title: Endothelial SRSF1 directly regulates ATF3-KLF2 pathway to promote ischemia-induced angiogenesis

first: Wenting

last: Zhu

summary: Peripheral artery disease is a severe ischemic vascular pathology without effective pharmacological approaches and improving angiogenesis to recover blood perfusion is a promising therapeutic strategy. Endothelial cells (ECs) are the primary cell type contributing to angiogenesis in response to ischemia. However, the molecular mechanisms regulating ischemia-induced angiogenesis remain elusive. We used a discovery-driven approach to identify elevated serine/arginine splicing factor 1 (SRSF1) expression in endothelial cells after ischemia. Proteomic analyses identified endogenous SRSF1 accumulated in ECs of the ischemic muscle and responded to hypoxia. Mice deficient in endothelial SRSF1 exhibited impaired blood flow recovery and impaired new capillary formation following HLI. Importantly, overexpression of SRSF1 enhanced blood flow recovery and angiogenesis after HLI. SRSF1 overexpression facilitated, while SRSF1 knockdown suppressed, the angiogenic function of human ECs. To explore the mechanism the mechanisms contributing to SRSF1-promoted endothelial angiogenesis,we conducted transcriptome RNA-seq searching for downstream molecules and verified it through RNA-pull down and chromatin immunoprecipitation qPCR analysis. Mechanistically, SRSF1 modulated ATF3 alternative splicing, wherein SRSF1 directly bound to ATF3 pre-mRNA and elevated full-length ATF3 transcript at the expense of truncated ATF3Δzip2 transcript, and in turn, ATF3 directly bound to the KLF2 promoter and suppressed KLF2-S1PR1 signaling, through which mediated the SRSF1-promoted endothelial tube formation and angiogenesis. Additionally, alprostadil, the prostaglandin E1 analog, could activate the SRSF1 signaling to improve endothelial angiogenesis in vitro and in vivo. Our findings identified SRSF1 as a novel regulator of ischemia-induced angiogenesis that enhances endothelial angiogenic functions by regulating the ATF3-KLF2/SIPR1 pathway. These results suggest that modulation of endothelial SRSF1 may represent a promising therapeutic approach for treating ischemic vascular diseases.

overall_design: To investigate the mechanisms contributing to SRSF1-promoted endothelial angiogenesis, we induced SRSF1 overexpression in HUVECs by infecting HUVECs with Ad-SRSF1-GFP, while using Ad-GFP-infected HUVECs as controls. We then performed gene expression profiling analysis using data obtained from RNA-seq of HUVECs infected with Ad-SRSF1-GFP or Ad-GFP.

first: Ning

last: Xie

first: Chun-Mei

last: Cao


title: Multi-omics Analysis and Evidence of IL1R1 as a Potential Biomarker for Diabetes-related Intervertebral Disc Degeneration

status: Public on Oct 27 2025

city: Beijing

first: Lixiang

last: Ding

state: None Selected

address: No. 10, Tieyi Road, Yangfangdian, Haidian District, Beijing

country: China

institute: Beijing Shijitan Hospital, Capital Medical University

type: Expression profiling by high throughput sequencing

summary: This study employs 10x Genomics single-cell RNA sequencing (scRNA-seq) to profile cellular changes within the intervertebral discs of diabetic mice. Diabetes mellitus is a systemic inflammatory disease known to drive pathological alterations in multiple organ systems. Here, we specifically investigate the process of diabetes-induced intervertebral disc degeneration. Our findings aim to provide novel insights into the pathogenic mechanisms underlying disc degeneration.

overall_design: We performed single-cell RNA sequencing on nucleus pulposus tissue from ten high-fat-diet-induced diabetic mice. To ensure sequencing data quality, tissues were pooled to create two biological replicate samples: one comprising 5 semaglutide-treated mice and the other comprising 5 untreated control mice.

first: Meng

last: Yi

first: Shijie

last: Liu

first: Yao

last: Zhang

first: Wancheng

last: Lin


title: Dedifferentiation-Driven Oncogenic Stemness Promotes Tumor-Sustaining Adaptability in the Intestinal Epithelium

city: Hoboken

first: ansu

last: perekatt

state: NJ

address: 507 River street

department: CCB

country: USA

institute: Stevens Institute of Technology

summary: Intestinal tumors can arise either from mutations sustained in Lgr5⁺ stem cells (bottom-up) or through dedifferentiation of lineage-committed epithelial cells (top-down). Using a Smad4 loss-of-function and β-catenin gain-of-function mutant mouse model, we show that dedifferentiation-derived stem cells sustain tumorigenesis more effectively than mutant Lgr5⁺ cells in the endogenous crypts. Bulk and single-cell RNA sequencing revealed transcriptional reprogramming and the emergence of oncogenic villus-derived stem-like cells. Complementary in vivo and organoid studies suggest that dedifferentiation-derived oncogenic stem cells possess superior tumor-initiating and sustaining capacity.

overall_design: RNA was harvested from the intestinal villi epithelia of both wildtype (control) mice and Smad4 loss-of-function and β-catenin gain-of-function conditionally mutant mice. Samples were collected from three biological replicates for each treatment.

first: Kylee

last: Zgeib

first: Thompson

last: Hui

first: Simon

last: Garcia

first: Zahra

last: Hashemi

first: Shima

last: Nejati

first: Amartya

last: Singh

first: Crystal

last: Lim

first: Atharva

last: Inamdar

first: Binfeng

last: Lu

first: Ansu

last: Perekatt


title: DNMT3A:R882H Is Not Required for Disease Maintenance in Primary Human AML, but Is Associated With Increased Leukemia Stem Cell Frequency [RNA-Seq]

city: Stanford

first: Thomas

last: Koehnke

state: CA

address: 265 Campus Drive

institute: Stanford University

summary: Genetic mutations are being thoroughly mapped in human cancers, yet a fundamental question in cancer biology is whether such mutations are functionally required for cancer initiation, maintenance of established cancer, or both. Here, we study this question in the context of human acute myeloid leukemia (AML), where DNMT3A:R882 missense mutations often arise early, in pre-leukemic clonal hematopoiesis, and corrupt the DNA methylation landscape to initiate leukemia. We developed CRISPR-based methods to directly correct DNMT3A:R882 mutations in leukemic cells obtained from patients. Surprisingly, DNMT3A:R882 mutations were largely dispensable for disease maintenance. Replacing DNMT3A:R882 mutants with wild-type DNMT3A did not impair the ability of AML cells to engraft in vivo, and minimally altered DNA methylation. Taken together, DNMT3A:R882 mutations are initially necessary for AML initiation, but are largely dispensable for disease maintenance. The notion that initiating oncogenes differ from those that maintain cancer has important implications for cancer evolution and therapy.

overall_design: Bulk RNA-Seq of human leukemic blasts 4 months after re-mutation or correction of DNMT3A:R882H

last: Köhnke

first: Daiki

last: Karigane

first: Ravindra

last: Majeti


title: CRISPR-free RNA Base Editing Mediated PTC-readthrough Restores Hearing in Humanized Mice with Otof Nonsense Mutation [PRAISE]

first: Wenqing

last: Liu

email: puyulwq@163.com, liuwq21@gamils.com

address: No. 5 Summer Palace Road, Haidian District, Beijing

institute: Peking University

summary: Today, the gene editing for hereditary deafness is becoming a promising field with global interest. Among them, AAV-mediated otoferlin overexpression is undoubtedly the most successful representative of deaf-related gene therapy. However, it remains challenging to realize physiological, endogenous pattern of otoferlin expression. Here, we firstly generated a humanized homology Otof c.1315C>T (p.R439X), equivalent to OTOF c.1273C>T (p.R425X) identified in human with profound hearing loss, nonsense mutation-caused deaf mice model. We then delivered the ‘RESTART v3’ system, which is a CRISPR-free RNA base editor for nonsense mutation suppression, into the cochlea of the mice model. We accomplished a physiological manner of otoferlin expression; moreover, the edited premature termination codon is precisely reversed back to the original amino acid. Using multiple assays to estimate hearing restoration, we identified a significant reduced thresholds of auditory brain response (ABR) and statistical enhanced amplitudes of behavioral auditory startle reflex (ASR) in the treated mice with the RNA editing system. Thus, our study presented a successful CRISPR-free RNA editing approach to significantly restore hereditary deafness carrying humanized Otof c.1315C>T (p.R439X) non-sense mutation, providing a great promise for future clinical translation.

overall_design: we examined the global modification level of cochlear tissues with or without RESTART treatment to evaluate the off-target events.


title: A novel subtype of astrocytes critical for HIV-associated pain pathogenesis

city: Stony Brook

first: Shao-Jun

last: Tang

state: New York

address: 101 Nicolls Road

institute: Renaissance School of Medicine at Stony Brook University

summary: Pathological pain is a frequent complication in HIV patients, yet the underlying mechanism remains elusive, and effective therapeutic targets have not been identified. Reactive astrocytes are specifically activated in the spinal dorsal horn (SDH) of HIV patients with pathological pain and required for HIV-associated pain development in mouse models. These findings suggest that reactive astrocytes play a crucial role in the pathogenesis of HIV-associated pain. However, due to the heterogeneity of reactive astrocytes, the pathogenic subtype remains unknown. In this study, we used single-nucleus RNA-seq (snRNA-seq) transcriptomic analysis to identify a novel subtype of HIV-pain-associated astrocytes (HIPAs) in the lumbar spinal cord of HIV-1 gp120 transgenic models and in the spinal cord of HIV patients. HIPAs express galectin 3 (Gal3) and exhibit transcriptomic signatures of phagocytosis and inflammation. We demonstrated that HIPAs phagocytose neuronal components and are associated with neuronal degeneration. We also found that knockout (KO) of Gal3 in gp120 transgenic mice reduced the number of HIPAs. Furthermore, Gal3 KO inhibited the expression of mechanical allodynia and HIV pain-related pathogenic processes, including neuronal degeneration and neuroinflammation, in gp120 transgenic mice. Our data collectively suggest that HIPAs are a novel Gal3-expressing astrocytic subtype that plays a critical role in gp120-induced pathogenesis in the spinal pain neural circuit. Targeting HIPAs may provide a potential therapeutic strategy for treating HIV-associated pain.

overall_design: All cell types from the lumbar spine of wild-type, gp120Tg, gal3KO, and gp120Tg/gal3KO mice were isolated and analyzed using snRNAseq

first: Junying

last: Zheng

first: Michael

last: Spurgat

first: Subo

last: Yuan

first: Xin

first: Benjamin

last: Gelman


title: Regulated TRESLIN-MTBP loading governs initiation zones and replication timing in human DNA replication [LD-OK-seq]

city: Mishima

first: Masato

last: Kanemaki

state: Shizuoka

address: Yata 1111

department: Department of Chromosome Science

country: Japan

institute: National Institute of Genetics

summary: Replication origins in human cells form clusters ranging from tens to hundreds of kilobases called initiation zones (IZs), typically located in intergenic regions between active genes. On a larger megabase scale, chromosomes replicate in a temporally defined replication timing (RT) pattern, where euchromatic regions replicate early, and heterochromatic regions replicate late. Although the stochastic model of IZ firing with a temporally regulated limiting factor can explain RT formation, this limiting factor in human cells remains unclear. To investigate the relationship between IZ and RT, we mapped the temporal firing pattern of IZs and examined the genome-wide distributions of replication licensing and firing factors in human cells. We identified TRESLIN-MTBP as the key limiting firing factor for replication initiation. Its loading onto phosphorylated MCM2–7 double hexamer (MCM-DH) is controlled by the opposing phosphorylation events on MCM-DH by Dbf4-dependent kinase and RIF1-Protein Phosphatase 1, which ultimately determine IZs and establish RT.

overall_design: Ligase-depleted Okazaki-sequencing (LD-OK-seq) was performed using HCT116 cells. LD-OK-seq experiments were conducted under both asynchronous and synchronous conditions. Synchronous LD-OK-seq was carried out at six time points across the S phase. The same LD-OK-seq experiments were also performed using HCT116 cells with RIF1 knockout (RIF1-KO). Additionally, RIF1-KO cells with reintroduction of either wild-type RIF1 (WT-RIF1) or a RIF1-ppC mutant were used. Furthermore, cells overexpressing TRESLIN-MTBP were also subjected to LD-OK-seq.


title: Regulated TRESLIN-MTBP loading governs initiation zones and replication timing in human DNA replication [RNA-Seq]

overall_design: RNA-seq were conducted using control HCT116 (LIG1/3-mAID2) and RIF1-KO HCT116 cells.


title: Inhibition of CDK10 triggers nucleic acid sensors-mediated anti-tumor immunity [RNA-Seq]

city: wuhan

first: Gaoshan

last: Xu

state: -- Please Select --

address: wuhan

institute: wuhan university

summary: Cdk10 knockout significantly reduced CT26 tumor growth in immunocompetent BALB/c WT mice.To elucidate the molecular mechanisms by which Cdk10 suppresses anti-tumor immunity, we conducted transcriptomic analyses on WT and Cdk10 KO CT26 cells

overall_design: Total RNA was isolated from sgControl and sgCdk10 CT26 cells using TRIzol Reagent, following the manufacturer's instructions.


title: Regulated TRESLIN-MTBP loading governs initiation zones and replication timing in human DNA replication [IP-seq]

type: Genome binding/occupancy profiling by high throughput sequencing

overall_design: TRESLIN-IP was performed using a cell line expressing 3xFLAG-TRESLIN and GINS4-mAID2. Cells were synchronized with nocodazole and released into early S-phase. 5-Ph-IAA or DDKi was added to the culture medium 2 hours after nocodazole release. A control experiment was conducted using a cell line without any FLAG-tagging.


title: Inhibition of CDK10 triggers nucleic acid sensors-mediated anti-tumor immunity [CRISPR screening]

summary: Given the high druggability of kinases, we utilized a targeted library focused specifically on the kinome for in vivo CRISPR/Cas9 knockout (KO) screening

overall_design: To conduct the screen, lentiviruses carrying the kinome-targeting sgRNAs were used to infect mouse CT26 cancer cells. Following a 7-day selection process to remove uninfected cells, the transduced cells were divided into three groups: in vitro two-dimensional (2D) culture, and in vivo experiments through subcutaneous injection to establish xenograft tumors in either immunodeficient BALB/c nude mice (BALB/c Nude) or immunocompetent wild-type BALB/c mice (BALB/c WT).


title: Off-Target RNA editing induced by RECODE in Mouse Quadratus Femoris Muscle

status: Public on Oct 26 2025

city: Fukuoka City

first: Soichi

last: Ozaki

state: Fukuoka

address: West Zone 5 #430, Kyushu University, 744 Motooka, Nishi-ku,

institute: EditForce, Inc.

summary: Programmable guide-RNA-dependent cytidine deaminase tools to convert cytidine-to-uridine (C-to-U) have limitations such as low editing efficiency, limited targetable sequence flexibility, and off-target RNA editing. Here, we report the editing activity of a novel guide-free C-to-U editing tool, named RECODE, based on RNA-binding pentatricopeptide repeat proteins. We show that RECODE, delivered via AAV, is functional in mice, demonstrating high editing efficiency and low off-target effects in quadratus femoris, a skeletal muscle. These results suggest its therapeutic potential for various diseases.

overall_design: RNA-seq to identify global C-to-U substitution events induced by RECODE-WW[F75F] and RECODE-PG[I83I] in the transcriptome of mouse quadratus femoris two months after infection.


title: Dominant intragraft plasma cells targeting bilirubin implicate local heme catabolism in human cardiac allograft vasculopathy

city: New York

first: Sarah

middle: B

last: See

address: 1300 York Ave

institute: WCM

summary: Cardiac allograft vasculopathy (CAV) is invariably associated with immune infiltrates around affected coronary arteries, including antibody-producing plasma cells (PC). We used single-cell-RNA-sequencing and IGHV profiling to characterize these infiltrating PC. We then generated 37 recombinant monoclonal antibodies (mAb) using rearranged immunoglobulin gene sequences from dominant intragraft PC clones as well as 24 control mab from peripheral blood PC. Unexpectedly, a majority (21/37) of mAb derived from intragraft PC, but none of the control mAb derived from blood PC, reacted to bilirubin, a by-product of heme degradation. Furthermore, we observed the deposition of bilirubin within lymphocytic aggregates of cardiac grafts with CAV but not in healthy heart tissue. Bilirubin also accumulated in the cytoplasm and the nuclei of smooth muscle cells in the tunica media of coronary arteries with CAV lesions. Lastly, expression of heme-oxygenase-1 and biliverdin reductases in graft-infiltrating macrophages as well as presence of Fe2+ ion in the media of arteries with hyperplasia further supported the implication of heme catabolism in CAV. Collectively, our findings suggest a key role for bilirubin in CAV. The enrichment of PC clones reactive to this metabolite within graft infiltrates indicates that it acts as a prominent target of local antibody responses.

overall_design: Cardiac tissue from explanted hearts with CAV were collected, dissociated and sorted for scRNA-seq profiling

first: Talita

last: Aguiar

first: Max

last: Dietzel

first: Mattea

last: Ausmeier

first: Hang

middle: T

last: Nguyen

first: Shunya

last: Mashiko

first: Laura

last: Donadeu-Casassas

first: Hector

last: Cordero

first: Poulomi

last: Roy

first: Lorea

last: Roson

first: Charles

middle: C

last: Marboe

first: Matthias

last: Szabolcs

first: Maryjane

last: Farr

first: Jose

last: González-Costello

first: Aleix

last: Olivella

first: Yoshi

last: Naka

first: Koji

last: Takeda

first: Rodica

last: Vasilescu

first: Kevin

last: Clerkin

first: Gilles

last: Benichou

first: Joren

last: Madsen

first: Rodney

middle: G

last: King

first: Oriol

last: Bestard

first: Evan

middle: P

last: Kransdorf

first: Emmanuel

last: Zorn


title: CRISPR-free RNA Base Editing Mediated PTC-readthrough Restores Hearing in Humanized Mice with Otof Nonsense Mutation [TargetSeq]

first: Wenqing

last: Liu

sample_organism:
Homo sapiens
Mus

platform_organism:
Homo sapiens
Mus

overall_design: we examined the modification level of the PTC sites in HEK293T cells or cochlear tissues with or without RESTART treatment to evaluate the eidting efficiency.


title: Single nucleic RNA sequencing after mouse spinal cord injury: Effects of athlete-derived extracellular vesicles

city: Nanjing

first: xuhui

last: Ge

address: Hanzhong Road

institute: Nanjing medical university

summary: Spinal cord injury (SCI), a disease with a very poor prognosis, poses a serious social hazard and therapeutic burden due to its high morbidity, disability, and mortality. Despite the availability of existing treatments including surgery and drugs, their limited efficacy necessitates the  development of innovative therapeutic approaches. As a new form of cell death, ferroptosis plays an important role in the context of SCI. Therapies targeting ferroptosis could offer novel approaches to improve neuronal survival after SCI. Additionally, as an important intercellular communication mediator, extracellular vesicles show potential therapeutic effects after SCI. In this study, we conducted single nucleic RNA sequencing (snRNA-seq) on mouse spinal cords after SCI followed by injection of athlete plasma extracellular vesicles (AEVs) or PBS to clarify multiple cell type-specific transcriptional responses, especially neuronal ferroptosis. Unbiased clustering identified 8 distinct cell clusters in the mouse spinal cords according to their expression of known cell-type-specific markers at day 7 post injury. AEVs treatment increased the frequency of viable neurons after SCI and significant alterations in ferroptosis-related gene expression within neuronal populations was identified. These data reveal the important role of AEVs in regulating neuronal ferroptosis after SCI at single-cell resolution.

overall_design: Six C57BL/6 mice (6-8 weeks old) were randomly assigned to two groups (n=3 per group): 1. PBS group 2. AEVs group. Mice underwent T10 spinal cord contusion injury and then received daily tail-vein injections of AEVs (containing 100 μg of total protein from EVs in 100 μL of PBS) or PBS alone as a control (100 μL) for five consecutive days. Mice were sacrificed at 7 days post-injury, and the spinal cord was dissected for scRNA-seq. One sample corresponds to three individual animal.

first: Yuluo

last: Rong

first: Xuhui

first: Jiaxing

last: Wang


title: Macrophage PD-1 protects against metabolic dysfunction via ULK1-dependent PD-1 phosphorylation by blocking FBXO38-mediated ubiquitination and IRE1α-mediated ER stress

city: Harbin

first: Mingming

last: Wu

state: Heilongjiang

address: 23 Youzheng St

department: Departments of Cardiology

institute: The First Affiliated Hospital of Harbin Medical University

summary: Immune checkpoint inhibitor (ICI) therapies are associated with an increased risk of metabolic syndrome; the underlying mechanisms remain elusive. We show here that anti-PD-1 antibody targets macrophage PD-1 to reduce energy expenditure without affecting food intake, leading to an augmented susceptibility of mice to high-fat diet (HFD)-induced obesity, and systemic metabolic disorders. Mechanistically, LPS activates Unc-51-like autophagy activating kinase 1 (ULK1) via Mechanistic target of rapamycin (mTOR)-dependent manner. Activated ULK1 phosphorylates PD-1 at Thr250 to inhibit FBXO38-mediated PD-1 ubiquitination and degradation by disrupting FBXO38-PD-1 binding. Phosphorylated PD-1 interacts with inositol-requiring enzyme 1α (IRE1α) and attenuates IRE1α autophosphorylation to suppress ER stress-mediated inflammatory responses. Inhibition or IRE1α knockout alleviates HFD-induced metabolic disorders in macrophage-specific PD-1 knockout mice by rescuing the reduced energy expenditure. Our findings highlight the critical role of macrophage PD-1 in the intersection of immune checkpoint blockade, energy expenditure, and metabolic dysfunction. The underscored moonlighting function of macrophage PD-1 in protecting against ER stress-driven systemic inflammation may provide new rational for combating both ICI therapy- and HFD-induced metabolic diseases.

overall_design: To investigate the role of macrophage PD-1 in regulating high-fat diet (HFD)-induced obesity and metabolic disorders, we established a mouse model with PD-1 depletion in macrophage by crossing PD-1fl/fl mice with Lyz2-Cre mice. Myeloid-specific PD-1 KO mice (PD-1fl/fl Lyz2-Cre) and their littermate controls (PD-1fl/fl) were fed on HFD for 16 weeks. Subsequently, gene expression profiling was performed using RNA-seq data derived from epididymal white adipose tissues (eWAT) of myeloid-specific PD-1 KO mice or littermate controls.

first: Zhi-Ren

last: Zhang

first: Ming-Ming


title: Chemically-inducible CRISPR/Cas9 circuits for ultra-high dynamic range gene perturbation

city: South San Francisco

first: Hetal

last: Patel

state: California

address: 1 DNA Way

institute: Genentech

summary: CRISPR/Cas9 technologies provide unique capabilities for modeling disease and understanding gene-to-phenotype connections. In cultured cells, chemical-mediated control of Cas9 activity can limit off-target effects and enable mechanistic study of essential genes. However, widely-used Tet-On systems often show “leaky” Cas9 expression, leading to unintended edits, as well as weak activity upon induction. Leakiness can be distinctly problematic in the context of Cas9 nuclease activity, which may result in cumulative DNA damage and degradation of the target cell genome over time. To overcome these deficiencies, we established transgenic platforms that minimize Cas9 functionality in the off-state along with maximized and uncompromised on-state gene editing efficiency. By combining conditional destabilization and inhibition of Cas9, we developed an all-in-one (one or multiple guide RNAs and Cas9) ultra-tight, Tet-inducible system with exceptional dynamic range (on vs. off-state) across various cell lines and targets. As an alternative to Tet-mediated induction, we created a branaplam-regulated splice switch module for low-baseline and robust Cas9 activity control. Lastly, for circumstances where DNA damage needs to be avoided, we constructed a dual-control, Tet-inducible CRISPRi module for tight and potent transcriptional silencing. This upgraded suite of inducible CRISPR systems has broad applications for numerous cell types and experimental conditions.

overall_design: Time course of inducible knockout of target gene CD81 in 293T stable lines expressing inducible Cas9 by pTET, pTET:Ultra-tight, Xon, Xon:Ultratight.  The cells are either treated with the inducer drug(s) or left untreated. Additional samples and controls are also included: a constitutive Cas9 line targeting CD81 in 293T,  untreated or treated 293T cells for knockout of B2M,  untreated or treated HAP1 cells for knockout of CD81.

first: Rajini

last: Srinivasan

first: Tao

last: Sun

first: Azalia

last: Sandles

first: Diana

first: Liang

first: Ryan

last: Pabalate

first: Maram

last: Bader

first: Amy

last: Heidersbach

first: Clark

last: Ho

first: Shiqi

last: Xie

first: Andrew

last: Ng

first: Benjamin

last: Haley


title: Circadian clock control of ribosome composition promotes rhythmic  translation and termination fidelity [Ribo-Seq]

city: College Station

first: Deborah

last: Bell-Pedersen

state: TX

address: Mail Stop 3258, BSBW210

department: Biology

institute: Texas A&M University

summary: Ribosome composition can vary depending on cell type, stress or developmental state. Here we describe circadian clock-dependent changes in ribosome composition in Neurospora crassa. Mass spectrometry of ribosomes isolated at different circadian times identified six ribosomal proteins and one ribosome-associated protein with clock-controlled abundance. We confirmed clock control of eL31-HA abundance in purified ribosomes and found that deletion of eL31 altered translation of many mRNAs and inhibited translation rhythms for 42% of rhythmically translated mRNAs. Because eL31 homologs promote translation termination fidelity, we examined ribosome protected footprint (RPF)-seq reads mapping past stop codons over circadian time to reveal clock-dependent and eL31-enhanced rhythms in translation termination fidelity. We further discovered that eL31 promotes proper ion homeostasis and translation elongation fidelity. Taken together these studies demonstrated that the circadian clock governs daily changes in ribosome composition that control rhythms in translation and impact translation fidelity, likely through changes in intracellular Mg2+ levels.

overall_design: Ribosome protected footprint (RPF)-seq of Neurospora crassa ∆eL31(∆ncu08344) mycelia, grown over a circadian time course, 10 time points, 2 replicate experiments for a total of 20 samples

first: Teresa

middle: M

last: Lamb

first: Kathrina

middle: D

last: Castillo

first: Rachel

last: Porter

first: Cheng

first: Samuel

middle: O

last: Purvine

first: Griffin

last: Best

first: Ebimobowei

last: Preh

first: Lunda

last: Shen

first: Erika

last: Zenk

first: Scott

middle: E

last: Baker

first: Matthew

middle: S

last: Sachs


title: Nutrient stress diverts RRN3 from rRNA transcription to alternative polyadenylation of autophagy mRNAs in ovarian cancer [CRISPR Screen]

city: Tianjin

first: Jianying

last: Lv

state: Tianjin

address: weijin Road No.94

department: School of medicine

institute: Nankai University

summary: Stress-induced alternative processing of mRNA is emerging as an essential mechanism to drive almost every hallmark of cancer. Through a genome-wide screening based on an abnormal transcriptional readthrough event favoring the malignant progression of ovarian carcinoma (OC), we identified novel mRNA processing regulators including RRN3, an essential factor for the transcriptional initiation of rRNA. The long-read RNA sequencing and PAR-CLIP analyses revealed that RRN3 was involved in the usage of alternative polyadenylation (APA) sites, resulting in the altered stability of autophagy-related mRNAs. More interestingly, we discovered that nutrient-deprivation-induced phosphorylation of RRN3 at serine 199 was sufficient to divert RRN3 out of the nucleolus to the nuclear plasma, where RRN3 regulated the APA of autophagy mRNAs, such as OPTN, to enhance their stability and eventually promoted autophagy. Further in vivo experiments showed that nutrient-stress-triggered switch of RRN3 from rRNA transcription to APA regulation was essential for the growth and dissemination of OC in mice.

overall_design: The GeCKO (v2.0) CRISPR library made by Zhangfeng’s lab was purchased from Addgene and amplified using the recommended protocol. The library contains 122,411 sgRNAs targeting 19,050 protein-encoding genes, 1,864 miRNA and 1,000 non-targeting control sgRNAs. A dual-fluorescence transcriptional readthrough (TRT) reporter system was established using pLV-EF1a-MCS-IRES-Bsd vector for the establishment of stable SK-OV-3 cells harboring the TRT reporter sequence (SK-OV-3TRT), in which the majority encoding sequences of COMMD3 and BMI1 were replaced by the mCherry- and EGFP-encoding sequences, respectively, and the sequence in between (including the terminal part of exon 7, intron 7, exon 8 of COMMD3, and the exon 1, intron 1, initial part of exon 2 of BMI1) was kept in frame. Lentivirus carrying the library were prepared according to the manufacturer’s protocol and infected the SK-OV-3TRT cells. The viral titer was estimated and the transduction was performed at a low multiplicity of infection (MOI) around 0.3. After puromycin (2 μg/mL) selection, mCherry+ EGFP- cells were sorted by flow cytometry. The sorted cells (mCherry+ EGFP-) and the parental cells (unsorted cells) were subjected to genomic DNA extraction, PCR-based amplification of sgRNA-encoding sequences and the high-throughput DNA sequencing


title: The RNA-binding protein PRRC2B preserves 5' TOP mRNA during starvation to maintain ribosome biogenesis   during recovery

city: Rehovot

first: Tsviya

last: Olender

address: 234 Herzl St.

department: Molecular Genetics

country: Israel

institute: The Weizmann Institute

summary: PRRC2B is an intrinsically disordered RNA-binding protein that is part of the cell’s translation machinery. Here we show that PRRC2B has two alternatively spliced mRNA transcripts producing major long and minor short isoforms. Mass spectrometry-based interaction studies indicated that both isoforms associate with the 40S ribosomal subunit and translation initiation factors. Importantly, the long isoform also interacted with additional RNA-binding proteins through its unique Arg/Gly-rich region. Among these is LARP1, a regulator of 5' TOP mRNAs under conditions of mTOR inhibition. We discovered that like LARP1, PRRC2B is an important regulator of 5' TOP mRNA levels during starvation, particularly those encoding ribosomal proteins. Overall, our study elucidates a newly discovered function for PRRC2B as an RNA-binding protein that regulates ribosomal biogenesis, establishing it as a global regulator of translation in addition to its function in initiating specific target translation.

overall_design:
To invesigate the role of PRRC2B in TOP mRNA stabilty during starvation we first used CRISPR to reduce the levels of PRRC2B in HEK293T cells. We infected the cells with lentiviral particles carrying 2 guides targeting PRRC2B. As control we used non targeting guide rna.
We then replaced the medium of the cells to EBSS for 48hrs followed by total RNA extraction using a commercial kit. We then preformed gene profiling expression data based on 4 different biological repetitions

first: Nadav

last: Goldberg

first: Doron

last: Bril

first: Miriam

last: Eisenstein

first: Alon

last: Savidor

first: Shani

last: Bialik

first: Shmuel

last: Pietrokovsky

first: Adi

last: Kimchi


title: PPARγ agonists induce biased signaling in human adipocytes

city: Missoula

first: Travis

last: Hughes

state: MT

address: 32 Campus Drive, Skaggs BLDG RM 472

institute: University of Montana

summary: Biased signaling and ligand bias, often referred to as functional selectivity or selective nuclear receptor modulation, have been reported for many nuclear receptor partial agonists over the past 20 years. However, whether differences in nuclear receptor signaling produced by partial and full agonists results from less intense partial agonist modulation, off-target effects, or biased signaling remains unclear. To determine whether biased signaling can occur through nuclear receptors we compare the transcriptional effects of two full agonists (which also favor different coactivator peptides) in human adipocytes. We also test the signaling effects of a partial agonist relative to full agonists. Furthermore, whether biased coactivator peptide recruitment translates to biased signaling in cells has not been determined. In a step towards this goal, we show that these same full agonists induce biased recruitment of 100-300 residue regions of coactivators containing all their nuclear receptor binding motifs. Together these data support the idea that nuclear receptor agonists can induce biased signaling through differences in coactivator recruitment.

overall_design: To investigate differences in PPARγ agonist signalling, we treated human adipocytes differentiated from adult stem cells with PPARγ ligands at three and 24 hours.

first: Mariah

middle: L

last: Rayl

first: Michelle

last: Nemetchek


title: Endothelial SRSF1 directly regulates ATF3-KLF2 pathway to promote ischemia-induced angiogenesis [eCLIP]

first: Wenting

last: Zhu

address: No.95 Yong'an Road, Xicheng District

institute: Beijing Friendship Hospital, Capital Medical University

summary: Peripheral artery disease is a severe ischemic vascular pathology without effective pharmacological approaches and improving angiogenesis to recover blood perfusion is a promising therapeutic strategy. Endothelial cells (ECs) are the primary cell type contributing to angiogenesis in response to ischemia. However, the molecular mechanisms regulating ischemia-induced angiogenesis remain elusive. We used a discovery-driven approach to identify elevated Peripheral artery disease is a severe ischemic vascular pathology without effective pharmacological approaches and improving angiogenesis to recover blood perfusion is a promising therapeutic strategy. Endothelial cells (ECs) are the primary cell type contributing to angiogenesis in response to ischemia. However, the molecular mechanisms regulating ischemia-induced angiogenesis remain elusive. We used a discovery-driven approach to identify elevated serine/arginine splicing factor 1 (SRSF1) expression in endothelial cells after ischemia. Proteomic analyses identified endogenous SRSF1 accumulated in ECs of the ischemic muscle and responded to hypoxia. Mice deficient in endothelial SRSF1 exhibited impaired blood flow recovery and impaired new capillary formation following HLI. Importantly, overexpression of SRSF1 enhanced blood flow recovery and angiogenesis after HLI. SRSF1 overexpression facilitated, while SRSF1 knockdown suppressed, the angiogenic function of human ECs. To explore the mechanism the mechanisms contributing to SRSF1-promoted endothelial angiogenesis,we conducted transcriptome RNA-seq searching for downstream molecules and verified it through RNA-pull down and chromatin immunoprecipitation qPCR analysis. We performed enhanced crosslinking and immunoprecipitation (eCLIP)-seq to profile the global SRSF1-binding sites on RNAs in HUVECs. Mechanistically, SRSF1 modulated ATF3 alternative splicing, wherein SRSF1 directly bound to ATF3 pre-mRNA and elevated full-length ATF3 transcript at the expense of truncated ATF3Δzip2 transcript, and in turn, ATF3 directly bound to the KLF2 promoter and suppressed KLF2-S1PR1 signaling, through which mediated the SRSF1-promoted endothelial tube formation and angiogenesis. Additionally, alprostadil, the prostaglandin E1 analog, could activate the SRSF1 signaling to improve endothelial angiogenesis in vitro and in vivo. Our findings identified SRSF1 as a novel regulator of ischemia-induced angiogenesis that enhances endothelial angiogenic functions by regulating the ATF3-KLF2/SIPR1 pathway. These results suggest that modulation of endothelial SRSF1 may represent a promising therapeutic approach for treating ischemic vascular diseases.

overall_design: We performed enhanced crosslinking and immunoprecipitation (eCLIP)-seq to profile the global SRSF1-binding activities on RNAs in HUVECs infected with Ad-GFP or Ad-SRSF1.

first: Ning

last: Xie

first: Chun-Mei

last: Cao


title: IG_Sickle_ExRNA1

city: Boston

first: Ionita

middle: Calin

last: Ghiran

state: MA

address: 3 Blackfan Circle

department: Medicine

institute: Beth Israel Deaconess Medical Center

type: Non-coding RNA profiling by high throughput sequencing

summary: To investigate changes in small RNA quantity in Sickle cell disease in mice versus non-diseased mice with or without carbon monoxide treatment.

overall_design: 16 samples, 8 controls and 8 diseased, no replicates Please note that each raw data mate file (R1 and R2) was processed separately as indicated in the corresponding processed data file name (sample_*_R1_001_fastq_*CORE_RESULTS_v4.6.2.tar.gz,  sample_*_R2_001_fastq_*CORE_RESULTS_v4.6.2.tar.gz).

first: Moua

last: Yang

first: Vivian

last: Tran


title: Adenophages are an atypical macrophage population in exocrine glands sustained by ILC2-derived GM-CSF

city: Zurich

first: Burkhard

last: Becher

address: Winterthurerstrasse 190

country: Switzerland

institute: University of Zurich

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

overall_design: Refer to individual Series


title: Circ_0006949 as a potential non-invasive diagnosis and therapy biomarker promotes the proliferation of lung adenocarcinoma cells via miR-4673 /GLUL axis

city: 天津市

first: 娜

last: 赵

address: 河西区天塔街道卫津南路天津市肿瘤医院

institute: 天津医科大学肿瘤医院

summary: To find the circRNAs specifically upregulated in lung adenocarcinoma(LUAD), we uesed circRNA microArray analysis form Arraystar to examine the expression of circRNAs in sputum of healthy individuals,  patients with lung infection or with LUAD.

overall_design: sputum specimens were distributed into three experimental groups: normol  group,lung infection group and tumor(LUAD) group, the 10 specimens from each group were equally mixed, then perform circRNAs chips by Arraystar Human circRNAs chip (Arraystar)

first: Changsen

last: Bai

first: Chaomin

last: Wang

first: Jialei

last: Hua


title: Gene expression profiles of hematopoietic stem/progenitor cells after stimulating CCL2.

city: Minato-ku

first: Takaharu

last: Kimura

state: Tokyo

address: 4-6-1 Shirokanedai

institute: The University of Tokyo

summary: To characterize the hematopoietic stem/progenitor cells (HSPCs), we performed single-cell RNA sequence analysis of cultured HSPCs after stimulating CCL2. The proportions of sub-clusters comprising HSPCs were analyzed. In addition, Myeloid clusters exhibited high expression of Kras genes. This transcriptome analysis suggested that HSPCs after stimulating CCL2 promote myeloid cell differentiation via CCL2/CCR2 signaling.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: HSPCs from mouse bone marrow and human unbilical cord are used for experiments.

first: Satoshi

last: Yamazaki

first: Yo

last: Mabuchi

first: Chihiro

last: Akazawa


title: TCR signaling via NFATc1 constrains IL-15-induced NK-like activation of human memory CD8+ T cells

status: Public on Oct 25 2025

city: Daejeon

first: So-Young

last: Kim

state: Daejeon

address: GSMSE, 291, Daehak-ro, Yuseong-gu, Daejeon, Rep, E18-1

country: South Korea

institute: KAIST

summary: Here we investigated the regulatory mechanisms of TCR-independent bystander activation and NK-like cytotoxicity of human CCR7- memory CD8+ T cells. We found that TCR signals suppressed characteristic features of IL-15-induced CD8+ T-cell activation, including upregulated NKG2D expression and genes related to NK cytotoxicity and IFN response. Moreover, ionomycin suppressed IL-15-induced bystander activation and NK-like cytotoxicity, indicating that Ca2+-calcineurin signaling is responsible for TCR-mediated suppression of IL-15-induced bystander activation. NFATc1 suppressed IL-15-induced bystander activation via binding to AP-1 that is necessary for the IL-15-induced upregulation of NK cytotoxicity-related genes. Consistently, calcineurin inhibitors enhanced IL-15-induced NKG2D expression under concurrent TCR stimulation. Additionally, we defined genes upregulated by IL-15 and downregulated by concurrent TCR signals as an IL-15-induced bystander activation gene set, which was upregulated in bystander CD8+ T cells from patients with hepatitis A virus infection. Our findings open avenues for investigating bystander CD8+ T-cell activation and its regulation in pathological conditions.

overall_design: Total CD8⁺ T cells  isolated from patients at the acute and convalescent stages of HAV infection (n = 5) and from healthy donors (n = 3), were enriched using DNA barcode-labeled MHC class I dextramers to label HAV-, CMV-, EBV-, and IAV-specific CD8⁺ T cells. These cells were then processed for single-cell RNA sequencing.

first: Hoyoung

last: Lee

first: Su-Hyung

last: Park

first: Hyuk Soo

last: Eun

first: Eui-Cheol

last: Shin


title: Fruit derived superoxide dismutase nanozyme for sepsis-induced acute lung injury therapy

city: jinan

first: zhongyao

last: li

address: qingdao road 6699

institute: shandong first medical university

summary: As a life-threatening infectious disease in clinics, sepsis is characterized by uncontrolled systemic inflammatory response and multi-organ damage due to the excessive production of reactive oxygen species (ROS). The lung is the most frequently affected organ that is susceptible to developing acute lung injury (ALI) with high morbidity and mortality rates. Nevertheless, there are currently no effective drugs for the treatment of ALI. Herein, we developed a natural superoxide dismutase-mimicking carbon dots (H-CDs) derived from a fruit, hawthorn, with distinctive ROS scavenging ability for ALI therapy. The carboxyl/oxygen-based groups give the generated H-CDs excellent SOD-like activity of > 9000 U mg-1 and acted as a SOD-like nanozyme in the protection of cells from oxidative damage by scavenging ROS and ameliorating the levels of pro-inflammatory factors. Importantly, H-CDs reduced lung inflammatory response, oxidative damage, and histological severity in a lipopolysaccharide (LPS)-induced ALI mice model with minimal adverse effects. Mechanistically, H-CDs protected lung tissues by activating Nrf-2/HO-1 mediated antioxidant signaling and inhibiting NF-κB-dependent inflammatory responses. Collectively, this study establishes the potential of H-CDs as a promising therapeutic agent for the treatment of ALI.

overall_design: RAW264.7 cells were cultured in a 6 wells plate in LPS (1 ug/ml) containing medium for 6 h supplemented with or without H-CDs (400 ug/ml), and total RNA was extracted using the TRIzol reagent. RNA sequencing was conducted using the Bioanalyzer 2100 system (Agilent Technologies, CA, USA).

first: Zhongyao

last: Li


title: Construction of Human nucleus basalis Organoids and Cholinergic Projections in nbM-cortical Assembloids

first: Yan

last: Liu

state: Jiangsu

address: 101Longmian Avenue, Jiangning District

department: School of Pharmacy

institute: Nanjing Medical University

summary: The nucleus basalis, also known as the nucleus basalis of Meynert (nbM), which is considered to be one of the major cholinergic output of basal forebrain, have been found to dynamically modulate activity in the cortex. Dysfunction of nucleus basalis-cortical cholinergic circuit led to cognitive impairment, such as Alzheimer's disease (AD) and Down syndrome (DS). Human nucleus basalis cholinergic neurons derived from human pluripotent stem cells provide powerful tools to study cholinergic neurons-associated diseases and cell therapy. Previous studies reported the generation of 2D human basal forebrain cholinergic neurons which failed to recapitulate the spatial organization, cellular diversity, and crosstalk between different regions. Therefore, a better model to recapitulate human nucleus basalis and cholinergic projections in nbM-cortical is desired. Here we developed a approach for differentiating human pluripotent stem cells into nucleus basalis of Meynert organoids (hnbMOs). We reconstructed hnbM-cortex cholinergic projection by transplanting hnbMOs into immunodeficiency mice to construct chimeric brains and coculturing with human fetal brain. Then we fused hnbMOs with cerebral cortex organoids (hCOs) to form hnbMO-hCO assembloids. We validate the structural and functional connectivity of basal forebrain cholinergic neurons to the cortex in assembloids. An assembloid-chimeric brain was constructed innovatively by transplanting corresponding organoids in the cortex and nbM region to establish a complete human cholinergic projection system. Futhermore, we identified the defects in projection of cholinergic neurons at the morphological and transcriptomic level in Down syndrome patient iPSC-derived assembloids as well as Down syndrome fetal brain tissue. Our work establishes new approach for the study of neurological disorders associated with nbM and nbM-cortical cholinergic neuron circuit.

overall_design: The study presents a novel method for creating nucleus basalis organoids from human pluripotent stem cells to model cholinergic connections to the cortex, helping to investigate neurological disorders linked to this system, particularly in the context of Down syndrome and Alzheimer's disease.


title: CRISPR-free RNA Base Editing Mediated PTC-readthrough Restores Hearing in Humanized Mice with Otof Nonsense Mutation [RNA-seq]

city: Beijing

first: Wenqing

address: No. 5 Summer Palace Road, Haidian District, Beijing

institute: Peking University

summary: Today, the gene editing for hereditary deafness is becoming a promising field with global interest. Among them, AAV-mediated otoferlin overexpression is undoubtedly the most successful representative of deaf-related gene therapy. However, it remains challenging to realize physiological, endogenous pattern of otoferlin expression. Here, we firstly generated a humanized homology Otof c.1315C>T (p.R439X), equivalent to OTOF c.1273C>T (p.R425X) identified in human with profound hearing loss, nonsense mutation-caused deaf mice model. We then delivered the ‘RESTART v3’ system, which is a CRISPR-free RNA base editor for nonsense mutation suppression, into the cochlea of the mice model. We accomplished a physiological manner of otoferlin expression; moreover, the edited premature termination codon is precisely reversed back to the original amino acid. Using multiple assays to estimate hearing restoration, we identified a significant reduced thresholds of auditory brain response (ABR) and statistical enhanced amplitudes of behavioral auditory startle reflex (ASR) in the treated mice with the RNA editing system. Thus, our study presented a successful CRISPR-free RNA editing approach to significantly restore hereditary deafness carrying humanized Otof c.1315C>T (p.R439X) non-sense mutation, providing a great promise for future clinical translation.

overall_design: we examined the global modification level of cochlear tissues with or without RESTART treatment to evaluate the off-target events.


title: Pharmacological Targeting of ALDH1a2 in Dendritic Cells Enhances Antigen Presentation, T Cell Activation and Anti-tumor immunity

city: Plainsboro

first: Yong

last: Wei

address: LTL227 Wathington Road

department: Molecular Biology

institute: Princeton University

summary: Dendritic cell (DC) vaccines have been proposed as cancer immunotherapies due to their role as crucial antigen-presenting cells that regulate T cell functions. Despite considerable efforts to optimize ex vivo priming of DCs, DC vaccines have rarely been successful, suggesting the presence of unknown inhibitory factors. Here, we examined DC differentiation dynamics and discovered that ALDH1a2-produced retinoic acid (RA) acts as a bottleneck factor, initiating negative feedback to inhibit DC maturation. Removing this inhibition through either genetic knockout or pharmacological blockade using KyA33, an ALDH1a2 inhibitor we developed, significantly enhances DC maturation, phagocytosis, antigen presentation, and T cell activation, in part by downregulating glucose metabolism. KyA33 demonstrates favorable drug-like properties, including low toxicity, high membrane permeability, and low cell efflux rate. Its non-covalent binding to ALDH1A2 was also validated through X-ray crystallography. Importantly, application of KyA33 generates more robust DCs vaccines, promoting anti-tumor immunity through enhancing antigen-specific T cell responses. Our investigation highlights the intricate interplay between retinoid signaling, dendritic cell maturation, and immune metabolism, offering promising avenues for enhancing cancer immunotherapies.

overall_design: To characterize BMDC subpopulations of high vs low maturation, two groups of BMDC were generated by bone marrow cell culture w/ and w/o retino acid and sorted by CD11c/MHC/CD86 using FACS. Three biological replicates were produced per group for bulk RNAseq analysis. To further characterize the role of Aldh1a2 in the process, BMDC from Aldh1a2 wild type and KO mice were generated for bulk RNAseq analysis. Two models were used to achieve the KO of Aldh1a2 in BMDC cells by Cre/Flox system, either CD11c-Cre to achieve DC specific KO and UBC-CreERT2 combined with tamoxifen to induce whole body Aldh1a2 KO. In both model, littermate mice without Cre are used as wild type control. Bulk RNA sequencing was performed to compare gene expression between Aldh1a2 KO and WT BMDCs in each model.

first: Cao

last: Fang

first: Mark

last: Espisito

first: Yibin

last: Kang


title: Bovine Formative Embryonic Stem Cell Plasticity in Embryonic and Extraembryonic Differentiation

city: Storrs

first: Yue

last: Su

address: 1390 Storrs Road

institute: University of Connecticut


title: Inflammation perturbs hematopoiesis by remodeling specific compartments of the bone marrow niche

city: New York

first: James

last: Swann

state: NY

address: 650 W 168th Street

institute: Columbia University

summary: Hematopoietic stem and progenitor cells (HSPC) are regulated by interactions with stromal cells in the bone marrow (BM) cavity, which can be segregated into two spatially defined central marrow (CM) and endosteal (Endo) compartments. However, the importance of this spatial compartmentalization for BM responses to inflammation and neoplasia remains largely unknown. Here, we extensively validate a combination of scRNA-seq profiling and matching flow cytometry isolation that reproducibly identifies 7 key CM and Endo populations across mouse strains and accurately surveys both niche locations. We demonstrate that different perturbations exert specific effects on different compartments, with type I interferon responses causing CM mesenchymal stromal cells to adopt an inflammatory phenotype associated with overproduction of chemokines modulating local monocyte dynamics in the surrounding microenvironment. Our results provide a comprehensive method for molecular and functional stromal characterization and highlight the importance of altered stomal cell activity in regulating hematopoietic responses to inflammatory challenges.

overall_design: Combined stromal and hematopoietic cells isolation Mice were euthanized in a rising concentration of carbon dioxide, followed by cervical dislocation. For each mouse, both femurs, tibiae, hemipelves, and humeri were dissected and thoroughly cleaned using KimWipes (Kimtech) and were used for combined isolation of stromal and hematopoietic cells. To isolate central marrow (CM) stromal cells, the proximal and distal epiphyses were removed from 1 femur. Intact marrow plugs were flushed with Hank’s balanced saline solution (HBSS) without calcium or magnesium into 5 ml polypropylene tubes by inserting a 3 ml syringe with 22G needle into the distal end of the femur. The plugs were digested with 1 ml of a solution of 3 mg/ml type I collagenase (Worthington) dissolved in HBSS for 10 min at 37 C with 110 rpm shaking. The tube was then vortexed briefly and the supernatant was removed into a new tube through a 100 m mesh, taking care not to disturb the marrow plug. A further 1 ml of 3 mg/ml type I collagenase solution was then added to the marrow plug and the digestion repeated. After the second incubation, the plug was dissociated by pipetting up and down with a P1000 pipette before passing the cell suspension through a 100 m mesh into the tube containing the previous digestion supernatant. To isolate endosteal (Endo) stromal cells and hematopoietic cells, the flushed femur was combined with the other bones, gently crushed up to 10 times using a mortar and pestle, and thoroughly washed with 10 ml HBSS until all the non-adherent BM cells were removed and collected in a separate 15 ml polypropylene tube. The bone chips were then digested in a 15 ml polypropylene tube with 3 ml of 3 mg/ml type I collagenase solution for 1 hour at 37 C with 110 rpm shaking. After digestion, the tube was vortexed briefly and the cell suspension was filtered through a 100 m mesh into a new tube. The bone chips were then washed with HBSS, and the washing was collected into the same tube as the digestate. For cell suspensions acquired for both stromal compartments, red blood cells were removed by adding 1 ml ACK lysis buffer (150 mM NH4Cl and 10 mM KHCO3) and incubated on ice for 3 minutes before washing with HBSS containing 4% fetal bovine serum (HI FBS, Gibco). During the optimization of the protocol outlined above, the digestion with 3 mg/mL type I collagenase was performed in parallel to and compared with 3 mg/mL type I collagenase + 4 mg/mL type II Dispase (Roche), 250 μg/mL Liberase DL (Roche) + 200 U/mL DNAse I (Sigma), and mechanical dissociation with P1000 pipette. The BM cells collected from crushed bones were resuspended in HBSS with 2% FBS and RBCs were removed by lysis with ACK buffer. The BM cell suspension was further purified on a density gradient (Histopaque 1119, Sigma-Aldrich), by layering 2 ml of the cell suspension under 2 ml of Histopaque solution. For profiling of CM plug and Endo marrow, only femurs were used and were flushed once with a 3 ml syringe and 21G needle to collect BM plugs, while the flushed bones were crushed to collect the remaining Endo-associated BM cells. No ACK RBC lysis was performed for these analysis. Both stromal and hematopoietic cells were finally counted using a Vicell automated cell counter (Beckman Coulter).  RNA sequencing - Droplet-based scRNA-seq for stroma and hematopoietic analyses For Endo and CM stromal cell preparations, 30,000–50,000 Ter119-/CD45- cells were sorted on purity mode into 1.5 ml tubes containing 300 μl of 50% FBS and 50% HBSS using a BD FACS Aria II instrument (gating shown in Supplementary Fig. 2a, 3b). For whole BM cells, 40,000 live (propidium iodide negative) cells from n=2 mice were sorted into similar tubes from PBS or pIC-injected mice. Cells were rested for 1 hour on ice then pelleted at 350 x g for 5 minutes at 4°C before the supernatant was removed down to a volume of 40 μl.  For sequencing of defined HSPCs, 8,000 cells of 10 different populations (HSC, stHSC, MPP2, FcγR- MPP3, FcγR+ MPP3, MPP4, GMP, MkP, CLP, CFU-E, see Supplementary Fig. 1 for sorting scheme) were sorted separately into 1.5 ml tubes. In this experiment, there were 2 biological replicates with BM from n=5 mice, and 2 technical replicates were generated per sample. Sorted cell populations were labelled with 0.05 μg of oligo-hashed antibodies specific for MHC class I and CD45 (BioLegend, TotalSeq B1-10) for 30 minutes on ice before washing three times with HBSS containing 2% FBS. All 10 populations were then pooled for subsequent partitioning. Cells were rested for 1 hour on ice then pelleted at 350 x g for 5 minutes at 4°C before the supernatant was removed down to a volume of 40 μl.  For sequencing of purified stromal populations, samples of central marrow or endosteal stromal cells pooled from n=7 mice were labelled with oligo-hashed antibodies at a concentration of 0.375 μg of antibody per 106 cells. These antibodies were mixed with the fluorophore-conjugated surface antibodies, and cells were then washed three times with HBSS and 4% FCS. Specific stromal cell populations were then sorted by FACS (gating scheme shown in Fig. 1e), and mixed according to the following scheme to produce two combined samples, each containing a total of 25-40,000 cells: Sample type Oligo hash antibody Sample 1 (cells) Sample 2 (cells) CM TotalSeq B1 MSC-L (6899) SEC (2228) CM TotalSeq B2 mMPr (2775) SEC (2097) CM TotalSeq B3 mMPr (2406) SEC (2529) CM TotalSeq B4 MPr (847) SEC (1041) CM TotalSeq B5 MPr (802) SEC (1353) CM TotalSeq B6 MSC-L (7642) AEC (438) CM TotalSeq B7 - AEC (496) Endo TotalSeq B8 MPr (10048) MSC-S (8212) Endo TotalSeq B9 AEC (5046) MSC-L (2319) Endo TotalSeq B10 SEC (3537) mMPr (6823)  Cells were rested for 1 hour on ice then pelleted at 350 x g for 5 minutes at 4°C before the supernatant was removed down to a volume of 40 μl. GEM generation and 3' RNA library preparation was performed according to 10X Genomics protocol CG000315 Rev E, targeting 5000 cell data recovery. RNA libraries were pooled 1:1:1:etc, sequenced on an Illumina NovaSeq 5000, and alignment was performed using Cellranger (v.7.0.1) to mouse genome mm10. For hashing samples, hashing libraries were pooled with RNA libraries at a ratio of 1:4. Library concentrations and fragment sizes were evaluated using Qubit dsDNA HS assay kit (ThermoFisher Scientific) and TapeStation D5000 DNA ScreenTape analysis (Agilent).  RNA sequencing - Plate-based SMART-seq for stromal cells and MSC-L For Smart-seq analyses of Endo (AEC, MSC-S, mMPr, MPr) and CM (SEC, MSC-L) stromal cells isolated from young WT mice, we utilized our own published data (Mitchell et al., 2023) that was processed using the same analytical pipeline. For Smart-seq of iMSC-L from pIC-injected young WT mice and aged 24-month-old WT mice, we utilized the SMART-Seq Single Cell PLUS kit (96 reactions, Takara Bio). Single iMSC-L were sorted into 96 well plates (see Supplementary Fig. 3c for gating strategy) containing 12.5 μl of CDS sorting solution provided in the kit. Preparation and amplification of cDNA and single cell libraries were completed according to the manufacturer’s instructions. Single cell libraries that passed quality control (n=43 from pIC-injected mice and n=30 from aged mice) were pooled at a 1:1:1etc ratio before sequencing on an Illumina NextSeq 500/550 instrument using a version 2 kit (Illumina). Fastq files were generated for each library in Illumina BaseSpace.

middle: W

first: Emmanuelle

last: Passegue


title: TCR signaling via NFATc1 constrains IL-15-induced NK-like activation of human memory CD8+ T cells [RNAseq_CD3_48H]

overall_design: Sorted CCR7- memory CD8+ T cells from healthy donors (n = 10) were stimulated for 48 hours with IL-15 (10 ng/ml), anti-CD3 (coated, 1 µg/ml), or the combination of both. Total RNA was isolated and RNA-seq was performed.


title: Gene expression profile at single cell level of Normal Colonic Epithelial Stem Cells (NCE-SC)

city: Kyoto

first: Makoto

middle: Mark

last: Taketo

state: Kyoto

address: Yoshida-Honmachi, Sakyo

department: Kyoto University Hospital

institute: Kyoto University

summary: NCE-SC are inferred as a homogeneous population cultured in the eL-WRN medium (Miyoshi H, Oncotarget 2018;9(31):21950-64.). We used single-cell RNA sequencing (scRNA-seq) to confirm the homogeneity of NCE-SCs.

overall_design: Cultured NCE-SC of human colon were trypsinized, filtered through a strainer and analyzed using scRNA-seq.

first: Fumihiko

last: Kakizaki

first: Hiroyuki

last: Miyoshi

first: M


title: MIF Nuclease Inhibition Protects CNS Neurons following Inflammation.

status: Public on Oct 24 2025

city: Baltimore

first: Matthew

last: Smith

state: MD

address: 600 N Wolfe Street

department: Neurology

institute: Johns Hopkins University


title: Colon Cancer Patient-Derived Spheroids and their Stem Cell-Intrinsic mRNA Expression Yield Novel Prognostic Signatures

summary: We characterized colorectal cancer stem cell (CRC-SC)-intrinsic subtypes based on mRNA expression of 57 patient-derived SC lines. We compared their expression profiles with those of normal colonic epithelial stem cells (NCE-SC), and obtained genes relatively underexpressed or overexpressed. First, we found a subgroup of CRC-SC lines in which seven genes were overexpressed despite very low expression in the remaining CRC-SC lines. Importantly, cross-reference of the expression-survival relationship in public databases indicated the Lucky Seven genes to constitute a signature for improved outcome. Likewise, five lead genes were expressed at higher levels than in NCE-SC, representing non-overlapping subtypes. While their expression affected patient outcome singly in each subtype, other related genes overexpressed within the subtype were also associated with better or poorer prognosis. When combined, collective signatures provided novel predictive signatures with much stronger statistical significance. Finally, we integrated the six signatures into a comprehensive signature, which yielded a novel survival prognostic indicator.

overall_design: Six prognostic signatures for colorectal cancer, three favorable and three unfavorable were revealed as a consequence of the 57 patient-derived cancer stem cell lines. Critically, we devised a predictor applicable to any patient via the integration of the six signatures that can be determined using the whole tumor tissue sample.


title: Effect of matenral immune activation on lung immune cells during house dust mite induced allergic inflammation. [scRNA-Seq]

first: Myoung Seung

last: Kwon

state: Yuseong-gu

address: 291, Daehak-ro, Yuseong-gu, Biomedical research center (E7)

summary: To investigate the effect of maternal inflammation during pregnancy on allergic reactions in offspring, we inoculated house dust mite and ovalbumin into offspring born to either control or LPS induced maternal immune activation dams and analyzed their lung immune cells.

overall_design: We analyzed gene expression changes in 5,000 CD4-positive and 5,000 CD4-negative cells among CD45+ cells In lung.

first: Myoungseung


title: Soluble αKlotho interacts with Hsp90aa1 to inhibit the chaperone machinery-mediated Hif1α stabilization and alleviate CKD-induced vascular calcification

city: Jinan

first: Fengyang

last: Xu

state: Shandong

address: Wen Hua Xi Road

department: Qilu Hospital

institute: Shandong

summary: Vascular calcification is the major risk factor for increased cardiovascular morbidity and mortality in patients with CKD, especially those with advanced renal diseases. However, once mineral deposits form in vascular tissue, no specific therapies can completely arrest or reverse the progression of calcification. Here, we identified that decreased serum soluble αKlotho levels are characteristic events in the development of vascular calcification in patients with early CKD. We employed a transcriptomicsapproach, as well as in vitro and in vivo vascular calcification models, and determined that soluble αKlotho specifically suppressed Hsp90aa1 activation-mediated osteogenic transdifferentiation of VSMCs and vascular calcification. Moreover, we revealed that the phosphorylation of Hsp90aa1 at Thr5/7 modulated its chaperone activity to stabilize Hif1α, thereby playing a causative role in the pathogenesis of vascular calcification in response to CKD. Upregulation of soluble αKlotho expression in VSMCs enhanced the interaction with Hsp90aa1, which abolished Hsp90aa1-Hif1α axis activation in response to osteogenic induction. Together, these results identify a key pathway in VSMCs that mediates vascular calcification, which could be used as a potential therapeutic target.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for MA-VSMCs after treatment with calcification medium or without it, combined with or without αKlotho lentiviral vector transfection

first: Jialin

last: Guo

first: Yunyun

first: Jiaxin

last: Ma

first: Wentao

last: Sang

first: Xiangkai

last: Zhao

first: Han

last: Du

first: Dehui

last: Hou

first: Jian

last: Zhang

first: Hongwei

last: Yue

first: Kehui

last: Yang

first: Sumei

last: Cui

first: Tonghui

first: Feng

first: Yuguo

last: Chen


title: Large scale cell-based transcriptome data treated with Jakyakgamcho-tang in HepG2 cell

first: Su-Jin

last: Baek

address: 1672, Yuseong-daero

institute: KOREA INSTITUTE of ORIENTAL MEDICINE

summary: Jakyakgamcho-tang (JGT) is the simplest form of herbal medicine composed of Paeoniae Radix (PR) and Glycyrrhizae Radix at Rhizoma (GR), and has been used to treat muscle-related diseases and inflammation. However, the pharmacological effects of JGT may vary depending on the ratio of the constituent herbs and the main preparation conditions of the extraction method. Nevertheless, no gene expression dataset systematically reflecting these various variables has existed to date. In this study, a total of 498 transcriptome profiles were constructed with three concentrations and three replicates of RNA-seq data. It is structured to enable multidimensional analysis of the effects of various preparation conditions on gene expression by combining the ratio of PR:GR (2:1, 1:1, 1:2), solvent (water, 70% ethanol), and extraction method (combined decoction vs. separate decoction) of JGT, targeting three cell lines (HepG2, C2C12, and PC12). All raw and preprocessed data are deposited available through GEO, and standardized metadata and ingredient data are also provided. This dataset provides a foundation for exploring how traditional herbal medicine formulations affect cellular transcriptomic responses and can serve as a baseline for scientific optimization of herbal medicines.

overall_design:
Whole transcriptomic analysis in HepG2 cells was conducted, comparing cells treated with a vehicle control to those treated with three different concentrations of JGT-E and JGT-W. All experiments were performed in triplicate.
We constructed the pharmacological transcriptome data obtained through combinatorial conditions, such as extraction orders (individual herbs extracted or the herb-mixed JGT extracted), combination ratios (two herbs into 2:1, 1:1, and 1:2), two extracting methods (hot water or 70% ethanol).

first: Seongwon

last: Cha

first: Jin-Mu

last: Yi


title: Epithelial Reprogramming by OM-89 Enhances Antibiotic Clearance of Uropathogenic E. coli in a Bladder Organoid Model

city: Lausanne

first: Kathrin

last: Tomasek

address: Rte Cantonale

institute: EPFL

summary: Recurrent urinary tract infections (UTIs) are a major clinical burden, often driven by the ability of uropathogenic Escherichia coli (UPEC) to persist intracellularly within bladder epithelial cells, protected from antibiotics and host immune responses. While vaccine and antibiotic alternatives are being explored, strategies that enhance epithelial cell-intrinsic defense mechanisms remain sparse. Here, we show that OM-89 (Uro-Vaxom®), an E. coli lysate, reprograms epithelial antimicrobial responses to enhance intracellular bacterial clearance. Using murine bladder organoids and epithelial cell monolayers, we demonstrate that OM-89 enhances lysosomal acidification and cathepsin activity, promoting trafficking of UPEC into degradative compartments. Unexpectedly, OM-89 also increases epithelial uptake of antibiotics, increasing their intracellular potency and reducing UPEC regrowth after treatment. This dual effect, on epithelial immunity and antibiotic accessibility, was conserved across multiple UPEC strains and antibiotic classes. Our findings position bladder epithelial cells as active players in the host defense and highlight OM-89 as a tool to amplify their antimicrobial potential, offering a promising adjunctive strategy against intracellular bacterial persistence.

overall_design: RNA profiling of mouse bladder organoids comparing the response to OM-89 treatment paired with e.coli infection.

first: Kristina

last: Skurvydaite

first: Gauri

last: Paduthol

first: Allison

last: Burns

first: Léa

last: Schlunke

first: Christian

last: Pasquali

first: Mario

last: Romani

first: John

last: McKinney


title: RNA sequencing analysis of wild-type and ITF2357 or I-BET151 or JQ1 treated mouse pancreatic cancer KPC cells

city: Indianapolis

first: Xiongbin

last: Lu

state: Indiana

address: 980 W. Walnut Street, R3-C218D	MMGE

institute: Indiana University School of Medicine

summary: The ITF2357, I-BET151 and JQ1 promote antigen presentation and sensitize pancreatic cancer to T cell cytotoxicity, in combination with Immune checkpoint blockage therapy suppress tumor cell growth

overall_design: Comparison of total RNA expression profile of wild-type and ITF2357 or I-BET151 or JQ1 treated mouse pancreatic cancer KPC cells

first: Zhuolong

last: Zhou


title: Gene expression profiling of Trophoblast Stem Cells with Progesterone Receptor (PGR) knockdown

city: Shanghai

first: Xiaoya

last: Zhao

state: -- None selected --

address: No. 910, Hengshan Rd

institute: School of Medicine, Shanghai Jiaotong University

summary: As medical abortion has established itself as the significant method of terminating pregnancies, many studies in recent years has focused on Mifepristone, a principal agent used in this field. It can be found that much effort has been devoted to the research on the impact of Mifepristone on the decidua and embryo implantation process in the mechanism of abortion and contraception; the mechanism of its impact on trophoblast cells, however, still remains unclear. Therefore, the present study aims to fill the gap by investigating the effects of Mifepristone on villous trophoblasts in medical abortion patients. We have found that Mifepristone affects trophoblast cell proliferation and invasion through the progesterone receptor. We established PGR-knockdown trophoblast stem cells and collected these cells for comprehensive RNA-seq analysis.

overall_design: Trophoblast stem cells extracted from normal human placental villi underwent gene knockdown of PGR (shPGR) and luciferase (Control). Subsequently, transcriptome sequencing was performed on both cell groups.

first: Jie

first: Jian

last: Zhang


title: Spatiotemporal characterization of human early intervertebral disc formation at single-cell resolution

first: Taifeng

state: Massachusetts

address: 188 Longwood Avenue, REB514B

department: Developmental biology

institute: Harvard University

summary: Intervertebral disc (IVD) performs as a fibrocartilaginous joint to anchor adjacent vertebrae (VB) and is essential for the mechanical stabilization of axial skeleton. Although several studies have demonstrated the cellular heterogeneity of adult mature IVD, a single-cell transcriptomic atlas mapping early IVD formation is still lacking. Here, we generated the first spatiotemporal and single-cell based transcriptomic atlas of human IVD formation at embryonic stage, as well as a comparative mouse transcript landscape. The notochord (NC)/nucleus pulposus (NP) cells, vertebral chondrocytes, and annulus fibrosus (AF) cells were characterized. Our single-cell transcriptomic atlas that resolves spatiotemporally regulated cellular heterogeneity underlying early IVD development will be leveraged to develop preventative and regenerative strategies for IVD degeneration.

overall_design: Three embryonic mouse axial skeleton at E11.5-13.5 were cryosectioned to a thickness of 10 μm, followed by spatial transcriptome analysis via 10x Genomics Visium.

first: Yu

first: Zhiheng

last: Liao

first: Ren

last: Xu

first: Peiqiang

last: Su


title: Development of a split-toxin CRISPR screening platform to systematically identify upstream regulators of human myoblast fusion

city: Athens

first: Pengpeng

last: Bi

state: GA

address: 315 Riverbend Rd

institute: University of Georgia

summary: Muscle defects are a common feature in human developmental disorders and often lead to severe functional impairment. These defects arise from intricate tissue crosstalk and rare genetic mutations, underscoring the need to systematically identify cell-autonomous mechanisms regulating human myogenesis. Despite the clinical significance, our understanding of human development remains limited, due in part to the absence of a scalable genetic approach to study this process. Here, we introduce a rationally designed high-throughput CRISPR screening platform that integrates human myoblast models, muscle-specific CRISPR knockout libraries, and a split-toxin strategy that acts as a functional readout for myoblast fusion—a fundamental step of human myogenesis. This screening strategy enables selection of CRISPR-induced fusion-defective myocytes in a quantifiable manner. Leveraging this platform, our initial genetic screen uncovered a large group of new hits essential for human myoblast fusion. The majority of these hits converge into 23 protein complexes, most of which have not previously been functionally linked to myogenesis in any species. Notably, mutations in 41 of our fusion screen hits cause human diseases presenting abnormal skeletal muscle morphology. Applying a new single-cell CRISPR & RNA-seq approach, we show that majority of these hits control human myoblast fusion as well as influence early-stage myogenic differentiation. Together, this work presents a new systematic approach to study human myogenesis and uncovers promising candidates governing human muscle differentiation and fusion. A broader application of this split-toxin based CRISPR screening platform would accelerate the study of cell-autonomous mechanisms of human muscle development and diseases at scale.

overall_design:
RNA-Seq profile of human myoblast at different stages, including myoblast in growth medium (GM), differentiate for 2 days (DM2d), differentiate for 6 days (DM6d).
Split-pool based single cell CRISPR screen data.


title: Dual inhibition of FACT and RRM2 suppresses the progression of pancreatic ductal adenocarcinoma driven by the oncohistone H2BG53D

city: Hong Kong

first: Tiantian

last: QIN

address: Tat Chee Avenue, Kowloon

institute: City University of Hong Kong

summary: Pancreatic Ductal Adenocarcinoma (PDAC) is a highly lethal cancer with frequent genetic mutations. We previously reported the identification of the histone H2BG53-to-D mutation in PDAC patients and revealed that the mutation destabilizes nucleosomes and promotes cell migration. However, how H2BG53D impacts chromatin and the significance in PDAC development remain largely unknown. Using the genetically engineered mouse models, we demonstrate that H2BG53D accelerates PDAC initiation and metastasis. Moreover, H2BG53D mutation elevates the interaction between nucleosomes and the Facilitates Chromatin Transcription (FACT) complex. FACT depletion reverses transcriptional changes and oncogenic effects induced by H2BG53D mutation. In search of a novel therapeutic strategy for H2BG53D-PDAC, our genome-wide CRISPR/Cas9 screen identified Ribonucleotide Reductase Regulatory Subunit M2 (RRM2) as a druggable target. Remarkably, dual inhibition of RRM2 and FACT exhibits potent cytotoxic effects on PDAC cells in vitro and significantly prolongs the survival of H2BG53D-PDAC mice. This study deciphers the roles and mechanisms of H2BG53D in PDAC progression and proposes a promising therapeutic strategy for this aggressive cancer.

overall_design: mouse RNA-seq: KPC and G53D-KPC mice were generated to study the roles of H2BG53D mutation in PDAC development. Primary PDAC tumors and metastases from KPC and G53D-KPC mice were used for this experiment. Gene expression was profiled by RNA-seq. KI cell line RNA-seq: FLAG tagged Wildtype (WT) H2B and H2BG53D knockin PDAC cells were used to study the mechanisms of  H2BG53D on transcriptional alteration and oncogenic phenotype. RNA-seq  were conducted in  in two WT and two H2BG53D cells with or without SPT16 (one subunit of FACT)  depletion. KI cell line CUT&RUN seq: FLAG tagged WildtypeH2B (H2B_WT-FLAG)and H2BG53D-FLAG knockin PDAC cells were used to study the mechanisms of  H2BG53D on transcriptional alteration and oncogenic phenotype.  CUT&RUN seq (targeting FLAG and H2A) were conducted in parental, two WT and two H2BG53D cells with or without SPT16 (one subunit of FACT)  depletion. KI cell line CRISPR screen: FLAG tagged WildtypeH2B (H2B_WT-FLAG) and H2BG53D-FLAG knockin PDAC cells were used to identify the synthetic lethal partner for H2BG53D mutation in PDAC.

first: Kui Ming

last: CHAN


title: C-to-U RNA editing with RECODE in human cells

city: Fukuoka City

first: Soichi

last: Ozaki

state: Fukuoka

address: West Zone 5 #430, Kyushu University, 744 Motooka, Nishi-ku,

institute: EditForce, Inc.

summary: Guide-RNA-dependent cytidine deaminase tools to convert cytidine-to-uridine (C-to-U) have limitations such as low editing efficiency, limited targetable sequence flexibility, and off-target RNA editing. Here, we report the editing efficiency in HEK293T human cells of a novel guide-free C-to-U editing tool, named RECODE, based on RNA-binding pentatricopeptide repeat proteins naturally fused to a C-terminal DYW cytidine deaminase domain. The RECODE specificity domain was engineered to enable retargeting, while its length and sequence were optimized to reduce off-target effects. Further optimization of the C-terminal catalytic region increased both the editing activity and the translation of the edited RNA. The programmability, high editing efficiency, reduced neighboring nucleotide preference and gRNA-free design establish the RECODE technology as a versatile platform with potential applications beyond cellular systems, such as therapeutic approaches for repairing disease-causing mutations and modifying protein functions to enable novel treatments.

overall_design: RNA-seq to identify global C-to-U substitution events induced by RECODE or RESCUE-S, 48 hours after transfection.


title: Circadian clock control of ribosome composition promotes rhythmic  translation and termination fidelity [RNA-Seq]

city: College Station

first: Deborah

last: Bell-Pedersen

state: TX

address: Mail Stop 3258, BSBW210

institute: Texas A&M University

summary: Ribosome composition can vary depending on cell type, stress or developmental state. Here we describe circadian clock-dependent changes in ribosome composition in Neurospora crassa. Mass spectrometry of ribosomes isolated at different circadian times identified six ribosomal proteins and one ribosome-associated protein with clock-controlled abundance. We confirmed clock control of eL31-HA abundance in purified ribosomes and found that deletion of eL31 altered translation of many mRNAs and inhibited translation rhythms for 42% of rhythmically translated mRNAs. Because eL31 homologs promote translation termination fidelity, we examined ribosome protected footprint (RPF)-seq reads mapping past stop codons over circadian time to reveal clock-dependent and eL31-enhanced rhythms in translation termination fidelity. We further discovered that eL31 promotes proper ion homeostasis and translation elongation fidelity. Taken together these studies demonstrated that the circadian clock governs daily changes in ribosome composition that control rhythms in translation and impact translation fidelity, likely through changes in intracellular Mg2+ levels.

overall_design: RNA-seq (done in parallel with ribosome profiling) of Neurospora crassa ∆eL31(∆ncu08344) mycelia, grown over a circadian time course, 10 time points, 2 replicate experiments for a total of 20 samples

first: Teresa

last: Lamb

first: Kathrina

last: Castillo

first: Rachel

last: Porter

first: Cheng

last: Wu

first: Samuel

last: Purvine

first: Griffin

last: Best

first: Ebimobowei

last: Preh

first: Lunda

last: Shen

first: Erika

last: Zenk

first: Scott

last: Baker

first: Matthew

last: Sachs


title: Lineage-determining transcription factors constrain cohesin to drive multi-enhancer oncogene regulation

city: Philadelphia

first: YEQIAO

last: ZHOU

state: PA

address: 421 Curie Boulevard

department: Pathology

institute: University of Pennsylvania

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

overall_design: Refer to individual Series

first: Yeqiao

last: Zhou

first: Robert

last: Faryabi


title: The carbon fixation and secretion of economic seaweed Gracilariopsis lemaneiformis under high CO2 and nitrate treatment: a CDR solution in aquaculture

city: Xiamen

first: Jichen

last: Chen

state: None Selected

address: Fujian province xiamen city

institute: Xiamen University

summary: Gracilariopsis lemaneiformis is an economic seaweed. It can play critical role in carbon dioxide removal in aquaculture. In this study, we presented physiological and transcriptomic methods to analyzed the response of G. lemaneiformis to high CO2 concentration, especially the carbon fixation and secretion.

overall_design: The Gracilariopsis lemaneiformis was collected from the macroalgal farm (119.31 °E, 26.39 °N) in Ningde city, Fujian province of China in April 2023. A detail clean of sediment, fragment and other impurities were performed. The culture medium was coastal seawater obtained from Jinjiang city, Fujian province of China. The seawater was standing for 30 days, preserved in 17°C and filtered by ultrafiltration water purifier with 0.1 μm pore diameter (MUF1580-2T, Midea Group Co., Ltd). Before culturing, the G. lemaneiformis was cultured in nature seawater with two antibiotics (Penicillin, 400 U·mL-1; Streptomycin, 0.4 mg·mL-1) for reducing the bacterial abundance in the two days. After that, the G. lemaneiformis were shifted to new medium for the wipe off of residual antibiotics in another two days. Finally, the G. lemaneiformis was transfer to new medium for 7 days of starvation. The thalli were cultured with 0.5 g·L-1 in 1 L polycarbonate bottle, at the temperature of 17°C, the light tensity of 100 μmol·photons·m-2 s-1 with the light dark cycle of 12 h: 12 h. Moreover, two crossing independent variables (CO2 concentration and nitrate concentration) were set as 4 groups which was put as Table. 1. 50% of culture solution was exchange for fresh filtered seawater with equivalent nutrient and adequate aeration every 4 days. The nutrient was added in each two days.

first: Chi

last: Song

first: Guang

last: Gao


title: TCR signaling via NFATc1 constrains IL-15-induced NK-like activation of human memory CD8+ T cells [RNAseq_IONO_12H]

city: Daejeon

first: So-Young

last: Kim

address: GSMSE, 291, Daehak-ro, Yuseong-gu, Daejeon, Rep, E18-1

institute: KAIST

summary: Here we investigated the regulatory mechanisms of TCR-independent bystander activation and NK-like cytotoxicity of human CCR7- memory CD8+ T cells. We found that TCR signals suppressed characteristic features of IL-15-induced CD8+ T-cell activation, including upregulated NKG2D expression and genes related to NK cytotoxicity and IFN response. Moreover, ionomycin suppressed IL-15-induced bystander activation and NK-like cytotoxicity, indicating that Ca2+-calcineurin signaling is responsible for TCR-mediated suppression of IL-15-induced bystander activation. NFATc1 suppressed IL-15-induced bystander activation via binding to AP-1 that is necessary for the IL-15-induced upregulation of NK cytotoxicity-related genes. Consistently, calcineurin inhibitors enhanced IL-15-induced NKG2D expression under concurrent TCR stimulation. Additionally, we defined genes upregulated by IL-15 and downregulated by concurrent TCR signals as an IL-15-induced bystander activation gene set, which was upregulated in bystander CD8+ T cells from patients with hepatitis A virus infection. Our findings open avenues for investigating bystander CD8+ T-cell activation and its regulation in pathological conditions.

overall_design: Sorted CCR7- memory CD8+ T cells from healthy donors (n = 10) were stimulated for 12 hours with IL-15 (10 ng/ml), ionomycin (500 ng/ml), or the combination of both. Total RNA was isolated and RNA-seq was performed.

first: Hoyoung

last: Lee

first: Su-Hyung

last: Park

first: Hyuk Soo

last: Eun

first: Eui-Cheol

last: Shin


title: The E3 ubiquitin ligase LTN1 downregulates RNF10 level and uS3 ubiquitination

city: Minato-Ku

first: Satoshi

last: Hashimoto

address: 4-6-1 shirokanedai

institute: The University of Tokyo

summary: During gene expression, ribosome stalling frequently occurs and can lead to detrimental effects on cellular homeostasis. Several quality control mechanisms, including RQC and NRD, have been identified to resolve these aberrant translation events. While the molecular mechanisms of each pathway have been extensively characterized, the mechanisms underlying the mutual regulation of these factors' expression remain to be elucidated. In this study, we employed a series of knockout mouse and human cell lines to investigate the crosstalk between translational quality control factors. Our findings revealed that LTN1 suppresses RNF10 expression in a manner dependent on the RING domain of LTN1. This discovery offers new insights into the coordination of translational surveillance pathways.

overall_design: Riboseq of MIN6, expressing sgNT or sgLTN1. 80% confluent cells were washed twice with PBS and lysed in Ribo-seq lysis buffer. 40 µg of RNA was mixed with RNase I at 25 °C for 45 min. Ribosome fraction was purified by sucrose cushion with ultracentrifuge at 542,715 × g for 1 h at 4 °C in a TLA-110 rotor. The ribosomal pellet was resuspended in 120 µL splitting buffer to dissociate ribosomes from mRNA fragments, then passed through a 100 kDa MWCO Amicon Ultra-0.5 filter. Approximately 100 µL of the non-ribosomal flow-through was mixed with 300 µL TRIzol, and RNA fragments were purified.

first: Yuxi

last: Huang

first: Sota

last: Ito

first: Chisato

last: Kikuguchi

first: Miho

last: Hoshi

first: Kiyoshi

last: Yamaguchi

first: Yoichi

last: Furukawa

first: Toru

last: Suzuki

first: Toshifumi

last: Inada


title: MIF Nuclease Inhibition Protects CNS Neurons following Inflammation

first: Matthew

last: Smith

summary: In its early phases, multiple sclerosis (MS) is characterized by relapses mediated by immune cell infiltration into the central nervous system, which can mostly be managed with existing therapies. However, there is concomitant axonal injury and neuronal loss associated with these exacerbations which gradually accumulate and cause worsening of disability. This neurodegenerative pathology is not directly targeted by existing therapies. The precise mechanism(s) of cell death that neurons undergo in MS remains unclear. Here we show that parthanatos, a recently described caspase-independent, DNA damage-induced cell death program contributes to neuron death in a mouse model of autoimmune neuroinflammation, experimental autoimmune encephalomyelitis (EAE). We show that DNA damage increases in neurons during EAE, and that they are progressively lost over the disease duration. Furthermore, at early timepoints the inflamed neurons express markers of parthanatos, including accumulation of cytoplasmic poly-ADP-ribose (PAR) and nuclear macrophage migration inhibitory factor (MIF). Genetic or pharmacologic blockade of the final step in parthanatos, MIF nuclease, reduces neuron loss and disease severity. Transcriptomic characterization of these neurons reveals parthanatos-dependent differences in response to EAE. Together, this work establishes parthanatos as a key mechanism of neuron cell death during neuroinflammation.

overall_design: Mice with a homozygous point mutation neutralizing MIF nuclease activity (MIF-E22Q) as well as wild type controls were immunized with MOG35-55 to induce experimental autoimmune encephalomyelitis (EAE). At day 14 lumbar spines were collected from WT EAE (n=3), MIF-E22Q EAE (n=3), WT CFA controls (no mog peptide) (n=2), and MIF-E22Q CFA controls (n=2). Nuclei were isolated and subjected to probe based single nucleus transcriptomics.

first: Sachin

last: Gadani

first: Jackson

last: Mace

first: Peter

middle: A

last: Calabresi


title: Maresin 1 Modulates Cardiac and Renal Lipid Profiles in Hypertensive Rats

city: Tampa

first: Olha

last: Kravtsova

state: Florida

address: 560 Channelside Dr

institute: University of South Florida

summary: BACKGROUND: Chronic inflammation contributes significantly to hypertension and associated target organ damage, particularly in the heart and kidneys. Specialized pro-resolving mediators (SPMs), a class of bioactive lipids primarily derived from omega-3 fatty acids, play key roles in resolving inflammation and maintaining tissue homeostasis. Among them, Maresin 1 (MaR1) has been implicated in cardiovascular regulation and blood pressure control. We hypothesized that MaR1 may mitigate salt-induced hypertension and its related effects in Dahl salt-sensitive (SS) rats. This study evaluated the impact of MaR1 on blood pressure progression, cardiac function, fibrosis, lipid metabolism, gene expression, and circadian rhythms in SS rats fed a high-salt diet. METHODS: SS rats were fed a high-salt diet and treated with MaR1. Mean arterial pressure (MAP) and heart rate (HR) were continuously monitored. Echocardiography and histology were used to assess cardiac structure, contractility, and fibrosis. Lipidomic profiling quantified inflammation-resolving lipid mediators, and transcriptomic analysis identified organ-specific gene expression changes. RESULTS: MaR1 treatment did not significantly alter MAP, HR, or cardiac structure and function. Echocardiographic and histological evaluations showed no significant changes in cardiac remodeling, contractility, or collagen deposition in the heart or kidney. However, lipidomic profiling revealed shifts in inflammatory lipid mediators, suggesting immunomodulatory and metabolic effects of MaR1. Transcriptomic analysis demonstrated organ-specific gene expression changes, with upregulation of circadian and metabolic pathways in the heart and modulation of immune signaling in the kidney. Notably, MaR1 influenced circadian blood pressure rhythms, enhancing amplitude and shifting the acrophase, consistent with altered expression of circadian clock genes. CONCLUSIONS: Although MaR1 did not affect hypertension development directly, its modulation of lipid metabolism, inflammatory pathways, and circadian regulation suggests therapeutic potential. Future studies should explore extended treatment durations and combination strategies to fully evaluate its role in cardiovascular and renal disease management.

overall_design: Male animals were used for the study.  At 8 weeks of age animals were implanted with a telemeter to measure chronic blood pressure. After a few days of recovery animals were switched to high salt (HS, 4% NaCl) diet for 3 weeks. Maresin 1 (MaR1, No. 1268720-28-0, Cayman Chemical Company) was administered subcutaneously via an implanted osmotic minipump (2ML4, ALZET Osmotic Pump) to deliver either the vehicle control or Maresin 1 at a dosage of up to 1.3 µg/kg/day continuously over 3 weeks HS challenge. After 3 weeks on HS, animals were anesthetized with isoflurane and kidneys were collected. Kidney cortex from one kidney and left ventricle of heart  were snap-frozen and used for transcriptomic analysis to compare gene expression in vehicle and MaR1-treated groups.

first: Alexander

last: Staruschenko


title: Neuronal hyperactivity becomes mTORC1 independent due to epigenetic changes in Tuberous Sclerosis Complex [RNA-seq]

city: Boston

first: Kellen

last: Winden

state: MA

address: 300 Longwood Ave

institute: Boston Children's Hospital

summary: Tuberous Sclerosis Complex (TSC) is caused by variants in either TSC1 or TSC2, which cooperate to inhibit the mechanistic target of rapamycin complex 1 (mTORC1). TSC is associated with neurological disorders that have been attributed to disinhibition of mTORC1, but mechanisms connecting dysregulation of mTORC1 to molecular and physiological changes in neurons remain unclear. In this study, we sought to understand transcriptional changes in TSC and identified down-regulation of the immediate early gene, EGR1, in TSC2-deficient excitatory neurons. Furthermore, we found that activity-dependent transcription is impaired in TSC due to abnormalities in epigenetic maturation. Finally, we determined that mTORC1 inhibition started late in neuronal maturation of human neurons is only partially effective in reversing gene expression changes and ineffective in reducing spontaneous neuronal hyperactivity in TSC. These data demonstrate a critical window in early brain development where mTORC1 dysregulation leads to epigenetic changes that contribute to persistent neuronal abnormalities.

overall_design: iPSC neurons of three different genotypes (TSC2+/+, TSC2+/-, and TSC2-/-) were differentiated using induced neurogenin-2 induction. Neurons were re-plated with astrocytes on 48 well multielectrode arrays. Cells were treated with either rapamycin 20nM or DMSO starting at either day 10 or 30 of differentiation. Spontaneous neuronal activity was monitored, and then on day 40 of differentiation, cells were lysed and RNA was extracted.

first: Wardiya

last: Afshar-Saber

first: Juan

last: Ruiz

first: Isabel

last: Gisser

first: Ziqin

last: Yang

first: Leena

last: Mehendale

first: Nicole

last: Teaney

first: Madison

last: Glass

first: Truc

last: Pham

first: Elizabeth

last: Bainbridge

first: Mustafa

last: Sahin


title: Neuronal hyperactivity becomes mTORC1 independent due to epigenetic changes in Tuberous Sclerosis Complex

overall_design: Control (Tsc2fl/fl SynCre-) and mutant (Tsc2del/fl SynCre+) mice were sacrificed at P30. The frontal cortex was dissected out and processed to obtain nuclei. NucSeq was performed using the inDrops platform.


title: Screening for the telomerase RNA component in Caenorhabditis elegans by FLAG-TRT-1 eCLIP.

city: Kobe

first: Hiroki

last: Shibuya

state: Hyogo

address: 2-2-3 Minatojima-minamimachi

department: BDR

institute: RIKEN institute

summary: The telomerase RNA component has been unidentified in C. elegans. To identify the telomerase RNA, we generated a C. elegans strain overexpressing FLAG-TRT-1, the catalytic subunit of telomerase, and biochemically screened for FLAG-TRT-1-binding RNAs by enhanced crosslinking and immunoprecipitation (eCLIP) assay.

overall_design: eCLIP studies were performed by Eclipse Bioinnovations Inc (San Diego, www.eclipsebio.com) according to the published single-end CLIP protocol.

first: Jingjing

last: Zhang

first: Masahiro

last: Onoguchi

first: Michiaki

last: Hamada


title: Effect of LPS induced inflammation on placenta [RNA-Seq]

first: Myoung Seung

last: Kwon

address: 291, Daehak-ro, Yuseong-gu, Biomedical research center (E7)

summary: To investigate the effect of maternal inflammation during pregnancy on placenta, we injected LPS to pregnant dams at embryonic day 14 and analyzed gene expression.

overall_design: Placenta were havested four hours after PBS or LPS injection. Total RNA were anaylzed.

first: Myoungseung


title: Novel lipid nanoparticle adjuvants vaccine for porcine reproductive and respiratory syndrome (PRRS) using in-vitro macrophage assays

city: Vancouver

first: Robert

middle: E. W.

last: Hancock

state: British Columbia

address: 2259 - Lower Mall Research Station

department: Department of Microbiology & Immunology

country: Canada

institute: University of British Columbia

summary: A novel triple adjuvant (TriAdj) platform formulated in various cationic lipid nanoparticles (L-TriAdj) was tested in-vitro for use in porcine reproductive and respiratory syndrome virus (PRRSV) vaccines. Gene expression changes in a RAW 264.7 macrophage cell line were evaluated (compared to media control) after 12-hour stimulation with: (i) TriAdj alone, (ii) lipid formulations alone, (iii) L-TriAdj (lipid:TriAdj ratio of 3:2 or 2:1), and (iv) L-TriAdj + PRRSV antigen. Tested L-TriAdj formulations were comprised of (i) dimethyldioctadecylammonium bromide (DDAB) and 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE) in a 55:45 ratio (abbreviated as DDAB/DOPE), or (ii) DDAB/DOPE with 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC), and cholesterol in a 40:30:20:10 ratio (abbreviated as DDAB/DOPE/DSPC/CHOL).

overall_design:
Cultured RAW264.7 macrophage cells were stimulated for 12 hours with TriAdj, L-TriAdj, lipid carriers alone, inactivated PRRSV and whole vaccines (adjuvant+PRRSV).
RNA was isolated and sequenced to evaluate gene expression changes after 12 hour stimulation.

first: Aderonke

last: Ayanniyi

first: Bhavjinder

middle: K

last: Dhillon

first: Adrienne

last: Nagy

first: Volker

last: Gerdts

first: Ellen

last: Wasan