split into tubes containing 30,000–80,000 nuclei for reverse crosslinking.  Reverse-crosslinking, gap filling and in vitro transcription After the third ligation, reverse cross-linking buffer (50 mM Tris-HCl, pH 8, 250 mM NaCl, 1 mM EDTA, and 1% SDS) and 0.22 mg/mL proteinase K were added to each tube, following previous publications 42, 43. The reaction mixture was incubated in a thermomixer at 1,200 rpm overnight at 65°C. The next day, a second proteinase K digestion was performed at 37°C for 2 hours at 850 rpm to ensure complete protein digestion. DNA was then purified using the Zymo ChIP DNA Clean & Concentrator Kit, following the manufacturer’s instructions. For gap filling, an equal volume of NEBNext High-Fidelity 2X PCR Master Mix was added to the eluted DNA, and the reaction mixture was incubated at 72°C for 8 minutes in a thermal cycler. The sample was then purified using the Qiagen MinElute PCR Purification Kit, following the manufacturer’s instructions. Further DNA purification was carried out using the 1X SPRI beads method to remove fragments shorter than 150 bp. Finally, the DNA was eluted in nuclease-free water. For in vitro transcription (IVT), a T7 RNA synthesis kit (New England Biolabs) was used. The following reagents were mixed: • 12.4 μL of template DNA • 2.5 μL of 10X T7 buffer • 1 μL of DTT • 2 μL each of ATP, CTP, UTP, and GTP • 1 μL of T7 mix • μL of RNase inhibitor (Thermo Fisher Scientific)  The IVT mixture was incubated at 37°C overnight (16 hours), followed by DNase treatment and RNA purification using the ZYMO RNA Purification Kit (RNA Clean & Concentrator-5), according to the manufacturer’s instructions. Finally, RNA was eluted in 20 μL of nuclease-free water. The RNA concentration was measured using NanoDrop, and the sample was either immediately used for DNA library preparation or stored at −80°C for future use.  Single cell FFPE-ATAC DNA library preparation Single-stranded cDNAs are prepared from 0.5–1 μg of purified IVT transcripts using the FirstStrand_cDNA oligo (DNA oligo sequences are provided in Supplementary Table 1). The reaction mixture (11.25 μL), containing 500ng-1 μg of RNA, 1 μL of 100 μM random octamer primer stock, and 0.25 μL of RNase Inhibitor (20 U/μL), is heated at 70°C for 3 minutes, then immediately cooled on ice. Next, the master mix is added, which consists of: • 4 μL of 5× First-Strand Buffer • 2 μL of dNTP Mix • 2 μL of 100 mM DTT • 0.25 μL of RNase Inhibitor (20 U/μL) • 0.5 μL SMART MMLV RT The reaction is gently mixed and incubated at 42°C for 60 minutes and then at 70°C for 15 minutes. Next, 2.2 μL of 10× RNase H buffer and 0.2 μL of RNase H (5 U/μL) are added to each reaction and incubated at 37°C for 20 minutes. The RNAClean XP beads (1.8X) are used to purify the cDNA reaction mixture, and the cDNA is eluted in Qiagen Elution Buffer. A sequencing library mix is prepared using purified cDNA, NEBNext high-fidelity 2× PCR master mix, and a unique forward (i5) and reverse (i7) primer combination (DNA oligo sequences are provided in Supplementary Table 1). PCR amplification is run with the following cycling program: • 98°C for 30 seconds • 12 cycles of: • 98°C for 20 seconds • 63°C for 20 seconds • 72°C for 1 minute The sample is purified using the Qiagen MinElute PCR Purification Kit and run on a 6% PAGE gel. The gel region corresponding to 250–800 bp is cut, and the gel is made into fine pieces, which are incubated overnight at 55°C in crush-soak buffer (500 mM NaCl, 1 mM EDTA, 0.5% SDS) at 1,200 rpm. The DNA is purified from the gel using Costar spin-X centrifuge tubes and the Zymo ChIP DNA Clean & Concentrator Kit, following the manufacturer's instructions. A high-sensitivity DNA bioanalyzer chip is used to check the DNA size distribution and quantify the DNA concentration of the DNA library. Finally, deep sequencing is performed with NovaSeq  6000 on the mouse and human scFFPE-ATAC samples for single-cell analysis.

first: xingqi

last: chen

first: Ram

last: Yadav

first: Pengwei

last: Xing


title: DEK function in silencing of Polycomb-target genes

first: Miyuki

last: Nakamura

address: Ulls Grand1

institute: Swedish University of Agricultural Sciences

summary: Chromatin-bound protein DEK is potentially related to a DNA topology change. In Arabidopsis, mutations in the DEK genes enhance the aberrant morphology in the background of lhp1 (Like Heterochromatin Protein) mutant.  LHP1 is responsible for the silencing of Polycomb target genes. We investigated whether defects in DEK genes affect transcriptional profiles in the presence and absence of the lhp1 mutation. Topoisomerase 1α is also known that affects DNA topology and interact with the DEK3 protein (Waidmann et al., 2014). ATRX is known as one of the LHP1 interacting proteins (Wang et al., 2018). By comparing these mutant transcriptomes, we addressed how these mutants are related to each other. We found that there is a synergistic effect on the misregulation of Polycomb target genes in the multiple mutant lhp1;dek2;3;4.

overall_design: Seedling sample of 5 different genetic background including wild-type (Wild-type x3, lhp1 x 3, dek1;2;3;4 quadruple x3, atrx x 3, lhp1;dek2;3;4 x2 biological replications)


title: A low protein diet drives short and long-term improvements in metabolic health in a mouse model of sleeve gastrectomy

city: Madison

first: Cara

middle: L

state: WI

address: 2500 Overlook Terrace

institute: University of Wisconsin-Madison

summary: Despite the largely beneficial impact of bariatric surgery on obesity and metabolic disease, many patients continue to suffer from obesity after surgery and weight recurrence is common. The success of bariatric surgery may be impacted by diet. While bariatric surgery guidelines recommend a high-protein diet following surgery based on the theory that this will preserve lean mass, emerging evidence suggests that both humans and mice are metabolically healthier when consuming low protein diets. Here, we assess the effect of varying dietary protein levels on post-surgical weight loss and weight regain in a mouse model of bariatric surgery, sleeve gastrectomy. We find that a low protein diet optimally drives post-surgical weight loss, boosting energy expenditure and improving blood glucose regulation. Using a multi-omics approach, we identified clusters of differentially expressed genes and metabolites that correlated with these phenotypes and find that diet heavily influences the molecular response of the liver to sleeve gastrectomy. These results suggest that current post-surgical dietary guidelines recommending a high protein diet may limit both the short and long-term benefits of surgery and lead us to conclude that a low protein diet may improve patient outcomes.

overall_design: Five-week-old C57BL/6J male mice were fed a high-fat, high sucrose Western Diet for 12 weeks to induce obesity and glucose intolerance. They were then randomized into weight-matched groups to receive either Sleeve Gastrectomy or Sham operations. To examine the interaction between post-operative dietary protein consumption and metabolism following SG, both SG and Sham mice were further randomized following surgery to either continue consuming a WD, or to consume a lower-fat diet with either High protein (36% calories from protein), Medium (21% calories from protein), or Low protein (7% calories from protein). The three lower-fat diets were isocaloric, with 20% calories from fat and variable carbohydrate to replace any caloric deficit as previously reported. Given the critical role the liver plays in regulating metabolism, we conducted a detailed transcriptomic analysis of liver specimens from sham and SG mice across the three post-operative protein diets in response to feeding. Prior to euthanasia and tissue collection, mice were fasted for 16 hours followed by a 4-hour refeeding period with their respective diets.

first: Julia

last: Illiano

first: Alina

last: Chao

first: Grace

last: Zhu

first: Luiz

last: Lopez

first: Odin

last: Schaepkens

first: Cholsoon

last: Jang

first: Dudley

last: Lamming

first: David

last: Harris


title: Impact of Sirt2 deficiency on downstream TCR signaling in mouse melanoma tumor-infiltrating T lymphocytes

city: Tampa

first: Dongliang

last: Du

address: 111 Magnolia

institute: Moffitt

summary: The T-cell receptor (TCR) signaling pathway is a highly regulated system designed to prevent autoimmunity while ensuring effective immune responses. This regulation is typically mediated through phosphorylation, dephosphorylation, and ubiquitination. Sirt2, is a cytoplasmic NAD+-dependent histone deacetylase with anti-aging and antioxidant properties. Here we reveal acetylation modulated by Sirt2 as a previously unrecognized process controlling TCR signaling. In our study, we found that the loss of Sirt2 in T cells leads to enhanced TCR signaling following T cell activation. Furthermore, we demonstrated that inhibiting Sirt2 in both murine and human tumor-infiltrating lymphocytes restores TCR responsiveness and enhances their anti-tumor activity. This suggests Sirt2 targeting as a potential therapeutic approach for reactivating exhausted T cells within the tumor microenvironment.

overall_design: To investigate the impact of Sirt2 deficiency on pathways downstream of T-cell receptor (TCR) signaling within the tumor microenvironment, we conducted RNA sequencing analysis on tumor-infiltrating T lymphocytes from wild-type and Sirt2 knockout mice bearing B16F10 subcutaneous tumors.

first: Imene

last: Hamaidi

first: Pingyan

last: Cheng

first: Min-Hsuan

last: Wang

first: Alak

last: Manna

first: Min

middle: G

last: Zhang

first: Odesha

middle: O

last: Taylor

first: Bin

last: Fang

first: Bradford

last: Perez

first: Ben

middle: C

last: Creelan

first: Andriy

last: Marusyk

first: Dongjun

last: Shin

first: Tae-Hyun

last: Hwang

first: Anders

last: Berglund

first: Virginia

last: Shapiro

first: Haitao

middle: M

last: Ji

first: Sungjune

last: Kim


title: An intracellular magnetic torque-regulator drives mechano-sequential antitumor immunity [RNA-seq]

first: Rui

last: Gao

overall_design: The underlying mechanism of mechano-stimulation for dendritic cell (DC) activation post low-frequency megneic field (LMF) were explored for the analysis. The RNA sequencing was performed on bone marrow-drived dendritic cells (BMDCs) collected at 6 h after Control, magnetic rod-shaped micromotors (MagRMs), and LMF treatments.

first: Chang

last: Xu

first: Zhenyan

last: Zhu

first: Chen

last: Liu

first: Xueyan

last: Wei

first: Peng

last: Ning

first: Wei

first: Yingze

last: Li

first: Junwu

last: Shen

first: Yajuan

first: Yifan

last: Tong

first: Feng

last: Tao

first: Xiaolei

last: Chen

first: Xiaoyi

last: Jiang

first: Huan

last: Ren

first: Cheng

last: Lv

first: Gustavo

middle: R

last: Plaza

first: Jason

last: Miska

first: Haiming

last: Fan

first: Yichao

last: Tang

first: Yalei

last: Dai

first: Weiwei

last: Yang

last: He

first: Yu


title: Integrated landscape of renal metabolism in domestic cats with spontaneous chronic kidney disease: a multi-omics study

city: Lausanne

first: James

middle: Holzwarth

last: A

address: EPFL Innovation Park

country: Switzerland

institute: Nestlé

summary: Chronic kidney disease (CKD) is the leading cause of mortality in aged cats. After injury, feline kidneys undergo extensive metabolic reprogramming, but a comprehensive evaluation is lacking. We integrate serum metabolome from 15 early stages, 6 late stages CKD and 14 healthy control cats, with renal cortex and medulla transcriptome and proteome. Our analysis reveals spatiotemporal patterns of gene and protein expression changes. In the early stages, there are ~2000 differentially expressed genes in the medulla, but 6 in the cortex. The number in the cortex increases to >4000 in the late stages. We provide evidence of deranged bioenergetics in CKD: circulating fatty acids and acylcarnitines accumulate, while genes and proteins involved in fatty acid transport and oxidation are downregulated. Glucose and pyruvate metabolism is altered in CKD. Impaired glutamine metabolism contributes to both energy deficiency and acid-base imbalance. Additionally, there is a downregulation of redox enzymes, and overexpression of proinflammatory and profibrotic markers in CKD. Finally, hypoxia signaling pathway is upregulated, and negatively correlated with SGLT2 expression. These data unveil profound metabolic abnormalities in feline CKD.

overall_design: 40 samples total, 22 kidney cortex and 18 kidney medulla from 23 separate cats, with 6 non-CKD control, 10 CKD stage 1/2 and 7 CKD stage 3/4 cats.

first: Qinghong

first: Ornella

last: Cominetti

first: James A

last: Holzwarth

first: Stacie

last: Summers

first: Loïc

last: Dayon


title: Maternal obesity programs cardiac remodeling in offspring via epigenetic, metabolic, and immune dysregulation

city: Portland

first: Brett

last: Davis

state: Oregon

address: 3181 SW Sam Jackson Park Rd

institute: Oregon Health and Science University

summary: Maternal obesity during pregnancy significantly increases the offspring’s risk of later-life cardiovascular disease. This study investigated cardiometabolic perturbations utilizing a mouse model of maternal high-fat diet (HFD)-induced obesity that recapitulates metabolic abnormalities observed in humans. We have previously shown that offspring of HFD-fed mothers (Off-HFD) exhibit a progression of obesity, hypertension, dyslipidemia, and metabolic inflexibility when compared with offspring of regular diet-fed mothers (Off-RD). The present investigation of cardiac function further identified significant functional, metabolic, and immune perturbations in adult Off-HFD mice. Specifically, Off-HFD mice presented progressing cardiac hypertrophy with reduced ejection fraction and fractional shortening, increased accumulation of fibrotic tissue, mitochondrial dysfunction, and altered immune complexity including increases in cardiac resident macrophages and decreases in CD4+ and Cd8+ T-cells. While these alterations may not be immediately catastrophic, they likely predispose offspring to heightened sensitivity to nutritional, psychological, or environmental stressors. To explore the timing and mechanisms by which maternal environment impacts offspring cardiac function, we examined DNA methylation in the hearts of newly-weaned Off-RD and Off-HFD mice. This analysis revealed numerous differentially methylated CpGs and regions within genes associated with cardiac development, hypertrophy, mitochondrial function, and immune response. Thus, our study reveals that cardiovascular dysregulations in offspring of obese mothers originate early in development and persist into adult life. These findings uncover potential windows of opportunity for preventive therapy and early therapeutic interventions.

overall_design: RRBS methylation profiling of mouse heart tissue from newly-weaned offspring from High Fat Diet (HFD) or Regular Diet (RD) mothers.

first: Elysse

middle: A

last: Philips

first: Yem

last: Alharithi

first: Tim

last: Wilsoni

first: Cameron

last: Broberg

first: Sheryl

last: Koc

first: Lucia

last: Carbone

first: Jack

last: Rubinstei

first: Susan

middle: B

last: Gurle

first: Sandra

last: Rugonyi

first: Sushil

last: Kumar

first: Alina

last: Maloyan


title: Single-cell analysis uncovers preserved prostate cancer lineages and universally altered pathways in Matrigel-free patient-derived organoids

city: Basel

first: Romuald

last: Parmentier

address: Schönbeinstrasse 40

department: Department of Biomedicine

institute: University Hospital Basel

summary: The application of patient-derived organoids (PDOs) in prostate cancer (PCa) research has been hampered by poor take-rates and benign overgrowth. We highlight the limitations of existing culture conditions and identify extracellular matrix (ECM) composition as a determinant of organoid outcome. Single-cell RNA sequencing (scRNA-seq) reveals that Matrigel-free PDOs preserve patient-specific PCa cells with active AR signaling, and enrich in intermediate cells. In contrast, Matrigel fails to maintain primary PCa cells and produces in vitro basal-like cells divergent from patient samples. Furthermore, we redefine cell type-signatures, identify biomarkers discriminating tumor versus all other cell types, and show that expression of laminin-binding integrins is a hallmark of Matrigel-derived organoids. Finally, integrating previously published datasets with our data, we generate a Prostate PDO single-cell atlas (PPScA). The PPScA captures a spectrum of cellular identities, and reveals pathways universally altered in vitro. Our study provides methodological improvements for short-term cultures and cellular biology insights.

overall_design:
Samples from prostate cancer patients (radical prostatectomy, biopsy, or transurethral resection of the prostate (TUR-P) were collected and dissociated into single cells following established protocols. Single-cell suspensions were used to generate organoids in either Matrigel-free or Matrigel-containing cultures, as described in Dolgos et al. Successful growth, assessed by visual inspection after approximately 7–14 days, was followed by organoid dissociation and fixation for inclusion in multiplexed library preparation batches of four samples. When feasible, single cells from the original patient samples were also fixed for later inclusion in batches. In addition, previously established long-term patient-derived organoid (PDO) lines were cultured under both Matrigel and Matrigel-free conditions using the same methodology, and were also included in the study.  Integrated analyses combining newly generated data with previously published datasets were performed in multiple configurations (e.g., early-passage PDO sets, matched tissue–PDO sets, and broader multi-study atlases). Selected comprehensive integrated datasets are provided as supplementary processed files, and the complete list of files used to assemble all other combinations is detailed in the accompanying README.

Raw files for human/patient samples are being made available in EGA (https://www.ebi.ac.uk/ega/) for controlled access to the personally identifiable sequence data.

Re-used or re-analyzed data are provided as a list of GEO accession numbers when available, or as direct links when the data were not originally deposited in GEO.

first: Robin

last: Dolgos

first: Jing

last: Wang

first: Raphaëlle

last: Servant

first: Arnoud J.

last: Templeton

first: Tobias

last: Zellweger

first: Alastair D.

last: Lamb

first: Kirsten D.

last: Mertz

first: Svetozar

last: Subotic

first: Tatjana

last: Vlajnic

first: Helge

last: Seifert

first: Ashkan

last: Mortezavi

first: Cyrill A.

last: Rentsch

first: Lukas

last: Bubendorf

first: Clémentine

last: Le Magnen


title: Analysis of microRNA-transcript regulatory networks in the hippocampus of the BTBR mouse model of autism

city: Rome

first: Valerio

last: Licursi

address: via degli Apuli, 4

department: c/o Dept. Biology and Biotechnologies "C. Darwin", Sapienza University of Rome

institute: Institute of Molecular Biology and Pathology (IBPM), National Research Council (CNR)

summary: Autism spectrum disorder (ASD) is a heterogeneous neurodevelopmental condition with unknown etiology. Currently, the role of post-transcriptional mechanisms in ASD remains unclear. microRNAs (miRNAs) are small non-coding regulatory RNAs that mediate mRNA destabilization and/or translational repression. To explore the potential role of miRNAs in ASD, we performed miRNA expression profiling in the hippocampus of the BTBR ASD mouse model and age-matched C57BL/6J mice. Alongside, we analyzed the BTBR hippocampal transcriptomic profile to identify differentially expressed transcripts (DETs). By integrating differentially expressed miRNA (DEmiRNA) and DET lists, we discovered mRNA transcripts putative targets of BTBR DEmiRNAs and with an anti-correlated differential expression in the BTBR hippocampus. They represent potential regulatory networks related to gene transcription regulation, and synaptic structure and function relevant for ASD. Thes include miR-200 family members, miR-200a-3p, miR-200b-3p, miR-200c-3p, and miR-429, and the experimentally validated target Zeb2. Moreover, we identified a set of non-canonical interactions characterized by extensive pairing between BTBR DEmiRNAs and DETs, potentially triggering Target-directed miRNA degradation (TDMD). Our findings support a role for miRNA dysregulation in the pathophysiology of ASD.

first: Silvia

last: Gasparini

first: Carlo

last: Presutti

first: Cecilia

last: Mannironi