title: Catalytic inhibition of KAT6/KAT7 enhances the efficacy and overcomes primary and acquired resistance to Menin inhibitors in MLL leukaemia [barcode-seq SG12]

city: Melbourne

first: Mark

last: Dawson

address: 305 Grattan Street

department: Cancer Research

institute: Peter MacCallum Cancer Centre

summary: Targeting the MYST acetyltransferases are an exciting therapeutic opportunity in acute myeloid leukaemia (AML). Here we define the individual and combined contribution of KAT6A, KAT6B and KAT7, in range of AML models showing that although KAT6A/B inhibition is efficacious in some pre-clinical models, simultaneous targeting of KAT7, with the novel inhibitor PF-9363, markedly increases efficacy. KAT7 interacts with Menin and the MLL complex and is co-localised at chromatin to co-regulate oncogenic transcriptional programs. Focusing on MLL fusion oncoprotein (MLL-FP) AML, we show that inhibition of KAT6/KAT7 provides an orthogonal route to targeting Menin to disable the transcriptional activity of the MLL-FP. Combined inhibition rapidly evicts the MLL-FP from chromatin, potently represses oncogenic transcription and overcomes primary resistance to Menin inhibitors. Notably, KAT7 remains an important targetable dependency in acquired genetic/non-genetic resistance to Menin inhibition providing the molecular rationale for rapid clinical translation of combination therapy, particularly in MLL-FP AML.

overall_design: DNA barcode sequencing of spleen and bone marrow samples extracted from mice transplanted with SPLINTR barcoded Molm-13 cells containing an Ubc-SLPI-3xFlag-miRFP680 or Ubc-MCS-miRFP680 vectors and treated with Vehicle, or combination of 5 mg/kg PF-9363 and 10 mg/kg SNDX-5613.

first: Shellaina

last: Gordon

first: Yih-Chih

last: Chan


title: Wnt signaling promotes inflammation and EMT-associated gene expression in mesenchymal TNBC

status: Public on Jul 01 2025

city: Paris

first: Elodie

last: Girard

address: 11-13 rue Pierre et Marie Curie

country: France

institute: Institut Curie

type: Expression profiling by high throughput sequencing

summary: Aberrant activation of the Wnt signaling pathway in triple-negative breast cancer (TNBC) is linked to treatment resistance and recurrence, yet its role in tumoral heterogeneity remains unclear. We developed Wnt reporter cell lines from two mesenchymal TNBC models (MDA-MB-231 and MDA-MB-436) using a Tcf/Lef-eGFP vector and observed intra- and inter-tumoral variation in Wnt activity. Paired Wnt-positive and Wnt-negative TNBC cell lines were established and profiled by RNA-Seq. Integrative analyses revealed that Wnt-positive cells consistently upregulate genes involved in epithelial-to-mesenchymal transition, inflammation (e.g., IL6/JAK/STAT3, TNFα via NF-κB), and extracellular matrix remodeling. A 55-gene Wnt signature common to both nutrient conditions captured these features. Wnt-related gene sets were also enriched in the Mesenchymal-like Immune-Altered subtype of 699 primary TNBC tumors. These findings highlight the role of basal Wnt activity in driving pro-tumorigenic transcriptional programs in TNBC and provide new insight into its contribution to subtype-specific disease features.

overall_design: Two mesenchymal triple-negative breast cancer (TNBC) cell lines, MDA-MB-231 and MDA-MB-436, were stably transfected with the 7TGP Wnt reporter vector, which drives eGFP expression under the control of TCF/LEF-responsive elements. Fluorescence-activated cell sorting (FACS) was used to isolate and expand paired Wnt-positive and Wnt-negative sublines from each model. These reporter-defined cell lines were cultured under two serum conditions: low serum (0.5% FBS) for 3 days and standard serum (10% FBS) for 6 days. Total RNA was extracted from two biological replicates per condition for each cell line for subsequent bulk RNA sequencing.

first: Ramon

last: Garcia-Areas

first: Hamza

last: Lasla

first: Virginie

last: Raynal

first: Amit

last: Kimar Panday

first: Nicolas

last: Servant

first: Pascal

last: Jézéquel

first: Thierry

last: Dubois


title: Catalytic inhibition of KAT6/KAT7 enhances the efficacy and overcomes primary and acquired resistance to Menin inhibitors in MLL leukaemia [scRNA-Seq]

country: Australia

overall_design: Single-cell RNA-sequencing of SPLINTR barcoded Molm-13 cells from the T0 and those harvested from the bone marrow and spleen of mice treated with vehicle or combined PF-9363 and SNDX-5613.

middle: J

middle: A


title: Sex-dependent amelioration of laser-induced choroidal neovascularization by Short-Chain Fatty Acid treatment in a mouse model

city: Berlin

first: Susanne

middle: A.

last: Wolf

state: Berlin

address: Augustenburger Platz 1

department: Klinik für Augenheilkunde

country: Germany

institute: Charite

summary: Age-related macular degeneration (AMD) is a leading cause of blindness worldwide, with a clinical presentation that varies between sexes. In late-stage AMD, choroidal neovascularization (CNV) triggers retinal inflammation and degeneration, processes that are exacerbated by an overactive response of retinal microglial cells. Short-chain fatty acids (SCFAs) have emerged as potential treatments for AMD due to their anti-inflammatory properties. In this study, we investigate the effects of SCFA treatment in a laser-induced CNV mouse model, focusing on sex-dependent differences in disease progression and microglial response. Our findings demonstrate distinct sex-specific patterns in the development of CNV and associated pathological hallmarks. SCFA treatment resulted in a slight increase in density of Iba1+ microglial cells in females at 3 days post-laser (3dpl), while it prevented a massive increase in males at 7 dpl, with both sexes showing enhanced microglial ramification. The dynamics of microglial density were linked to protective effects on CNV, leakage size, and inflammation, which occurred earlier in females and later in males. In vitro assays revealed that SCFAs reduced microglial phagocytosis of retinal debris, suggesting a potential mechanism for their anti-inflammatory action. This study underscores the importance of considering sex-specific responses in the development of AMD treatments, such as SCFAs, and highlights the need for personalized therapeutic strategies.

overall_design: Bulk RNAseq of laser-induced CNV retinas from untreated andSCFA-treated male and female mice.

first: Chufan

last: Yan

first: Caio Andreeta

last: Figueiredo

first: Inga-Marie

last: Pompös

first: Bilge

last: Ugursu

first: Paula

last: Arribas-Lange

first: Sergej

last: Skosyrski

first: Seulkee

last: Yang

first: Petra

last: Althoff

first: Norbert

last: Kociok

first: Antonia M.

last: Joussen

first: Susanne A.


title: Catalytic inhibition of KAT6/KAT7 enhances the efficacy and overcomes primary and acquired resistance to Menin inhibitors in MLL leukaemia [ChiP-seq]

first: Mark

last: Dawson

type: Genome binding/occupancy profiling by high throughput sequencing

overall_design: Chromatin Immunoprecipitation sequencing (ChIP-seq) of Menin and MLL1 in Molm-13 cells follwiung treatment for 6 hours with Vehicle, 2.5 uM PF-9363, 100 nM VTP50469 or combination of 2.5 uM PF-9363 and 100 nM VTP50469 (Combination/Combo).

first: Shellaina

last: Gordon

first: Yih-Chih

last: Chan


title: Catalytic inhibition of KAT6/KAT7 enhances the efficacy and overcomes primary and acquired resistance to Menin inhibitors in MLL leukaemia [barcode-seq SG05]

overall_design: DNA barcode sequencing of spleen and bone marrow samples extracted from mice transplanted with SPLINTR barcoded Molm-13 cells and treated with Vehicle, 5 mg/kg PF-9363, 10 mg/kg SNDX-5613 or combination of 5 mg/kg PF-9363 and 10 mg/kg SNDX-5613.


title: Lifestyle shapes premorbid social and microglial deficits in an Alzheimer’s disease mouse model

city: Dresden

first: Fanny

last: Ehret

address: Fetscherstraße 74

department: Medical faculity

institute: TU Dresden

summary: In this study we used a knock-in mouse model of Alzheimer's disease, App-NL-G-F mice. As a reductionistic model for lifestyle factors, we used a novel enrichment (ENR) paradigm in which App-NL-G-F mice and controls (App-NL) were continuously tracked until 7 months of age. In comparision mice from both genotypes were housed in standard (STD) cages to identify the effect of social housing and exploratory activity. Despite minimal plaque burden and no memory impairment, these mice displayed progressive deficits in social parameters — such as following behavior, social interaction, and exploration – suggesting early behavioral vulnerability. Altered correlations between adult neurogenesis and social parameters emerged by 7 months, linking neural plasticity to premorbid behavior. Morphological analysis of microglia indicated increased microglia coverage in more socially active animals. Exploratory active controls, but not App-NL-G-F mice, exhibit more ramified and less amoeboid microglia, suggesting that this relationship is disturbed by pathology. At the end of the experiments randomly selected animals were processed for single-cell RNA sequencing of hippocampal microglia. In App-NL-G-F mice enrichment dampened interferon-responsive microglia, which typically increase as amyloidosis advances. A shifted immune response was measured by reduced transcription of genes related to antigen processing and presentation and by increased chemokine signaling. Our study demonstrates  interwoven preclinical alterations in an AD mouse model involving multiple domains, including brain, behavior and immune system, and, thus, provides experimental access to the complex interplay between pathology and behavior (at premorbid stages of Alzheimer’s disease).

overall_design: Following enviromental enrichment or standard housing for 7 months, hippocampal microglia from the two differenct genotypes and two different environmental conditions were isolated by flourescence activated cell sorting (FACS) and anlayzed using 10x Genomics scRNASeq.

first: Birte

last: Doludda

first: Hang

last: Liu

first: Sindi

last: Nexhipi

first: Hao

last: Huang

first: Fabian

last: Rost

first: Annette

middle: E

last: Rünker

first: Rupert

middle: W

last: Overall

first: Warsha

last: Barde

first: Michael

last: Sieweke

first: Andreas

last: Dahl

first: Mirko

middle: H

last: Schmidt

first: Gerd

last: Kempermann


title: Xin-Zi-Sheng-Wan Decoction Alleviates Hyperuricemia-Induced Renal Injury by Modulating SPP1-CD44-Mediated Macrophage Infiltration and SRC/FAK/β-Catenin Signaling Pathway

city: Wenzhou

first: Liting

last: Zhu

address: Chashan Street

country: China

institute: Wenzhou Medical University

summary: Aim of the study This study aimed to evaluate the therapeutic efficacy of XZSWD against HUA-induced kidney injury and to elucidate the molecular pathways involved. Methods UPLC-QTOF/MS was used to identify compounds in XZSWD and medicated serum. A HUA mouse model was induced by potassium oxonate (PO) and hypoxanthine (HX) to evaluate the therapeutic effects of XZSWD on renal injury. RNA-Seq and CIBERSORT were applied to analyze gene expression and immune infiltration. Key signaling pathways were validated by qRT-PCR, Western blot, immunofluorescence, and flow cytometry. A THP-1/HK-2 co-culture system combined with SPP1 overexpression and knockdown was used to explore the molecular mechanisms underlying XZSWD’s anti-inflammatory and anti-fibrotic effects. Results 99 compounds were identified in XZSWDS. XZSWD reduced serum uric acid, improved renal function, and suppressed inflammation in HUA mice. RNA-Seq and CIBERSORT revealed downregulation of macrophage-related genes and enhanced mixed M1/M2 polarization. XZSWD inhibited the SPP1–CD44 axis and SRC/FAK/β-Catenin signaling, reducing Epithelial–Mesenchymal Transition (EMT) and Extracellular Matrix (ECM) deposition. In vitro, XZSWD suppressed uric acid-induced EMT and macrophage migration, polarization; these effects were reversed by SPP1 overexpression and enhanced by SPP1 knockdown, indicating SPP1 as a critical therapeutic target. Conclusions The XZSWD alleviates hyperuricemia-induced renal injury by effectively suppressing macrophage-mediated inflammation through coordinated regulation of the SPP1–CD44 axis and the SRC/FAK/β-Catenin signaling network.

first: Linjie

last: Mu

first: Xiaoqiu

last: Ni

first: Feng

last: Wang

first: Xinyi

last: Peng

first: Yujiu

last: Gao

first: Tingting

last: Wei

first: Congcong

last: Zeng

first: Yongheng

last: Bai

first: Fangyan

first: Zhengzhong

last: Yuan

first: Jinguo

last: Cheng


title: MTA1 Rewires Prostate Cancer Transcriptomes: Direct RNA Interactions Govern Oncogenic Gene Expression and Splice Variant Networks [RIP-Seq]

city: Hefei

first: Lei

last: Chen

address: No. 678, Furong Road, Shushan District

institute: The Second Affiliated Hospital of Anhui Medical University

summary: MTA1, a well-characterized epigenetic modifier, plays pivotal roles in cancer progression. However, its involvement in RNA-mediated regulatory mechanisms remains poorly understood. This study unveils MTA1’s dual functionality as a chromatin remodeler and RNA-binding protein (RBP) in prostate cancer (PCa) pathogenesis. By integrating RNA sequencing (RNA-seq) and formaldehyde-crosslinking RNA immunoprecipitation sequencing (fRIP-seq) in PC-3 cells, we demonstrate that MTA1 knockdown dysregulates 1,248 genes and modulates 2,367 alternative splicing events (ASEs). Functional annotation links MTA1-associated differentially expressed genes (DEGs) to extracellular matrix (ECM) remodeling, PI3K-Akt signaling, and hypoxia responses, while ASEs implicate spliceosome activity and RNA processing pathways. fRIP-seq identifies direct MTA1-bound RNAs enriched with GC-rich motifs, with integrative analyses revealing significant overlaps between MTA1 RNA targets and DEGs/ASEs. Our findings establish MTA1 as a multifunctional RBP bridging transcriptional and post-transcriptional networks, offering novel mechanistic insights into PCa progression.

overall_design: This study utilized prostate cancer PC-3 cells as the model system to systematically investigate the role of MTA1 in transcriptional and post-transcriptional regulation through MTA1 knockdown (KD) combined with RNA-seq and fRIP-seq analyses. RNA-seq identified 1,248 differentially expressed genes (DEGs) and 2,367 alternative splicing events (ASEs). Functional analysis revealed that DEGs were primarily enriched in ECM remodeling, PI3K-Akt signaling, and hypoxia response, while ASEs were associated with spliceosome activity and RNA processing. fRIP-seq identified MTA1 directly bound RNAs, which exhibited a preference for GC-rich motifs, and integrative analysis demonstrated significant overlap between MTA1 RNA targets and DEGs/ASEs. These findings confirm MTA1's dual functionality as an RNA-binding protein (RBP) in transcriptional and splicing regulation, providing novel mechanistic insights into prostate cancer progression.

first: Guanlin

last: Qu

first: Fuhao

last: Li

first: Bin

last: Wu

first: Enlai

first: Shen

last: Xu


title: Cell-intrinsic Tim-3 is Required for Optimal CD8 Response to Acute LCMV Infection [TCRseq]

city: Pittsburgh

first: Lawrence

middle: P.

last: Kane

state: PA

address: 5051 Centre Ave.

department: Immunology

country: USA

institute: University of Pittsburgh

summary: Tim-3, or transmembrane immunoglobulin and mucin domain-3, is a type I membrane protein expressed by various immune cell types and which is best known as a negative regulator of anti-tumor immunity. However, Tim-3 also has a co-stimulatory role in T cells under some conditions, via enhancement of PI3K signaling. We hypothesize that Tim-3 signaling enhances CD8+ T cell activation during acute infection and contributes to enhanced CD8+ effector function. To test this hypothesis, we used LCMV-specific TCR transgenic (P14) mice expressing a truncated, non-signaling, allele of Tim-3 or CD8-specific deletion of Tim-3. CD8+ T cell effectors were analyzed by flow cytometry after acute infection with LCMV Armstrong. At the effector stage, endogenous Tim-3+ T cells had increased expression of effector CD8 markers, compared with Tim-3 negative cells. Tim-3 knockout mice also showed a significantly lower number of LCMV-specific T cells. Additionally, endogenous Tim-3+ T cells were also significantly better at cytokine production and had increased cytotoxicity. Tim-3 expression correlated with better cell survival in homeostatic conditions after the virus had been cleared. Bulk RNA sequencing and TCR sequencing revealed that Tim-3-expressing effector CD8+ T cells had a distinct pattern of effector gene upregulation, with increased expansion of a subset of TCR clones. Mechanistically, Tim-3 signals enhances phosphorylation of Foxo1, inhibiting its entry into the nucleus, conferring enhanced effector function. Thus, Tim-3 signaling contributes positively to an effective CD8+ T cell response during acute infection. These findings may lead to a better understanding of CD8+ T cell function in different settings, including in tumors.

overall_design: We FACS sorted endogenous Tim-3lo vs. Tim-3hi antigen-experienced (CD44+CD62L-) CD8+ T cells from spleen at day 8 after LCMV armstrong infection. These cells were processed for bulk RNAseq and TCRseq. In the second experiment, P14 TCR Tg (WT, Tim-3 T2 or Tim-3 KO) were transferred into separate cohorts of WT C57BL/6 mice. These mice were infected with LCMV Armstrong and on day 8, P14 cells were FACS sorted and processed for bulk RNAseq,

first: Priyanka

last: Manandhar

first: Lawrennce

middle: P


title: Single-cell RNA Sequeuncing of Healthy and Diseased Rat Temporomandibular Joint Condyle Cartilage and Disc

first: Alejandro

last: Almarza

address: 335 Sutherland Dr

summary: We have performed single-cell RNA sequencing using 10x Genomics to study the cellular landscape of both healthy and osteoarhtritic temporomandibular joint (TMJ) condyle cartilage and disc tissues. Few studies have performed scRNA-seq on TMJ tissue, and those that have utilized mouse model or human embryo tissues. Here, we are the first to do so using a rat model and using an incisor splint OA-induced model. Understanding the cellular phenotypes within TMJ tissues and how these phenotypes respond to stress is a crucial step towards therepeutic advancements in the field.

overall_design: TMJ condyle cartilage and disc tissue were isolated from the left condyles of healthy(n=8)  rats and four-week-long bite-shifted (via incisor splint) rats (n=8). Disc tissue was isolated by removing it from the condyle, and cartilage tissue was scraped from the condyle using a scalpal blade. Each tissue was pooled separately with their respective experimental group and CMO tagged, resulting in 4 samples sequenced.

first: Sara

last: Trbojevic

first: Xudong

last: Dong

first: Robert

last: Lafyatis

first: Michael

middle: S

last: Gold

first: Juan

middle: M

last: Taboas


title: High-Purity Production of Endothelial Cells from Human Pluripotent Stem Cells

city: Kyoto

first: Koki

last: Yoshimoto

state: Kyoto

department: iCeMS

country: Japan

institute: Kyoto university

summary: Endothelial cells (ECs) are essential for vascular network formation and tissue homeostasis, yet the fields of tissue engineering and vascularized organoid generation still relies heavily on human umbilical vein ECs (HUVECs), which are venous, allogeneic, and difficult to mature fully. Human pluripotent stem cells (hPSCs) offer an autologous, developmentally flexible alternative, but most differentiation protocols require fluorescence-activated cell sorting, limiting scalability. Here we present a streamlined method that produces highly pure ECs directly from human embryonic stem cells (hESCs) without cell sorting. Extending Wnt activation with CHIR99021 to three days maximizes mesoderm induction, and brief Notch blockade with DAPT during specification suppresses smooth-muscle commitment. The result is over 90 % CD31⁺ CD144⁺ ECs that display classic cobblestone morphology, robust DiI-acetylated LDL uptake, and capillary-like sprouting comparable to HUVECs. Bulk RNA barcoding and sequencing segregates the hESC-derived ECs from both HUVECs and undifferentiated hESCs and uncovers an arterial-skewed transcriptome: NOTCH1, DLL4, and CXCR4 are up-regulated, whereas venous markers (EPHB4, NRP2) are reduced. Enrichment of Notch-responsive pathways further supports an arterial-like identity. Although several adult functional genes (e.g., vWF, NOS3) are expressed at lower levels than in HUVECs, the protocol delivers a scalable source of developmentally relevant ECs ideal for vascularizing organoids derived from the same hPSCs and for future applications in drug screening, disease modeling, and cell-based vascular therapies.

overall_design: Expression profiling by high throughput sequencing

first: Shiho

last: Terada

first: Ken-ichiro

last: Kamei


title: MTA1 Rewires Prostate Cancer Transcriptomes: Direct RNA Interactions Govern Oncogenic Gene Expression and Splice Variant Networks [RNA-Seq]

first: Lei

first: Guanlin


title: Profiling Endothelial Cells from Heart Regions and Liver Subpopulations

city: Shanghai

first: Zhicong

last: Liu

state: Shanghai

address: Chinese Academy of Sciences

institute: Center for Excellence in Molecular Cell Science, Chinese Academy of Sciences

summary: Bulk RNA sequencing was performed to investigate gene expression differences among endothelial cells isolated from distinct regions of the mouse heart and tdTomato-positive and -negative endothelial populations in the liver. Heart endothelial cells were collected from adult male wild-type C57BL/6 mice (10 weeks old), while liver endothelial cells were isolated from Ki67-CrexER, Cdh5-DreER, and R26-RL-tdT mice treated with tamoxifen at 8 weeks and collected at 10 weeks. Endothelial cells were isolated by flow cytometry, total RNA was extracted, and mRNA libraries were constructed. Sequencing was performed on the MGISEQ-2000RS platform. This dataset provides a resource for studying regional transcriptomic heterogeneity in cardiac endothelial cells and transcriptional differences in liver endothelial subpopulations.

overall_design: RNA-seq was performed on wild-type cardiac endothelial cells from the apex, LV, VS, and OZ, and on tdTomato⁺/⁻ liver endothelial cells from Ki67-CrexER; Cdh5-DreER; R26-RL-tdT mice.

first: Maoying

last: Han

first: Zhixin

last: Kang

first: Xiuzhen

last: Huang

first: Nicola

last: Smart

first: Xuekun

first: Qiang

last: Shu

first: Bin

last: Zhou


title: Inhibition of AhR improves cortical bone and skeletal muscle function via preservation of neuromuscular junctions

city: Augusta

first: Meghan

middle: E.

last: McGee-Lawrence

state: Georgia

address: 1120 15th Street

department: Department of Cellular Biology and Anatomy

institute: Augusta University

summary: The aryl hydrocarbon receptor (AhR) is proposed to mediate the frailty-promoting effects of the tryptophan metabolite kynurenine (Kyn), which increases with age in mice and humans. The goal of the current study was to test whether administration of pharmacological AhR inhibitors, BAY2416964 and CH-223191, could abrogate musculoskeletal decline in aging mice. Female C57BL/6 mice (18 months old) were treated with vehicle (VEH) or BAY2416964 (30 mg/kg) via daily oral gavage 5 days/week for 8 weeks. A second AhR antagonist, CH-223191, was administered to 16-month-old male and female C57BL/6 mice via intraperitoneal injections (3.3mg/kg) 3 days/week for 12 weeks. While grip strength declined over time in VEH-treated mice, BAY2416964 preserved grip strength in part by improving integrity of neuromuscular junctions, an effect replicated during in vitro studies with siRNA against AhR. Cortical bone mass was also greater in BAY2416964- than VEH-treated mice. Similarly, CH- 223191 treatment improved cortical bone and showed beneficial effects in skeletal muscle, including reducing oxidative stress as compared to VEH-treated animals. Transcriptomic and proteomic data from BAY2416964-treated mice supported a positive impact of BAY2416964 on molecular targets that affect neuromuscular junction function. Taken together, these data support AhR as a therapeutic target for improving musculoskeletal health during aging.

overall_design: RNA-seq profiling of tibialis anterior (TA) muscle and cortical bone shafts in vehicle- and BAY2416964-treated aged mice. For TA muscles, bulk RNA-seq was performed using total RNA extracted by Maxwell RSC simplyRNA Tissue kit; libraries were generated with Illumina Stranded mRNA Prep kit. For cortical bone, RNA was extracted with TRIzol. Sequencing was performed on Illumina platform. Transcript counts were aligned to the mm39 genome using STAR aligner and analyzed using Partek Flow and DESeq2. n=4 mice per group.

first: Kanglun

last: Yu

first: Jie

last: Chen


title: IL-26 from ILCs regulates early-life gut epithelial homeostasis by shaping microbiota composition I

first: Yazan

last: Salloum

address: 11-13 Rue Pierre et Marie Curie

summary: Animals host symbiotic microbial communities that shape gut health. However, how the host immune system and microbiota interact to regulate epithelial homeostasis, particularly during early development, remains unclear. Human interleukin-26 (IL-26) is associated with gut inflammation and has intrinsic bactericidal activity in vitro, yet its in vivo functions are largely unknown, primarily due to its absence in rodents. To examine the role of IL-26 in early life, we used zebrafish and found that gut epithelial cells in il26-/- larvae exhibited increased proliferation, faster turnover, elevated  and DNA damage response, and altered cell population abundance. This epithelial dysregulation occurred independently of the IL-26 canonical receptor and resulted from dysbiosis in il26-/-. Moreover, IL-26 bactericidal activity was conserved in zebrafish, suggesting a potential role of this property in regulating microbiota composition. We further identified innate lymphoid cells (ILCs) as the primary source of IL-26 at this developmental stage. These findings establish IL-26 as a central player in a regulatory circuit linking the microbiota, ILCs, and intestinal epithelial cells to maintain gut homeostasis during early life.

overall_design: RNA-seq profiling of dissected guts from 5-day post-fertilization WT and il26-/- zebrafish larvae.

first: Pedro

last: Hernandez


title: O-GlcNAc Transferase Promotes Eomesodermin-mediated Exhaustion of CD8+ Cytotoxic T Lymphocytes

city: Beijing

first: Tao

last: Gao

state: Beijing

address: No.38 Academy Road, Haidian District

department: Chemical Biology

institute: Peking University

summary: Prolonged exposure to antigens drives CD8+ cytotoxic T lymphocytes (CTLs) to a hypofunctional state known as exhaustion, which compromises their ability to control infections and cancer. Here, we identify O-GlcNAcylation (O-GlcNAc)—a nutrient-sensitive post-translational modification—as a critical regulator for CTL exhaustion. Exhausted CTLs elevated nucleus O-GlcNAc levels, coupled with increased levels of the cellular uridine diphosphate GlcNAc (UDP-GlcNAc) relative to their effector counterparts. Using quantitative chemoenzymatic O-GlcNAcomic analysis, we identified a T-box transcription factor downstream of TCF-1-centered exhaustion transition and sustenance, eomesodermin (Eomes), which undergoes intensive O-GlcNAcylation. Mechanistically, mutation of the primary O-GlcNAc sites on Eomes abolished its glycosylation, but their phosphorylation and interactions with O-GlcNAc transferase (OGT) were unaffected. OGT/Eomes was indispensable for interaction with TCF-1. OGT stabilized Eomes and increased its accumulation in the nucleus, where T-bet-to-Eomes transcriptional transition drives functional exhaustion. These results uncover a novel regulatory relationship between OGT and the TCF-1/Eomes axis in promoting CTL exhaustion and impairing their protective function.

overall_design: RNA-seq profiling of OT1 splenic CD8+ T cells before and after single and repeat stimulation

first: Senlian

last: Hong


title: Preclinical and first-in-human of Purinostat Mesylate, a novel selective HDAC I/IIb inhibitor, in relapsed or refractory multiple myeloma and lymphoma

city: Chengdu

last: Qiu

state: Sichuan

address: No. 17, the Third Part of Renmin South Road

institute: SKLB

summary:
To investigate the effects of the I/IIb HDAC inhibitor, Purinostat Mesylate (PM, also called HL13) on gene expression in multiple myeloma. The MM.1S cells were treated with Ctrl (DMSO) or HL13 (5nM) for 24 hours. Then the cells were collected and total RNA were isolation by Trizol. The expression profiling by array generated following the recommended protocol of Affymetrix GeneChip Human Transcriptome Array 2.0.
To investigate the effects of the I/IIb HDAC inhibitor, Purinostat Mesylate (PM, also called HL13) on gene expression in multiple myeloma. The MM.1S cells were treated with Ctrl (DMSO) or HL13 (5nM) for 24 hours. Then the cells were collected and total RNA were isolation by Trizol. The expression profiling by array generated following the recommended protocol of Affymetrix GeneChip Human Transcriptome Array 2.0.

overall_design: MM.1S cells were treated with Ctrl (DMSO) and HL13 (5 nM) for 24 hours，and LBH589 used for control. And microarray analysis was used to analyze gene expression profiles.

first: Linyu

last: Yang

first: Yiguo

last: Hu

first: Lijuan


title: Single-cell RNA sequencing of PBMCs from patients with myasthenia gravis and healthy controls

city: Chiba

first: Hiroyuki

last: Akamine

address: chuoku inohana 1-8-1

institute: Chiba university

summary: T and B cells from PBMCs were labeled and processed for single-cell capture using the BD Rhapsody system.

overall_design: Peripheral blood mononuclear cells (PBMCs) were isolated from patients with myasthenia gravis and healthy controls. T cells and B cells were enriched and subjected to scRNA-seq. Simultaneously, surface protein expression was profiled using Abseq. In a subset of samples, immune repertoire analysis was also performed. Several samples were multiplexed using antibody-derived tags, and demultiplexing was conducted prior to downstream analyses.

first: T

last: Tanaka


title: Effects of β-catenin deficiency on adipose tissue physiology

city: Ann Arbor

first: Ormond

last: MacDougald

state: MI

address: 2800 Plymouth Rd

department: Molecular & Integrative Physiology

institute: University of Michigan

summary: Objective: Compelling evidence from investigation of preclinical models and humans links canonical Wnt/β-catenin signaling to regulation of many aspects of white adipose tissue development and physiology. Dysregulation of this ancient pathway alters adiposity and metabolic homeostasis. Herein we explore how disruption of adipocyte Wnt signaling affects gene expression and crosstalk between cell types within adipose tissue.  Methods: To investigate mechanisms through which adipose tissue attempts to maintain homeostasis in the absence of β-catenin in adipocytes, we employed standard methods of metabolic phenotyping as well as bulk RNA sequencing, flow cytometry, single-cell RNA sequencing, and isolation of secreted extracellular vesicles.  Results: Our experiments reveal that male, but not female Ctnnb1AdKO mice have an increase in adiposity and insulin resistance. Whereas metabolic processes including fatty acid metabolism were suppressed in adipocytes, mitochondrial metabolism of immune cells was made more efficient, resulting in reduced reactive oxygen species in macrophages and dendritic cells. Deficiency of β-catenin in adipocytes altered the transcriptome of numerous stromal-vascular cell populations including adipose stem and progenitor cells, macrophages, and other immune cells. Homeostasis in white adipose tissue of Ctnnb1AdKO mice is maintained in part by elevated expression of Ctnnb1 mRNA in endothelial cells and in secreted small extracellular vesicles.  Conclusions: Our studies demonstrate the importance of adipocyte Wnt signaling for regulation of lipid and mitochondrial metabolic processes in stromal-vascular cells and adipocytes in adipose tissues. This research provides further support for an intercellular Wnt signaling network with compensatory capability to maintain homeostasis, and underscores importance of Wnt/β-catenin signaling for understanding adipose tissue physiology and pathophysiology.

overall_design: RNA-seq profiling of Ctnnb1AdKO mice and their sex- and age-matched controls (Ctnnb1fl/fl)

first: Romina

last: Uranga

first: Akira

last: Nishii


title: Histone demethylase JMJD3 regulates cell cycle-related gene expression pattern on eargly stage of human erythropoiesis [CUT&Tag]

city: zhengzhou

first: Hengchao

last: Zhang

state: Henan

address: 河南省郑州市高新技术开发区科学大道100号郑州大学新校区

institute: zzu

summary: The Jumonji domain-containing protein D3 (JMJD3) is a histone demethylases that specifically demethylate di-and tri-methyl-lysine 27 on histone H3 (H3K27me2/3) to regulate correlation gene expression. JMJD3 promotes the differentiation and apoptosis of leukemia cells, and inhibits cell proliferation by regulating the methylation level of H3K27. JMJD3 promotes Zebrafish myeloid development and inhibits erythroid development. However, the function and mechanism of JMJD3 in human erythropoiesis still remains largely unknown. Here, we demonstrated that JMJD3 was expressed throughout the erythroid development process, with higher expression in the early stages. During the early erythropoiesis, JMJD3 deficiency caused cell cycle arrest in the G0/G1 phase, which impaired cell growth and erythroid progenitors differentiation, but did not result in cell apoptosis. Cut&tag sequencing and RNA sequencing analysis discovered that JMJD3 regulated the expression of cell cycle and erythrocyte differentiation associated factor lymphoid enhancer-binding factor 1(LEF1) dependently of H3K27me3 modifications. Our findings provide novel insights into understanding of the roles of JMJD3 and H3K27me3 in the regulation of erythropoiesis, and further indicates the significance of epigenetic regulation for the erythropoiesis.

overall_design: The JMJD3/UTX inhibitor GSK-J4 (Catalog #HY-5648B; MedChemExpress) was dissolved in DMSO and administered to cell cultures at a final concentration of 1 and 1.5 μM. After treated with 1μM DZNep for 5 days, we performed a H3K27me3 CUT&Tag sequencing of DMSO and GSK-J4 treated cells.

first: Ying

last: Cheng

first: Xiuyun

last: Wu


title: Effect of supplementation of a minimal culture medium with Raffinose compared with a glucose condition, on the gene expression of a set of bacteria of the intestinal microbiota

city: Jouy-en-Josas

first: Paul

last: Biscarrat

address: 78352 Rue de la Manufacture,

institute: INRAE

summary: The aim of this RNA-sequencing study is to measure differential gene expression in 8 intestinal bacteria (Bacteroides xylanisolvens, Bacteroides thetaiotaomicron, Bacteroides uniformis, Bifidobacterium adolescentis, Bifidobacterium catenulatum, Subdoligranulum variabile and Roseburia intestinalis, Agathobacter rectalis). The data highlight the coordinated action of genes within the same locus involved in the degradation of complex carbohydrates. These loci are well characterized in Bacteroidota species and referred to as polysaccharide utilization loci. In Bacillota and Actinomycetota species, these loci are not so clear-cut, athough the GP-PUL concept has already been proposed. Here we compare the differential gene expression in minimal culture medium supplemented with a complex carbohydrate with a minimal culture medium supplemented with glucose. This differential analysis reveals a source-specific genetic response and a coordinated expression of genes involved in carbohydrate transport, carbohydrate degradation and transcriptional activation of these complex enzymatic machineries.

sample_organism:
Bacteroides thetaiotaomicron
Bacteroides uniformis
Bifidobacterium adolescentis
Bifidobacterium catenulatum
Agathobacter rectalis
Roseburia intestinalis
Subdoligranulum variabile
Bacteroides xylanisolvens

overall_design: To investigate this differential expression, bacterial RNA was extracted after 12h of culture in minimum medium supplemented with one complex carbohydrate or glucose. RNA was extracted using RNAeasy Qiagen kit.

first: Claire

last: Cherbuy


title: Histone demethylase JMJD3 regulates cell cycle-related gene expression pattern on eargly stage of human erythropoiesis [RNA-Seq]

address: 河南省郑州市高新技术开发区科学大道100号郑州大学新校区

overall_design: The JMJD3/UTX inhibitor GSK-J4 (Catalog #HY-5648B; MedChemExpress) was dissolved in DMSO and administered to cell cultures at a final concentration of 1 and 1.5 μM. Experiment procedure before sequencing: 2 million cells were collected and washed twice with PBS. RNA was extracted and used as cDNA input, cDNA library was prepared by Illumina Truseq DNA library kit, and sequenced on Illumina NovaSeq 6000.


title: Anti-injury Mechanism and Molecular Regulatory Networks in Hezuo Pig under Cold Stress

city: Lanzhou

first: Jihong

last: Yan

state: Gansu

address: Yintan Road Subdistrict, Yingmen Village, No. 1

institute: Gansu Agricultural University

type: Non-coding RNA profiling by high throughput sequencing

summary: This study selected cooperative pigs and Baoma pigs as the experimental subjects. By integrating the whole-transcriptome sequencing technology, the ceRNA network related to cold stress-induced anti-injury in the livers of cooperative pigs was screened. The aim was to reveal the molecular mechanism of maintaining liver functional homeostasis in cooperative pigs under cold stress conditions, and to provide new theoretical basis and technical support for cold resistance breeding and liver injury protection of livestock.

overall_design: Using the cooperative pigs and Baoma pigs treated with cold stress for 15 days as the experimental subjects, the normal-temperature groups of both types of pigs were respectively set as controls. High-throughput sequencing technology was employed to analyze the expression differences of mRNA, LncRNA, circRNA and miRNA in the livers of the cooperative pigs and Baoma pigs, to explore their biological functions, and to screen out the key genes and ncRNAs related to cold resistance of the cooperative pigs.

first: Shuangbao

last: Gun

first: Pengfei

last: Wang


title: RNA-Seq in WT and Cav3.1 knock-out female mPFC with or without chronic stress

city: Kumamoto

first: Yasushi

last: Yabuki

state: Kumamoto

address: 2-2-1, Honjo, Chuo-ku

department: Genomic neurology

institute: Kumamoto University

summary: The bulk RNA-seq in WT and Cav3.1 knock-out (KO) female mPFC with or without chronic stress was done to detect gene transcripcts assiciated with chronic stress-induced depressive-like bahaviors in mice.

overall_design: Total RNA was extracted from mPFC in WT and Cav3.1 KO female with or without chronic stress using an RNeasy Kit. The library preparation, sequencing, and Fastq file processing and mapping were conducted as described previously (Matsuo K et al., Cell. 2024;187(24):6835-6848.e20.; Ikenoshita S et al., J Clin Invest. 2023;133(22):e164792.).

first: Norifumi

last: Shioda


title: ERK and PKA signaling drive the evolutionary expansion of the cortex

first: TongYe

last: Fu

address: 138 Yi Xue Yuan Road

institute: Fudan University

summary: The cerebral cortex, the anatomical foundation of human intelligence, underpins our advanced cognition and language. Deciphering the regulatory mechanisms driving its remarkable expansion is essential for unraveling the unique features that define the human brain and species. Here, we demonstrate that ERK and PKA signaling pathways coordinately maintain the neurogenic identity and lineage specification of cortical radial glia (RGs) by suppressing YAP and SHH signaling. Conversely, YAP signaling promotes the RG-ependymal glial cell lineage by inhibiting ERK, PKA, and SHH signaling, while SHH signaling facilitates the generation of cortical tripotential intermediate progenitor cells (Tri-IPCs) from RG by repressing ERK, PKA, and YAP signaling. Cortical Tri-IPCs exhibit a sequential differentiation potential, giving rise to cortical astrocytes, oligodendrocytes, and olfactory bulb interneurons. Importantly, we provide compelling evidence for the evolutionary conservation of these molecular mechanisms between mice and humans. Thus, ERK and PKA signaling in cortical RG establish a mutually reinforcing positive feedback loop, mediated through repressing gliogenic YAP and SHH signaling, which collectively promote human cortical expansion. This study identifies a unifying principle governing mammalian cortical neurogenesis, gliogenesis, expansion, and evolution.

overall_design: The whole mouse cortex was dissected under a microscope for single-cell RNA sequencing (scRNA-seq) analysis. Briefly, mouse embryos and postnatal brains were collected and immediately submerged in fresh ice-cold Hanks' Balanced Salt Solution (Gibco 12175-095). The cortex was then finely dissected, cut into small pieces, and dissociated into a single-cell suspension using a Papain Cell Dissociation Kit (Miltenyi Biotec, catalog no. 130-092-628) according to the manufacturer’s instructions. Single-cell RNA-seq libraries were generated using the Chromium droplet-based sequencing platform (10X Genomics) following the manufacturer’s protocol (manual document part number: CG00052 Rev C). After cellular quality filtering, the number of cells included in the dataset were as follows: E15.0 cortex: control (Map2k1/2 without Cre), 12,128 cells, 2,673 genes/cell; Emx1-Cre; Map2k1/2-dcko mice, 9,291 cells, 2,850 genes/cell. FlashTag labling at E15.5 and FACS at E16.5: control (SuperHippo without Emx1-Cre), 13,194 cells, 2,582 genes/cell; Emx1-Cre; SuperHippo mice, 14,641 cells, 2,616 genes/cell. FlashTag labling at E15.5 and FACS at E16.5: control (wild-type littermate) brains, yielding 14,800 cells, 1,504 genes/cell, hGFAP-Cre; SuperHippo brains, yielding 13,171 cells, 1,544 genes/cell. IUE (dnPKA) labling at E15.0 and FACS at E18.0: hGFAP-Cre; SmoF/F +dnPKA, 11,508 cells, 2,369 genes/cell. FlashTag labling at P0 and FACS P2: control (SmoF/F), 14,292 cells, 2,498 genes/cell; hGFAP-Cre; SmoF/F mice, 7,694 cells, 2,681 genes/cell. FlashTag labling at P0 and FACS P2: control (SuperHippo without hGFAP-Cre), 12,054 cells, 2,258 genes/cell; hGFAP-Cre; SuperHippo mice, 11,539 cells, 2,120 genes/cell. This dataset provides a comprehensive transcriptomic landscape of cortical cells at different developmental stages under various genetic and experimental conditions

first: Zhuangzhi

first: Zhejun

last: Xu

first: Tongye

first: Jialin

last: Li

first: Zhengang

last: Yang


title: The role of GSK3 in BRAF-mutant colorectal cancer

city: Leicester

first: Nicolas

last: Sylvius

address: University Road

department: CBS-Genetics

country: United Kingdom

institute: University of Leicester

summary: Valine to glutamate substitution at residue 600 of the BRAF oncogene (V600EBRAF  mutation) is prevalent in human colorectal cancers (CRCs) with a serrated  histopathology and is thought to be a founder mutation. Using a conditional knock-in  mouse model we have previously demonstrated that V600EBraf drives crypt  hyperplasia in the short term as well as shortened survival linked to increased tumour  burden in the long-term. These phenotypes are associated with induction of gene  signatures for E2F targets, MYC signalling, G2/M transition, canonical WNT signalling,  and cholesterol biosynthesis. Although these gene signatures are reverted by MEK  inhibition, there remains a lack of understanding of the signalling pathways involved,  particularly the mechanism of crosstalk between the MAPK and WNT pathways. Here,  we have examined a role for phosphorylation of GSK3α/β isoforms at residues S21/S9.  By introducing homozygous knock-in mutations for GSK3α/β S9A/S21A onto the  V600EBraf background, we unexpectedly show a marginal effect of these mutations on  further increasing crypt proliferation. However, this impact is lost in the long-term as  there are no significant differences in mouse survival, tumour burden or tumour grade.  Consistently, the GSK3α/β knock-in mutations do not change the transcriptional  programme induced by V600EBraf, except for 3 genes (Ephx4, Eif2b3 Ppp1r13l)  whose expression is significantly altered, potentially contributing to the short-term  increase in crypt hyperplasia. Overall, our data show that therapeutic strategies  targeting GSK3α/β phosphorylation at serines 21/9 are not worthwhile options for  V600EBRAF CRCs.

overall_design: Intestinal tissue samples:  GSK3 alpha and beta isoforms knock-in/knock-in genotype and V600E_knock-in/knock-in genotype.

first: Alessandro

last: Rufini

first: Pooyeh

last: Farahmand

first: Catrin

last: Pritchard

first: Paulina

last: Rzasa


title: Perioperative Testosterone Supplementation Improves Short-Term Outcomes of Anterior Urethroplasty in Hypogonadal Models: Insights from a Preclinical Rabbit Study

first: Jingdong

last: Xue

address: No.389 XInCun Road

institute: tongji university Tongji Hospital

summary: To investigate whether perioperative androgen supplementation could improve surgical outcomes of anterior urethroplasty, particularly in hypogonadal patients, we evaluated the effects and mechanisms of systemic androgen supplementation at different doses in a castrated New Zealand rabbit model of bulbar urethroplasty.

overall_design: Fifteen castrated and five non-castrated adult male New Zealand rabbits were randomized into four groups: non-castrated control (NC), castrated control (0 mg/kg), physiological dose (2.5 mg/kg), and supraphysiological dose (5 mg/kg) of testosterone propionate.

first: Yifan

last: Liu

first: Hanchu

last: Ye

first: Chao

last: Feng

first: Denglong

last: Wu


title: H3K4me3 perturbation in hematopoietic stem progenitor cells by high fat diet-induced obesity in mice

city: Denver

first: Pluto

last: Bio

state: CO

address: 3575 Ringsby Ct Suite 414

institute: Pluto Bio

summary: Obesity is associated with chronic inflammation. H3K4me3 is generally described as a transcription activating histone modification and implicated in trained innate immunity and gene signature during lineage commitment of developing cels. We studied the effect of diet-induced obesity on H3K4me3 marks in hematopoietic stem progenitor cells.

overall_design: Using high fat diet-induced obesity in B6 mice as a model of pre-diabetic obesity, we studied H3K4me3 in LSK (Lin-Sca1+cKit+) and GMP (granulocyte macrophage progenitor: FcRg+CD34+Lin-Sca1-cKit+) hematopoietic stem progenitor cell (HSPC) populations in the bone marrow, compared with sex and age matched low fat diet-fed control mice. Downstream analysis was performed using Pluto Bio software (https://pluto.bio).

first: Norifumi

last: Urao

first: Kentaro

last: Takahashi

first: Ian

last: Davis


title: FOXK2 in Skeletal Muscle Development: A New Pathogenic Gene for Congenital Myopathy with Ptosis

first: Wu

last: Peixuan

address: Dongan road

institute: Fudan university

summary: Congenital ptosis, a genetic disorder involving levator palpebrae muscle dysfunction, is often associated with congenital myopathy. The genetic causes of this condition remain poorly understood. In this study, we identified FOXK2 mutations in six pedigrees with congenital myopathy and ptosis through whole exome sequencing and Sanger sequencing. Zebrafish with foxk2 deficiency exhibited underdeveloped skeletal muscles and reduced mobility, while mice with Foxk2 deletion in skeletal muscle stem cells (MuSCs) showed generalized skeletal muscle abnormalities. Further analysis revealed that FOXK2 deficiency impaired myogenic differentiation in C2C12 cells and disrupted mitochondrial homeostasis in both mouse MuSCs and C2C12 cells. Rescue experiments confirmed the loss-of-function effects of FOXK2 mutation. Coenzyme Q10 treatment improved mitochondrial function and alleviated skeletal muscle development defects in Foxk2-deficient mice. Preliminary omics analysis suggested FOXK2 directly regulates the expression of mitochondrial function-related genes by modulating chromatin accessibility at its binding sites. Our study identifies FOXK2 as a novel pathogenic gene for congenital myopathy with ptosis and highlights its essential role in skeletal muscle development and mitochondrial homeostasis, offering insights for potential diagnostics and therapies.

overall_design: Wild-type C2C12 cells were used to detect the immunoprecipitation of FOXK2 chromatin. Transcriptome profiles and paired chromatin interactions are examined before and after FOXK2 knockout (replicate 3 groups) in C2C12 cells, respectively.

first: Peixuan

last: Song

first: Yang

last: Xiang


title: Reactivation of TET2 reverses vascular smooth muscle cell osteogenic differentiation and prevents medial aortic calcification

city: Sydney

first: Justin

middle: J-L

last: Wong

address: Charles Perkins Centre Lvl4 West, The University of Sydney, Camperdown, NSW, Australia

department: School of Medical Sciences, Faculty of Medicine and Health,

institute: University of Sydney

summary: Cardiovascular diseases often manifest with vascular calcification. Vascular calcification is an active process orchestrated by contractile vascular smooth muscle cell (VSMC) phenotypic switch to an osteoblast-like cell. Here, we identified that the DNA demethylase, Tet2 (Ten-eleven translation 2), safeguards VSMCs from transitioning into the osteogenic lineage and loss of Tet2 promotes development and progression of vascular calcification. Tet2 was among the most significantly downregulated epigenetic markers in calcified aortas. VSMC-specific loss of Tet2 promoted VSMC osteogenic differentiation and enhanced vascular calcification as evidenced histologically, molecularly, and hemodynamically. In vivo studies further indicated that Tet2 inhibits calcification-associated VSMC apoptosis and medial thinning. Notably, calcified regions were enriched in a Trem2hi (triggered receptor expressed on myeloid cells 2) macrophage subpopulation. Intervention studies using high-dose ascorbate to enhance Tet2 enzymatic activity resulted in significantly reduced medial aortic calcification and improved aortic structural integrity in mice. Ascorbate treatment in human aorta organ cultures was sufficient to reduce calcification development in diseased tissues, and restore contractile properties to the calcified aorta. This study highlights the potential clinical impact of modulating Tet2 activity in managing cardiovascular disorders associated with vascular calcification.

overall_design: RNA-seq profiling of the whole aortic tree of vehicle/Vitamin D-treated litter controls mice and mice with VSMC-specific Tet2 knockout

first: Bob

last: Lee

first: Joshua

last: Dunn

first: Cassandra

last: Liang

first: Grace

last: Chensee

first: Renhua

last: Malecki

first: Elizabeth

last: Robertson

first: Gavin

last: Sutton

first: Christopher

last: Stanley

first: Brett

last: Hambly

first: Xiangjian

last: Zheng

first: Paul

middle: G

last: Bannon

first: Wai

last: Tang

first: Renjing


city: Lyon

first: Frédéric

last: Flamant

state: France

address: 46 allée d'Italie

department: IGFL

institute: ENSL

summary: We present a cheap, rapid, and versatile assay called SOSHI-seq (Screening Of Self-transcribed Hormone Inducible response elements coupled to sequencing). The protocol is an adaptation of the STARR-seq method, which allows testing the capacity of thousands of synthetic DNA sequences to act as hormone-response elements. Using the nuclear receptors of thyroid hormone as an example, we show that SOSHI-seq is a suitable complement to ChIP-Seq analysis to identify at genome-wide scale the functional response elements occupied by nuclear receptors in chromatin.

overall_design: SOSHI-seq: a high-throughput screening assay to test the functionality of putative response elements for nuclear hormone receptors. SOSHI-seq: a high-throughput screening assay to test the functionality of putative response elements for nuclear hormone receptors. Banks are prepared with 1000 inserts in a plasmid vector in which sequences able to bind a nuclear receptor are inserted. Plasmid banks are transfected in HEK293 cells and RNA is extracted. Amplicons are prepared, corresponding to the variable part of the plasmid transcript. Amplicons are used to prepare sequencing libraries. Samples differ by : - the plasmid bank that is transfected (bank1 or bank2) - the type of nuclear receptor that is encoded by an additionnal plasmid that is cotransfected with the bank (TRα1, TRβ1 or  LXRα) - the presence or absence of agonist in the culture medium (thyroid hormone , or T3, for TRα1, TRβ1, T-compound for  LXRα). Some of the experiments are duplicated.

first: Shijia

last: Wu

first: Romain

last: Guyot

first: Yanis

last: Zekri

first: Whenzeng

last: Jiang


title: PbAP2-TR plays a role in regulating major transcriptomic changes from early to late trophozoite [PbAP2-TR_DIP-seq]

city: tsu

first: masao

last: yuda

state: mie

address: edobashi 2-174

institute: mie university

summary: To investigate the DNA-binding property of AP2 domains of PbAP2-TR, DNA immunoprecipitation followed by high-throughput sequencing (DIP-seq) analysis was performed. Recombinant AP2 domains 1–2 or AP2 domain 3 fused with maltose-binding protein (MBP) were mixed with the P. berghei genomic DNA fragmented via sonication, and protein-DNA complex was harvested using amylose resin. The obtained DNA fragments were sequenced via the next generation sequencing.

overall_design: DNA fragments encoding the AP2 domains 1–2 or AP2 domain 3 of PbAP2-TR were each cloned into the expression vector pMal-c5X (NEB). Escherichia coli strain DH5α transformed with these plasmids was cultured for 12 h at 37 °C. Next, expression of the MBP-fused protein was induced by adding isopropyl β-D-thiogalactopyranoside (final concentration of 200 nM) in the culture and incubating for 9 h at 25 °C. Recombinant AP2 domains fused with MBP was purified using amylose resin, and eluted with 10 mM maltose solution. Subsequently, the MBP-fused homeodomain was incubated with P. berghei ANKA genomic DNA fragments in binding/washing buffer (10 μM ZnSO4, 2 mM MgCl2, 2 mM Tris-HCl at pH 7.4, 100 mM KCl, and 10% glycerol) for 30 min. The protein/DNA solution was further mixed with amylose resin and incubated for 30 min. After incubation, the resin was washed three times with binding/washing buffer, and bound protein-DNA complexes were eluted with 10 mM maltose solution. A sequencing library was prepared from the DNA fragments and sequenced using Illumina NextSeq. Before their use for DIP, genomic DNA fragments were also sequenced as an input, the raw data of which have been already deposited on Gene Expression Omnibus under the accession number GSE159045.

first: Tsubasa

last: Nishi

first: Izumi

last: Kaneko

first: Shiroh

last: Iwanaga

first: Masao

last: Yuda


title: PbAP2-TR plays a role in regulating major transcriptomic changes from early to late trophozoite [pbap2-g_KO_RNA-seq]

summary: Plasmodium berghei transcriptomes of gametocyte-absent mutant parasites, the pbap2-g-knockout line (pbap2-g(-)), were analyzed from ring to trophozoite stages by high-throughput RNA sequencing (RNA-seq) analysis.

overall_design: Cell cycle of pbap2-g(-) parasites was synchronized, and the parasites were harvested for total RNA extraction at 4, 8, 16 h post-injection. Libraries were constructed from the total RNA and then sequenced by the next generation sequencing.


title: Development of a user-friendly next-generation epigenomic chip adaptable to automation workflows

city: New Haven

first: Timothy

last: McConnell

state: CT

address: 290 Congress Ave

institute: AtlasXomics

summary: This next-generation FlowGel platform for spatial epigenome assays paves the way for a new era in spatial epigenome biology. The platform improvements from this project make the analysis of cell-to-cell interactions accessible while reducing user error, hands-on time, and improving the quality and insight potential. Reducing the complexity of capturing the epigenetic state in tissue will advance our understanding of disease mechanisms, improve the identification of therapeutic targets, and lead to more precise and effective treatments. AtlasXomics manufactures the FlowGel chip by flowing liquid reagents carrying barcodes at elevated temperatures and then cooling to form a stable gel. The end user serially places two stamps onto the tissue, allowing the barcodes to be transferred into the tissue. FlowGel provides significant improvements in usability, performance, and reproducibility. Herein, AtlasXomics spatial CUT&Tag (H3K27ac) analysis of human lymph node using the V2 (96x96, 20 um array) chip.

overall_design: The next-generation FlowGel microfluidic chip was used to perform DBiT-Seq on human lymph node fresh frozen tissue section that was interrogated for H3K27ac marks using CUT&Tag

first: Jeffrey

last: Sabina

first: Gumaro

last: Rojas

first: Silas

last: Decker

first: Jennifer

last: Garbarino

first: Daniel

last: Massucci

first: Katelyn

last: Noronha

first: Machele

last: Ricco

first: Joshua

last: Barnett

first: Molly

last: Wetzel

first: James

last: McGann


title: PbAP2-TR plays a role in regulating major transcriptomic changes from early to late trophozoite [PbAP2-TR_ChIP-seq]

summary: To investigate the genome-wide binding sites of PbAP2-TR and PbMORC in the asexual blood stage of Plasmodium berghei, the chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq) analyses were performed. Parasites expressing GFP-fused PbAP2-TR or GFP-fused PbMORC were generated from the Cas9-expressing parasite (PbCas9), and pbap2-g was disrupted in these parasites (PbAP2-TR::GFPpbap2-g(-) and PbMORC::GFPpbap2-g(-)). Whole blood was harvested from mice infected with these transgenic parasites at 12 h post infection,and the parasites were then subjected to ChIP-seq experiments using anti-GFP antibody. Genome-wide binding sites of PbAP2-TR were determined from the sequence data.

overall_design: PbAP2-TR::GFPpbap2-g(-) and PbMORC::GFPpbap2-g(-) parasites were each cultured for 16 h, and schizonts formed in the cultures were injected into mice. Chromatins associated with GFP-fused PbAP2-TR or PbMORC were IPed with anti-GFP antibody, and DNA fragments purified from the IPed chromoatins were sequenced by the Next Generation Sequencing. Using non-IPed samples as a control (INPUT), binding sites of PbAP2-TR were determined from the sequence data.


title: DynaST-seq: A Transcriptome-wide Spatio-temporal Approach for Profiling Gene Expression Dynamics in Tissues

first: Rui

last: Cheng

address: No. 1630 East Road, Pudong New Area

institute: Shanghai Jiao Tong University

summary: Understanding the molecular mechanisms of spatio-temporal gene expression is crucial for elucidating the dynamic processes that maintain tissue integrity, regulate cellular states, and drive biological responses. Despite recent advances in spatial transcriptomics, most approaches lack temporal resolution, limiting insights into dynamic gene expression. Here, we present DynaST-seq, an advanced spatio-temporal transcriptomics technology that integrates metabolic RNA labeling with spatial transcriptomics, enabling high-resolution, transcriptome-wide profiling of RNA dynamics across space and time. The high-resolution capabilities of DynaST-seq reveal finer tissue structures, such as vasculature, and provide a detailed understanding of cellular composition and RNA dynamics at the single-cell level. With its high sensitivity to newly synthesized RNA, DynaST-seq facilitates early detection of cellular responses to injury, offering detailed insights into processes such as oxidative stress, mitochondrial dysfunction, lipid dysmetabolism, and inflammatory responses during acute kidney injury (AKI). Our analysis identified rapidly responding transcription factors regulating AKI-related biological processes and uncovered enhanced interactions in regions of elevated RNA metabolic activity. This approach offers novel insights into tissue architecture, cellular behavior, and the regulatory networks underlying dynamic biological processes, holding a great promise for a wide range of biomedical applications.

overall_design: As a control, mouse tissues were not labled by 4sU, and analyzed by DynasT-seq. In the experimental group, live mice were labeled by injection, and tissue was collected for DynasT-seq analysis. Tissues from ischemia-reperfusion-treated mice were injection-labeled and analyzed by DynasT-seq.

first: Kun

last: Yin

first: Yafei

last: Qin

first: Wenyao

last: Zhang

first: Pei

last: Liu

first: Chaoyong

last: Yang

first: Jia

last: Song

first: Shichao

last: Lin


title: IGFBP3 Promotes Mammary Stem Cells Activity and Tumorigenesis by Regulating IGF Bioavailability

first: Pan

last: Jingyu

address: Haiding Street

department: School of basic medicine sciences

institute: Tsinghua university

summary: Mammary stem cells (MaSCs) play important roles in mammary gland homeostasis. A better understanding of MaSC activity enhances our knowledge of breast cancer initiation and progression, particularly in the context of triple-negative breast cancer (TNBC). Until now, the molecular mechanisms of MaSCs have remained elusive. Our current research has unveiled a significant enrichment of IGFBP3 expression in normal MaSCs, highlighting its indispensable role in maintaining MaSCs stemness and mammary gland development. Moreover, we show that IGFBP3 promotes breast cancer tumorigenesis and progression. Mechanistically, IGFBP3 regulates IGF1/2 bioavailability under the action of MMPs (such as MMP3), activating the IGF1R receptor and its downstream AKT and ERK1/2 signaling pathways. In clinical samples, high expression of either IGFBP3 or MMP3 is associated with poor prognosis in ER-negative or TNBC patients. Future studies should investigate the potential therapeutic benefit of IGFBP3 in TNBC.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: RNA-seq profiling of mammary stem cells and luminal cells. RNA-seq profiling of SCP28 cells with IGFBP3 KD or not.

first: Jingyu

last: Pan

first: Luyang

last: Zhao

first: Tengjiang

first: Xuan

last: Li

first: Hanqiu

last: Zheng


title: Chromatin accessibility and cell cycle progression are controlled by the HDAC-associated Sin3B protein in murine hematopoietic stem cells [scRNA-Seq]

city: Farmington

first: Harshpreet

last: Chandok

address: 10 Discovery Dr

institute: The Jackson Laboratory

summary: Blood homeostasis requires the daily production of millions of terminally differentiated effector cells that all originate from hematopoietic stem cells (HSCs). HSCs are rare and exhibit unique self-renewal and multipotent properties, which depend on their ability to maintain quiescence through ill-defined processes. Defective control of cell cycle progression can eventually lead to bone marrow failure or malignancy. In particular, the molecular mechanisms tying cell cell re-entry to cell fate commitment in HSCs remain elusive. Previous studies have identified chromatin coordination as a key regulator of differentiation in embryonic stem cells. Here, we utilized genetic inactivation of the chromatin-associated Sin3B protein to manipulate cell cycle control and found dysregulated chromatin accessibility and cell cycle progression in HSCs. Single cell transcriptional profiling of hematopoietic stem and progenitor cells (HSPCs) inactivated for Sin3B reveals aberrant progression through the G1 phase of the cell cycle, which correlates with the engagement of specific signaling pathways, including aberrant expression of cell adhesion molecules and the interferon signaling program in LT-HSCs. In addition, we uncover the Sin3B-dependent accessibility of genomic elements controlling HSC differentiation, which points to cell cycle progression possibly dictating the priming of HSCs for differentiation. Our findings provide new insights into controlled cell cycle progression as a potential regulator of HSC lineage commitment through the modulation of chromatin features.

overall_design: To investigate the function of SIn3B on murine hematopoiesis, conditional knockouts (cKO) of Sin3B in the hematopoietic system were generated by crossing Sin3B-flox mice with Vav1-iCre mice on a C57BL/6 background. Two cKO mice and two wild type mice were humanely sacrificed as adults, and bone marrow was isolated and subjected to cKit enrichment with magnetic beads. Lineage-cKit+Sca-1+ cells were then sorted by FACS and then samples from each genotype were pooled. Cells were stained with TotalSeq Hashtag Oligonucleotide-antibodies against CD45 (BioLegend). Cells were then washed and the 10X Genomics Chromium Single Cell 3' Kit was used to generate single cell suspension and cDNA synthesis/library preparation. Libraries were sequnced on an Illumina NovaSeq and demultiplexed using 10X Genomics CellRanger software. For stress conditions, two mice of the same genotypes as above were given 5-Fluorauracil (100mg/kg) 9 days before harvest.

first: Calderon

last: Alexander

first: David

last: Gregory


title: PbAP2-TR plays a role in regulating major transcriptomic changes from early to late trophozoite [PbAP2-TR_RNA-seq]

first: masao

last: yuda

summary: Plasmodium berghei transcriptomes of pbap2-tr-DiCre, a parasite line in which pbap2-tr is disrupted by DiCre-mediated recombination in the presence of rapamycin, were analyzed at the trophozoite stage.

overall_design: pbap2-tr-DiCre parasites treated with 30 nM rapamycin or DMSO were harvested at 12 h post-injection (hpi), and total RNA was extracted from the parasites. Libraries were constructed from the total RNA and then sequenced by the next generation sequencing.

first: Tsubasa

last: Nishi

first: Izumi

last: Kaneko

first: Shiroh

last: Iwanaga

first: Masao

last: Yuda


title: scDUAL-seq Simultaneously Measures RNA Synthesis and Degradation Rates in Single Cells

city: Xiamen

first: Siying

last: Wang

state: Fujian

address: 422 Siming Road

institute: Xiamen university

summary: The interplay between RNA transcription and degradation tightly regulates the dynamic changes of RNA abundance. To unveil the underlying regulatory mechanism of RNA in shaping gene expression patterns, it is essential yet challenging to profile the coordinated regulation of RNA synthesis and degradation rates at the single-cell level during dynamic programs such as cell differentiation and cell state transition. Herein, we propose scDUAL-seq, a double metabolic RNA labeling-based single-cell RNA sequencing method to simultaneously measure RNA synthesis and degradation rates in thousands of single cells during dynamic processes. In a cortisol response model of colorectal cancer cells, scDUAL-seq identifies the RNA regulatory strategies for 5,930 genes, discovers key regulons driving the strategy transitions, and disentangles the interplay between RNA synthesis and degradation that control distinct strategy transitions during cortisol response. We anticipate that scDUAL-seq will be broadly applied to record RNA kinetic rates and the underlying regulatory mechanism in diverse dynamic processes.

overall_design: mRNA profiles of dual metabolically labeled HCT116 and HEK293T in bulk assays and single-cell levels.

first: Kun

last: Yin

first: Chaoyong


title: Studies on the mechanism of complement protein C4 in the regulation of aging [I]

city: Cheng Du

first: Yan

address: Northern Jianshe Road

institute: University of Electronic Science and Technology of China

summary: Learning and memory capability always decline with age in healthy people, and synapse loss may be an important factor contributing to this reduction. The complement pathway plays a crucial role in synaptic pruning during normal brain development, and the abnormal activation of the classical complement cascade is involved in the pathogenesis of certain neurological disorders. However, none of the past studies elucidated the effects of complement proteins in the aging of the nervous system. Here, we performed transcriptome sequencing in multiple tissues of mice at different ages in an attempt to find important regulators during aging. Our results show that the expression of complement protein C4 in the mouse brain increases with aging. In addition, we constructed different neuronal aging models for transcriptome analysis to identify the regulatory pathways upstream of complement C4, and explored the possible mechanism of complement C4 involved in synapse pruning by co-culture or immunofluorescence methods. This further demonstrates that complement C4 is a potential marker of aging. These findings suggest that NF-κB signaling pathway may contribute to age dependent complement C4 increase, which in turn affect learning and memory function via neuronal synaptic pruning regulation.

overall_design: To investigate the mechanism of aging, we selected  the cortex, hippocampus, kidney of 2m ,10m and 18m C57BL/6 mice, totally 9 mice for mRNA-Sequencing. We descript the universal functional characteristics and tissue-specific changes that occur during aging with various analysis strategies.

first: Ting

last: Liu

first: Lu-nan

last: Luo

first: Yi

first: Jing

last: Yang


title: Reduced antigen-triggered BCR signaling in Fcmr-deficient B cells

city: Shanghai

first: Zichao

last: Wen

address: 130 Dongan Road, Xuhui District

department: school of basic medical sciences

summary: Differential gene expression profile between WT and Fcmr KO splenic B cells after BCR stimulation

overall_design: Purified B cells from WT and Fcmr KO mice were cultured with 10 μg/mL F(ab’)2 anti-IgM and harvested at 1 or 6 h for RNA-seq


title: Provoking the Immunotherapy Efficacy of Renal Medullary Carcinoma [RNA-seq I]

status: Public on Jul 02 2025

city: Houston

first: Haoqiang

last: Ying

state: Texas

address: 1515 Holcombe Blvd

institute: M. D. Anderson Cancer Center

summary: Renal medullary carcinoma (RMC) predominantly afflicts young individuals of African descent harboring sickle hemoglobinopathies such as sickle cell disease (SCD) or sickle cell anemia (SCA).  RMC is refractory to targeted and immune therapy strategies used for other renal cell carcinomas. We have demonstrated that RMC is resistant to immune checkpoint inhibition despite harboring an inflamed tumor immune microenvironment. Hence, novel therapeutic strategies are urgently needed to reverse this resistance. We discovered that CD8+ T cells under SCD condition exhibit altered genomic architecture and ferroptosis, which significantly impairs the anti-tumor immunity of effector T cells. These observations may underscore a fundamental mechanism behind the distinct resistance of RMC resistance to immune checkpoint therapies. By leveraging the use of both mouse SCD model and humanized mice phenotypically representing SCD/SCA, as well as primary CD8+ T cells isolated from patients with SCA, we identified that the SLC7A11 gene is transcriptionally downregulated in the CD8+ T cells of SCA donors via genomic architecture alteration. The reduced expression of SLC7A11 partially accounts for the ferroptosis observed in the CD8+ T cells of SCD disease, via impairing the cystine transportation and synthesis of Glutathione (GSH).  Our findings also demonstrated that treatment using a hydrogen sulfide slow-releasing donor restored the expression of SLC7A11, antagonized the ferroptosis of CD8+ T cells under SCD/SDA condition and restored anti-tumor immunity in vivo. Hence, our research findings provide novel mechanistic insights into SCD and RMC and pave the way to develop innovative therapeutic strategies to sensitize RMC to immune therapies.

overall_design: primary CD8+ T cells were collected from healthy donors and SCA patients for in vitro treatment, RNA extraction and sequencing.

first: Zilong

last: Zhao

first: Benxia

last: Hu

first: Yalan

last: Deng

first: Melinda

last: Soeung

first: Jun

last: Yao

first: Tina

middle: K

last: Nguyen

first: Yaohua

last: Zhang

first: Chengcao

last: Sun

first: Zhou

last: Jiang

first: Jian

last: Gao

first: Andrew

last: Wu

first: Bora

last: Lim

first: Linghua

first: Qing

first: Li

last: Ma

first: Leng

last: Han

first: Dihua

last: Yu

first: Michael

last: Curran

first: Giannicola

last: Genovese

first: Boyi

last: Gan

first: Wenbo

first: Palvos

last: Msaouel

first: Liuqing

first: Chunru

last: Lin


title: An APE1 Inhibitor Reveals Critical Roles of APE1 Redox Function in Pathogenesis of Chronic Rhinosinusitis and Novel Therapeutics

city: Jinan

first: Zhaochao

last: Liu

address: Wenhua West Road

institute: Shandong University

summary: We demonstrated that the Apurinic/apyrimidinic endonuclease 1 (APE1) redox inhibitor, C10, effectively blocked Type 2 inflammation and improved chronic rhinosinusitis (CRS) pathology in a mouse model.

overall_design: Chronic rhinosinusitis (CRS) mice model To establish CRS mice model, mice in CRS group were treated intranasally with 20μg Papain (Sigma-Aldrich, P5306-25MG, USA) which was dissolved in 20μL phosphate buffer saline (PBS) on days 0-2 and 7-11. Drug treatment The treatment dosage of C10 (MCE, HY-19357, CHN) drug solution for intranasal administration in CRS mice were 10 mg/mL, with bilateral administration (10 μl/side, bilateral intranasal instillation). Starting from the initial administration of papain, treatment was continued for 10 consecutive days, with once-daily application. The concentration 10 ng/mL of IL-13 (MCE, HY-P72795, CHN) was used to stimulate Beas-2b and HNECs for 48 hours.


title: Resuscitation from Near Cell Death Unleashes Developmental Cues that Enhance Tissue Repair and Regeneration

city: Hyderabad

first: Nitesh

middle: Kumar

last: Singh

state: Telangana

address: Habsiguda

department: Bioinformatics

country: India

institute: Center for Cellular and Molecular Biology

type: Methylation profiling by high throughput sequencing

summary: Resuscitation from near-cell death conditions is an intriguing but not well-understood cell biology phenomenon. First, we conclusively demonstrate that cells could revive from the early cell death phase evoked by LLOMe and other lysosomotropic agents. The resuscitation phase was found to be a highly orchestrated cellular and molecular process, where the fragmented cell organelles of the death phase fully renewed themselves within a few hours. Remarkably, the initiation of the resuscitation phase was accompanied by the upregulation of pathways of embryonic development, tissue repair, regeneration, and stemness. This was followed by the induction of networks of genes of vital cellular and metabolic pathways. Prompted by in vitro results, we tested whether LLOMe could affect tissue repair, regeneration, and stemness in vivo. Indeed, LLOMe dramatically induced repair in skin wound and corneal alkali burns mouse models. Also, it stimulated the production of hematopoietic stem/progenitor cells in the Drosophila melanogaster, enhanced tadpole tail regeneration in the frog, and promoted axon regeneration in the Caenorhabditis elegans. Fascinatingly, LLOMe treatment of frog tadpoles resulted in accelerated developmental growth. Among others, NF-ĸB signaling was found to be most critical for the revival of cells as well as the regeneration process. Thus, for the first time, we demonstrate that resuscitation from near-cell death conditions is an elegantly programmed process and that this process could be harnessed for therapeutic advances in regenerative medicine.

overall_design: Distribution of 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC) using Oxford Nanopore sequencing with DNA isolated from control and LLOMe treated MEF cells (30mins, 3hours and 24hours).

first: Kollori

last: Dhar

first: Kautilya

last: Jena

first: Reuben

last: Mathew

first: Soumya

last: Kundu

first: Divya

middle: T

last: Sowpati

first: Santosh

last: Chauhan


title: Genome-wide maps of chromatin state in MHCC97L cells

first: Xianghuo

last: He

address: No 270, Dong An Road

institute: Fudan University

summary: We report the application of chromatin immunoprecipitation sequencing (ChIPseq) for high-throughput profiling of histone modifications in MHCC97L cells.

overall_design: ChIP-seq were performed with antibodies against acetylated H3K27 (H3K27ac), H3K4me1, and H3K4me3 to detect the genome-wide maps of chromatin state in HCC cells.

first: Zhiao

last: Chen


title: The mosquito transcriptome is shaped by the individual and interactive effects of warmer temperature and aging

city: Nashville

first: Julian

middle: F

last: Hillyer

state: TN

address: VU Station B, 35-1634

department: Department of Biological Sciences

institute: Vanderbilt University

summary: In mosquitoes, warmer temperature accelerates the progression of senescence, weakening their ability to survive and respond to an infection. Thus, we hypothesized that this phenomenon is driven by changes in gene expression. To test this hypothesis, we employed an RNA-sequencing approach to investigate how warmer temperature and aging, individually and interactively, shape the transcriptome of the female African malaria mosquito, Anopheles gambiae. Warmer temperature increases the expression of genes involved in metabolism and cell signaling but decreases the expression of genes involved in replication, transcription, translation, and protein processing. Aging decreases the expression of metabolism-related genes but increases the expression of protein processing and transcriptional genes. However, early and late aging have opposing effects on many biological processes, with 1-day-old mosquitoes having a largely different transcriptomic profile than older mosquitoes. Importantly, warmer temperature and aging interactively shape gene expression patterns, where, for example, aging-dependent changes in immune gene expression accelerate at warmer temperatures. Altogether, warmer temperature and aging, individually and interactively, alter the mosquito’s transcriptomic profile, reflecting changes in their ability to survive and overcome an infection.

overall_design:
We set out to investigate how warmer temperature and aging, individually and interactively, shape the transcriptome of naïve and immune-induced mosquitoes. To do so, we reared Anopheles gambiae (Giles sensu stricto) at 27°C, 30°C or 32°C, and allocated adult females that were 1, 5, 10, and 15 days post-eclosion to two immune treatment groups: (i) naïve and (ii) immune induced by intrathoracic injection of heat-killed Escherichia coli.
At 6 h post immune treatment we isolated RNA from pools of 15 whole-body female mosquitoes for each temperature-age combination. We completed 5 independent biological trials, resulting in 120 samples derived from 1772 mosquitoes.
We then conducted RNA-sequencing and analysis to identify how immune induction, warmer temperature, aging, and the interaction between warmer temperature and aging, alter gene expression profiles.

first: Lindsay

last: Martin

first: Jordyn

last: Barr

first: Tania

middle: Y

last: Estévez-Lao

first: Jean-Philippe

last: Cartailler

first: Shristi

last: Shrestha

first: Julián


title: Lung tissue-resident memory T cells optimize protection by IL-10 regulation of innate immunity

city: New York

first: Donna

last: Farber

state: NY

address: 701 W168th street

institute: Columbia University

summary: Respiratory viral infections establish tissue resident memory T (TRM) cells in the lung, which provide optimal protection against infectious challenge, though the underlying mechanisms remain incompletely understood. Here, using a mouse model of heterosubtypic influenza infection, we demonstrate that the lung-localized secondary immune response exhibits an attenuated innate immune response compared to the primary response due to TRM-mediated production of the immunoregulatory cytokine IL-10. Using reporter mice, we identify that lung TRM are the predominant producers of early IL-10 in response to viral challenge. Inhibition of early IL-10 signaling in secondary infections results in increased morbidity and lung pathology, and reveals pleiotropic effects of IL-10 in regulating macrophage inflammation and enhancing IFN-g production by memory CD8+T cells. Human influenza-specific TRM isolated from lungs recapitulate robust IL-10 expression associated with augmented effector responses of murine TRM. These data support a dual role for TRM in coordinating in situ secondary responses - augmenting effector responses for robust viral clearance while dampening inflammation to limit tissue damage.

overall_design: There are two experiments of data submitted here. The goals of these experiments were to determine the effect of either (1) previous infection or (2) IL-10 signaling blockade on the lung-localized immune response to subsequent viral infection. To study this, we prepared RNA from whole lung homogenate of mouse lungs undergoing respiratory infection (IAV). For experiment (1), we compared mice undergoing either primary infection or secondary infection. Mice undergoing primary infection are defined as naive mice that are seeing IAV (PR8) for the first time. For secondary infection, we primed mice by infection of one strain of IAV (X31), waited 5 weeks for recovery, and then infected them with PR8 at the same time as mice undergoing primary infection. Lungs were harvested at days 1, 2, and 3 post-infection. Uninfected mice at baseline were also prepared as controls. For experiment (2), we wanted to determine the effects of IL-10 signaling in secondary infections. For these, we prepared memory mice as previously described, and then simultaneously administed an IL-10R blocking antibody at the time of infection. Lungs were collected at 2 days post infection, and uninfected mice were used as control.

first: Alexander

last: Yang


title: Targeting myeloid-PCSK9 enhancing cardio-protection beyond LDL cholesterol-lowering

city: Ulsan

first: Hyowon

last: Ki

address: 50, UNIST-gil, Eonyang-eup

country: South Korea

institute: UNIST

summary: Circulating Proprotein Convertase Subtilisin/Kexin type 9 (PCSK9) levels, known for regulating plasma cholesterol by degrading LDL receptors, correlate with acute myocardial infarction (AMI) risk. Recent studies suggest that PCSK9 improves cardiac function beyond LDL cholesterol levels after cardiac ischemic injury, but the precise role of PCSK9 in this process is not clarified yet. Our study reveals that PCSK9 deficiency induces heterogeneous changes in myeloid cells and macrophages, potentially protecting the heart in AMI regardless of LDL cholesterol homeostasis. Single-cell RNA sequencing identifies PCSK9-dependent cardiac macrophages (PDCMs) as reparative macrophages enriched in activator protein-1 (AP-1) transcription factor (TF)-related pathways. PDCMs enhance VEGF-C secretion and activate Akt signalling in cardiac endothelial cells, improving cardiac remodelling. Indeed, PCSK9 inhibitor-treated coronary artery disease (CAD) patients show increased myeloid cells with PDCM-like features. In sum, targeting myeloid-PCSK9 may offer cardio-protective effects through increased AP-1 activity and VEGF-C expression, providing a novel approach to prevent cardiac dysfunction in AMI.

overall_design: Immune cells were isolated from infarcted hearts of sham- or LAD-operated Pcsk9+/+ and LAD-operated Pcsk9-/- mice. CD45+ immune cells were selectively isolated by Magnetic Activated Cell Sorting (MACS) for a library construction and analyzed by scRNA-seq.

first: Shin H

last: Moon

first: Hyo Won

first: Na Hyeon

last: Yoon

first: Katherine I

last: Chung

first: Huiju

last: Jo

first: Jing

last: Jin

first: Sejin

last: Jeon

first: Seong-Keun

last: Sonn

first: Seungwoon

last: Seo

first: Joowon

last: Suh

first: Hyae Yon

last: Kweon

first: Yun Seo

last: Noh

first: Seung-Jun

last: Lee

first: Chan Joo

first: Nabil G

last: Seidah

first: Sung Ho

last: Park

first: Goo Taeg

last: Oh


title: Pro-cognitive restoration of PV interneuron plasticity in neurodevelopmental disorders

city: Boston

first: Lai Ping

last: Wong

state: MA

address: 185 Cambridge St

department: Molecular Biology

institute: Massachusetts General Hospital

summary: To identify candidate regulators of experience-dependent PV IN plasticity genes or “XPGs” in CA3/CA2 PV INs, we designed an in vivo functional screen in adult mice to isolate translated mRNAs from CA3/CA2 PV INs that receive increased mossy fiber inputs, a trigger for experience-dependent PV IN plasticity.

overall_design: We virally downregulated a molecular brake of mossy fiber filopodia-PV IN connectivity, Ablim3 (Non-target shRNA (shNT) vs. shAblim3-RNA), in DG mossy fibers, to reproduce the effects of experience on mossy fiber filopodial inputs onto PV INs. We used PV-Cre; Rpl22HA/HA mice, in which expression of the hemagglutinin (HA) epitope-tagged ribosomal protein L22 (Rpl22) is restricted to PV INs. Following optimization of signal-to-noise for mRNA isolation from this extremely sparse population of PV INs , we compared mRNAs biochemically isolated from genetically tagged PV INs in the naïve state versus an experience-induced activated state in CA3/CA2. Using this approach we identified differentially expressed candidate XPGs associated with the PV IN activated state.

first: Yu-Tzu

last: Shih

first: Amar

last: Sahay


title: Effect of chronic hypoxia and role of stearoyl-CoA desaturase 1 (SCD1) on the inflammatory response of human primary macrophages

city: Frankfurt

first: Andreas

last: Weigert

address: Theodor-Sern-Kai 7

department: Medicine

institute: Goethe-University

summary: A hypoxic microenvironment in solid tumors frequently is associated with poor outcome due to metabolic adaptation of cells in the tumor microenvironment that supports tumor cells survival and proliferation. However, little is known about how macrophages (MФ) metabolically and phenotypically adapt to chronic hypoxia (CH) and how this in turn affects tumor-associated inflammation. Here, we subjected MФs to CH to compare their inflammatory phenotype with normoxic MФs. In addition, we transfected SCD1, the rate-limiting enzyme of fatty acid desaturation, to investigate its role in MФ adaptation to oxygen shortage and its contribution to the inflammatory function.

overall_design: Human primary macrophages under chronic hypoxia (1% oxygen for 5 days) and normoxia (21% oxygen) with and without stearoyl-CoA desaturase (SCD1) or control and harvested 5 days after transfection (n = 6).

first: Vanesa

last: Guerrero Ruiz

first: Bernhard

last: Brüne


title: Transcriptomic data from a non-human primate model of stroke reproducing endovascular thrombectomy

city: Bron

first: Emmanuelle

last: Canet Soulas

address: bât. B13, Groupement Hospitalier Est, 59 Boulevard Pinel

institute: Universite Claude Bernard Lyon 1

summary: Background and purpose: Rodent models have demonstrated the key pathogenic role of the lectin pathway (LP) of complement activation in brain ischemic injury. Here we assessed the LP activation and its association with stroke severity and outcome in a well-characterized model of stroke mimicking mechanical thrombectomy in non-human primates. Methods: Transient middle cerebral artery occlusion was performed in Macaca fascicularis randomly allocated in cyclosporine A or placebo group. Neurological status was serially assessed. The occlusion lesion, the ischemic penumbra, the final lesion, the lesion growth between occlusion and day 7 as well as the blood-brain barrier permeability using quantitative transfer constant Ktrans ratio and inflammation by quantitative [11C]PK11195 imaging were assessed using positron emission tomography–magnetic resonance (PET-MRI) imaging. Functional residual LP-specific C3b deposition assay was performed in plasma. C3b/iC3b, mannose binding lectin (MBL), MBL associated serine proteases-2 (MASP-2) and interleukin-6 were measured in plasma, cerebrospinal fluid and brain homogenates. Brain tissue was used to perform immunofluorescence and confocal analyses (C3b/iC3b, MBL and MASP-2 and glycocalyx) as well as transcriptomic analysis. Results: Twenty-three subjects were included with a mean age of 6.7 +/- 0.5 years. LP residual activation in plasma collected late after recanalization was significantly decreased compared to baseline conditions. C3b/iC3b peaked at day 7 and this increase correlated with a more severe neurological deficit at day 1 and day 7, a larger final lesion and lesion growth volumes as well as a greater Ktrans ratio. C3b/iC3b, MBL and MASP-2 were found in microvascular thrombi at day 30. At this time point, 760 genes were differentially expressed within the lesion, including 13 LP-related genes. Conclusions: Our results support that the LP of complement activation is involved in thromboinflammation in a non-human primate model of stroke mimicking mechanical thrombectomy in humans.

overall_design: Transient middle cerebral artery occlusion was performed in Macaca fascicularis randomly allocated in cyclosporine A or placebo group. Brains were obtained at day-30. Tissue samples were collected into (“lesion”) and around (“peri lesional”) the ischemic lesion using anatomical landmarks from FLAIR MRI at day 7 as well as in the contralateral mirror area (“contralateral”). RNA samples from snap-frozen data collection were sequenced. Quality and quantity checks were performed by means of Fragment Analyzer (Agilent). Library construction was carried out using NextFlex Rapid Directional mRNA-Seq v2.0 (Bioo-Scientific, PerkinElmer) according to the manufacturer protocol. Libraries were applied to an Illumina flow cell High Output v2.5 and run on the Illumina Nextseq 500 as a single end read for 75 pb. Image analysis and base calling was carried out using the NCS 4.0.1 and RTA 2.4.11 Illumina software suite implemented on the Illumina sequencing machine. Final file formatting, demultiplexing, and fastq generation were carried out using Bcl2fastq v2.17.1.14 and Cutadapt v1.9.1. For trimming the threshold quality score was set to 30, minimum length of 50 bases after trimming, the maximum rate of undetermined bases (“N”s) was set to 30% and adapters were trimmed with 3 bases overlap and a maximum error rate of 10%. Alignment and quantification were performed using Salmon v1.6.0 with Macaca fascicularis (Ensembl Release 106).

first: Laura

last: Mechtouff

first: Domenico

last: Mercurio

first: Juliette

last: Geoffray

first: Alessia

last: Valente

first: Serena

last: Seminara

first: Justine

last: Debatisse

first: Omer Faruk

last: Eker

first: Tae-Hee

last: Cho

first: Océane

last: Wateau

first: Christelle

last: Léon

first: Jean-Baptiste

last: Langlois

first: Guillaume

last: Becker

first: Gabriel

last: Bidaux

first: Norbert

last: Nighoghossian

first: Maria- Grazia De

last: Simoni

last: Canet-Soulas

first: Stefano

last: Fumagalli


title: Harnessing human iPSC-microglia for CNS-wide delivery of disease-modifying proteins in Alzheimer’s disease mice

city: Irvine

first: Mathew

last: Blurton-Jones

state: CA

address: 845 Health Sciences Road

department: Neurobiology and Behavior

institute: University of California, Irvine

summary: Many studies have sought to circumvent the protective qualities of the blood brain barrier to deliver therapeutic proteins to the brain for the treatment of neurodegenerative diseases. Yet, brain specific uptake, peripheral toxicity, off-target effects, and repeated dosing continue to present considerable challenges. As proof of principle, we sought to determine whether human iPSC-microglia (iMG) could be genetically engineered ex vivo to enable pathology-responsive delivery of amyloid-targeting proteins to the brain. Human iPSCs were CRISPR-edited to express the beta-amyloid degrading enzyme neprilysin (NEP) or secreted neprilysin (sNEP) under control of the endogenous CD9 promoter, a microglial gene that is specifically upregulated in plaque-associated microglia. Following adult-transplantation into xenotolerant, amyloid-accumulating mice (5x-MITRG), beta-amyloid peptides, synaptic markers, and off-target substrate levels were examined. To further determine whether increased engraftment of therapeutic microglia could provide additional disease-modifying efficacy, sNEP iMGs were further modified to confer resistance to CSF1R antagonists, enabling brain-wide engraftment of human microglia. Amyloid pathology and synaptic, inflammatory, and neurodegeneration biomarkers were then assessed. We demonstrate that microglia can be engineered to enable pathology-responsive delivery of therapeutic proteins to the brain. NEP and sNEP xenografted microglia reduced levels of amyloid peptides and oligomers, prevented pathology-associated reductions in synaptic markers, and lowered astrogliosis within the hippocampus and overlying cortex of 5x-MITRG mice without off-target degradation. Surprisingly, sNEP delivery by microglia adjacent to the injection site alone was sufficient to achieve almost all disease-modifying outcomes as effectively as CNS-wide sNEP-microglia engraftment. However, significant reductions in amyloid pathology, dystrophic neurites, and perineuronal nets within the plaque-dense subiculum of 5x-MITRG mice was only achieved following CNS-wide microglial replacement. Taken together, these results demonstrate that human microglia can be engineered as a promising new immune cell therapeutic platform to provide widespread and pathology-responsive delivery of biological therapeutics for the treatment of neurodegenerative disease.

overall_design: To identify human microglia populations that are altered in response to in vivo cuprizone treatment, immuno-deficient hCSF1 mice xenotransplanted with human iPSC-derived hematopoietic progenitor cells were treated with cuprizone for four weeks. scRNA-seq was preformed on mice that received normal chow (n=3) or cuprizone chow (n=3).

first: Jonathan

last: Hasselmann


title: Chromatin regulator HELLS mediates SSB repair and responses to DNA alkylation damage

city: Birmingham

first: Wioletta

last: Czaja

state: AL

address: 1918 University Blvd

department: Department of Genetics

institute: University of Alabama at Birmingham

summary: The SNF2 family chromatin remodeler HELLS has emerged as an important regulator of cell proliferation, genome stability, and several cancer pathways. Significant upregulation of HELLS has been reported in 33 human cancer types. While HELLS has been implicated in DNA damage response, its function in DNA repair is poorly understood. Here we report a new regulatory link between HELLS and single-strand break (SSB) repair in cellular responses to DNA alkylation damage. We found that loss of HELLS impairs SSB repair, and selectively sensitizes cells to DNA alkylating agents and PARP inhibitors (PARPi). Furthermore, we found that HELLS is co-expressed with PARP1 in cancer cells, and its loss is synthetic lethal with homologous recombination deficiency (HRD). This work unveils new functions of HELLS in modulating SSB repair and responses to clinically relevant DNA alkylation damage, thus offering new insights into the potential therapeutic value of targeting HELLS in cancer.

overall_design: RNA-seq profiling of wildtype HAP1 cells (C631) and their HELLS KO derivative.

first: Arko

last: Sen


first: Liu

last: Zhaochao


title: RNA-Seq in WT and Cav3.1 knock-out mPFC with or without chronic stress

city: Kumamoto

first: Yasushi

last: Yabuki

address: 2-2-1, Honjo, Chuo-ku

institute: Kumamoto University

summary: The balk RNA-seq in WT and Cav3.1 knock-out (KO) mPFC with or without chronic stress was done to detect gene transcripcts assiciated with chronic stress-induced depressive-like bahaviors in mice.

overall_design: Total RNA was extracted from mPFC in WT and Cav3.1 KO with or without chronic stress using an RNeasy Kit. The library preparation, sequencing, and Fastq file processing and mapping were conducted as described previously (Matsuo K et al., Cell. 2024;187(24):6835-6848.e20.; Ikenoshita S et al., J Clin Invest. 2023;133(22):e164792.).

first: Norifumi

last: Shioda


title: Enhancing single-cell ATAC sequencing with formaldehyde fixation, cryopreservation, and multiplexing for flexible analysis [ATAC-seq]

city: Freiburg im Breisgau

first: Thomas

last: Manke

address: Stuebeweg 51

department: Bioinformatics Core Facility

institute: Max-Planck-Institute of Immunobiology and Epigenetics

summary: The assay for transposase-accessible chromatin using sequencing (ATAC-seq) revolutionized the field of epigenetics since its emergence by providing a means to uncover chromatin dynamics and other factors affecting gene expression. The development of single-cell (sc) applications in recent years led to an even deeper understanding of cell type specific gene regulatory mechanisms. One of the major challenges while running ATAC-seq experiments, bulk or sc, is the need for freshly collected cells for successful experiments. While various freezing methods have already been tested and established for bulk and sc ATAC-seq, quality metrics for preserved cells are rather poor or dependent on sampling time when compared to fresh samples. This makes it difficult to conduct all sorts of complex experiments i.e. with multiple conditions, patients, or time course studies. Especially, accounting for batch effects can be difficult if samples need to be processed at different time points of collection. We tackled this issue by adding a fixation step prior to the freezing method. The additional fixation step improved library quality and yield data comparable to fresh samples. The workflow was also tested on multiplexed sc ATAC experiments, set-up for cost-efficient low input sample handling. Sample cross-in, typically encountered in Tn5-based multiplex approaches, were tackled with a computational procedure specifically developed for this approach.

overall_design: To investigate the performance of our sample preservation and multiplexing strategies, we harvested cultured HepG2 cells. The cells were fixed with 0.1-5% Formaldehyde and subsequently frozen by either flash freezing or cryopreservation. Then, ATAC-seq libraries were created and sequenced to analyze the influence of each preservation combination on data quality and comparability.

first: Hohl

last: Tobias

first: Boenisch

last: Ulrike

first: Manke

last: Thomas

first: Arrigoni

last: Laura