title: Astrocyte glucocorticoid receptor signaling restricts neuronal plasticity [SHARE-seq]

first: Bruno

last: Gegenhuber

address: 220 Longwood Ave

institute: Harvard Medical School

summary: Sensory experience refines neural circuits during critical periods of postnatal development. Although neuronal activity is known to orchestrate the circuit wiring that underlies this process, the environmental cues that restrain developmental plasticity as animals mature are less clear. Here, we examine the experience-dependent maturation of the mouse primary visual cortex (V1) across postnatal development using paired single-cell transcriptomic and chromatin accessibility sequencing. In addition to identifying the activity-dependent gene programs that emerge within each cortical cell type, we find that light exposure drives astrocyte maturation through cell type-specific recruitment of the glucocorticoid receptor (Nr3c1/GR) to chromatin. Astrocyte GR signaling activates an extensive gene regulatory program that is partially conserved in human brain development and promotes maturation processes that may trigger critical period closure. Collectively, these findings reveal that astrocyte GR signaling restricts neuronal plasticity. Glucocorticoid regulation of astrocyte maturation may also contribute to the effects of early-life stress across the brain, and the disruption of this process may increase susceptibility to neuropsychiatric disease.

overall_design: Simultaneous high-throughput ATAC and RNA expression with sequencing (SHARE-seq) was performed on V1 harvested from P7, P14, P21, P28, and P35 C57BL/6 mice raised in dark-reared (DR) or normally-reared (NR conditions (n = 4 mice per group).

first: Takuma

last: Sonoda

first: Lisa

last: Traunmüller

first: Christopher

last: Davis

first: Shon

last: Koren

first: Eric

last: Griffith

first: Chinfei

last: Chen

first: Michael

last: Greenberg


title: Astrocyte glucocorticoid receptor signaling restricts neuronal plasticity

status: Public on Mar 20 2026

city: Boston

state: Massachusetts

department: Neuroscience

country: USA

type: Expression profiling by high throughput sequencing

type: Genome binding/occupancy profiling by high throughput sequencing

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

overall_design: Refer to individual Series

middle: P

middle: A

middle: C

middle: E


title: Astrocyte glucocorticoid receptor signaling restricts neuronal plasticity [snRNA-seq]

overall_design: snRNA-seq was performed on V1 tissue collected from P21 GRfl/fl animals injected at birth with AAV2/5-GfaABC1D-ΔCre-T2A-EGFP-CAAX-4x6T-WPRE (GR-Con) or AAV2/5-GfaABC1D-Cre-T2A-EGFP-CAAX-4x6T-WPRE (GR-KO) and then reared in DR or NR conditions. 2 biological replicates were collected per condition.


title: Gene expression profiles at the single-nucleus level in heart tissue after myocardial infarction and saNppa-LNP treatment

city: New York

first: Kaiyue

last: Zhang

state: NY

address: 3960 Broadway

department: BME

institute: Columbia University

summary: This project aims to investigate the cellular and molecular mechanisms underlying the therapeutic effects of a self-amplifying RNA encoding atrial natriuretic peptide (saNppa) in myocardial infarction (MI). In this study, a myocardial infarction model was established in C57BL/6 mice by coronary artery ligation. On the day of surgery, mice received intramuscular injection of saNppa formulated in lipid nanoparticles (saNppa-LNP) or phosphate-buffered saline (PBS) as control. Single-nucleus RNA sequencing (snRNA-seq) was performed on left ventricular tissue collected at Day 7 post-MI and treatment. Three experimental groups were included: sham-operated mice, MI mice treated with PBS (MI+PBS), and MI mice treated with saNppa-LNP (MI+saNppa-LNP). This dataset provides a comprehensive single-nucleus transcriptional landscape of the post-infarct heart and enables cell type–specific analysis of transcriptional changes associated with myocardial infarction and saNppa self-amplifying RNA therapy.

overall_design: This study consisted of three experimental groups: sham-operated mice, myocardial infarction (MI) mice treated with phosphate-buffered saline (MI+PBS), and MI mice treated with saNppa lipid nanoparticles (MI+saNppa-LNP). Each group included three mice. Myocardial infarction was induced in C57BL/6 mice by coronary artery ligation, while sham-operated mice underwent the same surgical procedure without ligation. Left ventricular tissues were harvested at Day 7 post-surgery and treatment for all groups. For each group, left ventricular samples from three mice were pooled prior to single-nucleus isolation and downstream single-nucleus RNA sequencing. Pooling was performed to obtain sufficient nuclei yield and to reduce inter-individual technical variability.

first: Hongyan

last: Tao

first: Dashuai

last: Zhu

first: Ke

last: Cheng


title: Chronic stress induces neural stem cell quiescence in the hippocampus by repressing Ascl1 expression [RNA-seq]

city: Xi 'an

first: Yuan

middle: Jia

last: Lan

state: Shaanxi Province

address: Yanta West Road

country: China

institute: The First Affiliated Hospital of Xi 'an Jiaotong University

summary: Chronic stress inhibits neurogenesis, yet its impact on neural stem cells (NSCs) remains poorly understood. Here, using the 5-bromo-2′-deoxyuridine (BrdU) label-retaining assay, we found that chronic restraint stress (CRS) in adult mice promoted quiescence in hippocampal NSCs (i.e. radial-glial-like cells, RGLs). Long-term administration of the synthetic stress hormone agonist dexamethasone (Dex) to adult mice recapitulated the phenotype. Moreover, pre-administration of the antagonist mifepristone (RU486) prevented RGLs from quiescence. At the cellular level, Dex induced reversible quiescence in hippocampal NSCs (HpNSCs) in a manner similar to the quiescence-promoting signal BMP4 in vitro. However, Dex and BMP4 regulated overlapping yet distinct NSC quiescence programs, with their co-regulation primarily being synergistic. Mechanistically, Dex downregulated the expression of achaete-scute homolog 1 (Ascl1) by repressing a distal enhancer. These data suggest that chronic stress induces quiescence in RGLs by repressing the transcription of the quiescence regulator gene Ascl1, and that synthetic glucocorticoid antagonists may have therapeutic value in correcting abnormalities of NSC activity in conditions associated with elevated cortisol levels, including psychological and mental health conditions.

overall_design: To explore the functional differences between Dex and BMP4 in HpNSC, we treated HpNSCs with 10-6 M Dex and/or 20ng/mL BMP4 for 3 days. Three replicates were set for each treatment.

first: Wan

first: Jialan

last: Yuan

first: Hang

last: Peng

first: Huanhuan

last: Dong

first: Jing

last: Liu

first: Feng


title: Hyperinnervation inhibits organ-level regeneration in mammalian skin

city: Cambridge

first: Hannah

middle: T

last: Tam

state: MA

address: 7 Divinity Avenue

department: SCRB

institute: Harvard University

summary: Some mammalian tissues can replace lost cells within one lineage, but organ-level regeneration—restoring diverse cell types across lineages—remains rare. Here we show that late embryonic full-thickness skin injuries heal by regenerating epithelial, mesenchymal, neuronal, and vascular lineages with proper connectivity. However, this ability is lost soon after birth, leading to a failure to restore most cell types and hyperinnervation of the wound bed. Single-cell sequencing identified a postnatal wound-specific fibroblast (PWF) population absent after embryonic wounding. Through an in vivo screen, we discovered three PWF-enriched genes—Timp1, Cxcl12, and Ccl7—that inhibit organ-level regeneration and cause hyperinnervation when overexpressed in embryonic wounds. Reducing hyperinnervation by depleting Cxcl12 in fibroblasts or inhibiting synaptic release enables postnatal skin to regenerate diverse lineages after injury. Our study identifies mechanisms transitioning an organ from regenerative to non-regenerative, discovers fibroblast-driven hyperinnervation as a key barrier, and demonstrates removing this barrier unlocks organ-level regeneration.

overall_design: We first sought to identify the changes that drive the distinct regenerative responses after wounding at E16.5 vs. P5. For this, we performed scRNAseq on E16.5 wounded, P5 wounded, and unwounded age-matched controls skin (2 biological replicates each). We sequenced the wounds 3 days after wounding to capture changes that precede and can potentially lead to the distinct outcomes observed 7 days after wounding.  Second, we sought to test specific genes that might inhibit regeneration in vivo with an AAV-based strategy. To assess which cell types are transduced by AAV, we FACS-purified and single-cell sequenced AAV-positive and -negative cells collected 3 days after E16.5 wounding and AAV transduction.

first: Jingyu

first: Rebecca

last: Freeman

first: Yulia

last: Shwartz

first: Shlomi

last: Brielle

first: Sakshi

last: Garg

first: Siti

last: Rahmayanti

first: Stephen

last: Crocker

first: Devin

last: Coon

first: Ya-Chieh

last: Hsu


title: Chronic stress induces neural stem cell quiescence in the hippocampus by repressing Ascl1 expression [CUT&Tag]

first: Yuan

overall_design: Cleavage under Targets and Tagmentation (CUT&Tag) and sequencing was used to detect GR binding sites on the genome in HpNSC cells. IgG was served as the control.

last: Zhang


title: CarS2 mediated cysteine catabolism drives brown fat development and thermogenesis through persulfidating EBF2 [Cut & Tag]

city: Shanghai

first: Xuyun

last: Zhao

state: Shanghai

address: South Chongqing Road

department: Medical school

institute: Shanghai Jiaotong University

summary: Targeting transcriptional regulatory complex is a promising strategy to activate thermogenic fat and treat  obesity. In this study, we focus on identifying critical and special metabolic pathways enriched during brown adipocyte differentiation. By analyzing metabolomics and RNA sequencing results from precursor and mature brown and white adipocytes, we revealed cysteine catabolism pathway is a unique metabolic process that is exclusively increased in the brown adipocyte differentiation. Then, we demonstrated cysteine persulfidation synthase CarS2 is directly induced by EBF2 and mediates cysteine catabolism to produce cysteine persulfide and triggers brown fat protein persulfidation in vitro and in vivo. Loss of CarS2 in thermogenic fat blocks brown and beige fat formation and reduced thermogenesis and energy expenditure in mice. More importantly, CarS2 generates cysteine persulfide and its derivative H2S cell autonomously induces brown adipogenesis and enhances uncoupled respiration by persulfidating EBF2. Mechanistically, the persulfidated EBF2 facilitates its interaction with PPARγ and enhanced the recruitment of the EBF2-PPARγ complex on browning gene promoter, thus driving brown fat development and boosting thermogenic function. Furthermore, the treatment of CarS2 coenzyme PLP or H2S donor elevates brown adipocyte function and ameliorates obesity progression in mice under HFD feeding, indicating a novel metabolite-based strategy for the obesity treatment.

overall_design:
In our study, we found EBF2 is the substrate of CarS2, and was persulfidated in the BAC differentiation.To investigate the role of CarS2 on EBF2 transcription activity in  the BAC differentiation, we seperated and immortalized primary brown adipocytes from CarS2 floxed mice and expressed vector (BAC-V) for control and  a cre recombinase to knock out CarS2 (BAC-cre).
BAC-V and BAC-cre cells were cultured and differentiated to performe Cut & Tag assay by using EBF2 antibody (R&D, AF7006) and H3K27ac antibody(CST,4353).The DNA libiray was sequenced.
we performed promoter binding profile analysis using data obtained from Cut & Tag-seq of BAC-V and BAC-Cre.

first: Hongyong

last: Song

first: Xin


title: HnRNP C binding to inverted Alu elements protects the transcriptome from pre-mRNA circularization

city: Paris

first: Pierre

last: de la Grange

address: 29, rue du Faubourg Saint-Jacques

department: Paris Biotech Santé

country: France

institute: GenoSplice technology

summary: Back-splicing is a non-canonical splicing event that drives the biogenesis of circular RNAs (circRNAs). Although the molecular mechanisms underlying circRNA biogenesis have been partially elucidated, how this process is globally regulated in tumors that show a general repression of circRNAs, such as medulloblastoma (MB), has not been fully investigated. Herein, we uncover a hnRNP C-dependent mechanism that represses a broad repertoire of circRNAs in Group 3 MB. HnRNP C binds Alu elements and prevents non-colinear splicing, thus repressing circularization of pre-mRNA transcripts. Expression of hnRNP C modulates the balance between linear and circular splicing and guarantees efficient expression of genes that sustain the oncogenic phenotype of Group 3 MB cells. Remarkably, in the absence of hnRNP C, the introns flanking the circularizing exons generated cytoplasmic dsRNAs through base-pairing of inverted Alu elements and, consequently, triggered an interferon-induced antiviral defense mechanism. These findings unveil the role of hnRNP C as guardian of transcriptome integrity by repressing circRNA biogenesis and suggest that targeting hnRNP C in Group 3 MB may be used to trigger an inflammatory immune response, thereby boosting cancer surveillance.

overall_design: RNA-sequencing of D341 MB cells depleted or not for the expression of hnRNP C. Treatment of the RNA samples with RNaseR was performed to enrich for circRNAs while removing linear RNAs.

first: Alberto

last: Marini

first: Consuelo

last: Pitolli

first: Sabrina

last: Ciccone

first: Marco

last: Pieraccioli

first: Noémie

last: Robil

first: Chiara

last: Naro

first: Fernando

last: Palluzzi

first: Manuela

last: Giansanti

first: Gianpiero

last: Tamburrini

first: Luciano

last: Giacò

first: Francesca

last: Nazio

first: Claudio

last: Sette

first: Vittoria

last: Pagliarini


title: Collective unstructured interactions drive chromatin binding of transcription factors.

city: Berkeley

first: Robert

last: Tjian

state: CA

address: University of California, Berkeley 450 Li Ka Shing #3370

department: MCB

institute: UC Berkeley


title: CarS2 mediated cysteine catabolism drives brown fat development and thermogenesis through persulfidating EBF2

first: Xuyun

last: Zhao


title: Endometrial Insights: Unmasking Endometriosis through Single-Cell Profiling and AI-Based Prediction

city: Berne

first: Lea

last: Duempelmann

address: Murtenstrasse 35

department: DBMR

country: Switzerland

institute: University of Berne

summary: Endometriosis, affecting over 10% of women, presents treatment and diagnostic challenges. To address these issues, we generated the biggest single-cell atlas of endometrial tissue to date, comprising 466,371 cells from 35 endometriosis and 25 non-endometriosis patients without exogenous hormonal treatment. Detailed analysis reveals significant gene expression changes and altered receptor-ligand interactions present already in the endometrium of endometriosis patients, including increased inflammation, adhesion, proliferation, cell survival, and angiogenesis in various cell types. These alterations may enhance endometriosis lesion formation and offer novel therapeutic targets.  Using ScaiVision neural networks, we developed accurate models predicting endometriosis of varying disease severity, including a minimal 11-gene signature-based model. In conclusion, our findings illuminate numerous pathway and ligand-receptor changes in endometriosis endometrium, offering insights into pathophysiology, targets for novel treatments and accurate diagnostic models for enhanced outcomes in endometriosis management.

overall_design: Endometrial biopsies were acquired from participants with and without endometriosis. After dissociating the biopsies into single cells, the cellular content was captured using 10x Genomics technology and sequenced. Subsequently, the resulting data was compiled into an endometrial single-cell atlas.

first: Shaoline

last: Sheppard

first: Angelo

last: Duo

first: Jitka

last: Skrabalova

first: Brett

last: McKinnon

first: Thomas

last: Andrieu

first: Dennis

last: Goehlsdorf

first: Sukalp

last: Muzumdar

first: Cinzia

last: Donato

first: Ryan

last: Lusby

first: Wiebke

last: Solass

first: Hans

last: Bösmüller

first: Peter

last: Nestorov

first: Michael

middle: D

last: Mueller


title: Astrocyte glucocorticoid receptor signaling restricts neuronal plasticity [ATAC-seq]

first: Bruno

last: Gegenhuber

overall_design: ATAC-seq was performed on V1 astrocyte nuclei isolated from P21 GRfl/fl animals that were injected at P0-P1 with AAV2/5-GfaABC1D-ΔCre-T2A-EGFP-KASH-4x6T (GR-Con) or AAV2/5-GfaABC1D-Cre-T2A-EGFP-KASH-4x6T (GR-KO).

first: Takuma

last: Sonoda

first: Lisa

last: Traunmüller

first: Christopher

last: Davis

first: Shon

last: Koren

first: Eric

last: Griffith

first: Chinfei

last: Chen

last: Greenberg


title: Microtubule anchoring and coupling of CD20 to the RhoA/Rock1 pathway

city: Freiburg

first: Geoffroy

last: Andrieux

address: Breisacherstr 153

country: Germany

institute: University clinics Freiburg

summary: CD20 is a B cell-specific four-helix transmembrane protein and a prominent target of therapeutic anti-CD20 antibodies. We have recently described that CD20 is localized within a membrane nanocluster harboring the IgD class B cell antigen receptor (IgD-BCR) where it functions as a gatekeeper for the resting state of naïve human B lymphocytes. How CD20 exerts its gatekeeper function is not yet known. Using Ramos and human peripheral blood B cells, we show here that the serine/threonine kinase PKCδ, constitutively phosphorylates serine residues at the cytosolic tails of CD20. The phosphorylated CD20 becomes a target for 14-3-3 adaptor proteins that link CD20 to the RhoA GDP/GTP exchange factor GEF-H1 and the microtubule (MT) network controlling the stability of the IgD-BCR nanocluster. Binding of anti-CD20 antibodies results in MT disassembly and the replacement of the GEF-H1/CD20 complex by a RhoA-GTP/ROCK1/CD20 complex which drives actomyosin contractility. Our study suggests that CD20 not only maintains the resting state, but also orchestrates the MT/actin switch in active B lymphocytes. This could have implications for treatment with anti-CD20 antibodies and may help to optimize therapeutic protocols

overall_design: To investigate the gatekeeper function of protein kinase C delta (PKCδ) we generated two different Ramos PKCδ KO cell lines using CRISPR-Cas system, exploring the differences in regulation of activation on B lymphocytes.

first: Cindy

last: Lee

first: Claudio

last: Mussolino

first: Klaus

last: Warnatz


title: p53 safeguards chemical reprogramming of human somatic cells towards pluripotency [scRNA-seq]

city: Beijing

first: Jingxiao

last: Cao

state: Beijing

address: No.5 Yiheyuan Road, Yanyuan Street, Haidian District, Beijing

department: Academy for Advanced Interdisciplinary Studies, Peking University, Beijing, China.

institute: Peking University

summary: Cell fate manipulation is powerful for generating desired cell types through reprogramming, such as induced pluripotent stem cells (iPSCs), with broad biological applications. However, reprogramming can be risky due to dramatic changes in cell identity, necessitating strict regulation to ensure safety and efficacy. p53 is essential for maintaining genome stability to prevent abnormalities induced by changes of cell states, which however, functionally counters Yamanaka factors for efficient reprogramming. Thus, delicately balancing p53 activity for efficient reprogramming with highly quality has proven challenging. Here, we demonstrate that efficient human chemical reprogramming is safeguarded by p53, in distinction from Yamanaka factors-mediated reprogramming. Suppressing p53 activity unexpectedly and substantially hinders the generation of chemically induced pluripotent stem cells (CiPSCs). Notably, p53 precisely regulates the mesenchymalization occurred at the early reprogramming stage where cell plasticity is re-established and curbs excessive epithelial-mesenchymal transition (EMT). Furthermore, the retinoid acid receptor agonist, TTNPB, leverages p53’s anti-metastatic functions to facilitate CiPSC generation by activating BTG2. Our results demonstrate that p53 activity distinguishes chemical reprogramming from Yamanaka factors-mediated reprogramming, and offer a promising strategy that delicately balances p53 with reprogramming for both safe and efficient cell fate manipulation.

overall_design: To characterize samples during  human chemical reprogramming by scRNA-Seq.


title: ZNF638 represses HBV covalently closed circular DNA transcription involving HUSH complex epigenetic silencing

first: Sifan

last: Meng

address: Beijing

institute: Capital Medical University

summary: Hepatitis B virus (HBV) infection represents a significant public health burden worldwide. Covalently closed circular DNA (cccDNA) is the transcriptional template of HBV, which interacts with both host and viral proteins to form minichromosome in the nucleus and exhibits resistance to antiviral agents. Here, we provide evidence that ZNF638 mediates transcriptional silencing of HBV cccDNA. Mechanically, ZNF638 bound to HBV cccDNA, increasing the occupancy of HUSH complex to target loci, which subsequently recruited SETDB1, leading to enhanced H3K9me3 modification, thereby repressing cccDNA transcription. Our research findings improved the existing understanding of ZNF638- and HUSH-mediated epigenetic silencing on viral DNA, that (1) ZNF638 and HUSH complex inhibited HBV transcription, not only by epigenetic silencing extrachromosome cccDNA but also silencing the integrate viral DNA, and (2) ZNF638- and HUSH-mediated repression appeared to function as an epigenetic rheostat: occupancy of HUSH components at the locus inhibited transcription; release from the target locus activated transcription. In summary, the study demonstrates a novel epigenetic role of ZNF638 and HUSH complex in negatively regulating HBV cccDNA transcription by deposit repressive H3K9me3 histone marks and highlights the importance of epigenetic therapy targeting cccDNA, suggesting that ZNF638 could be a potential target for epigenetic therapy of viral diseases.

overall_design: Removal of ChIP Seq (ZNF638, MPP8) in HepAD38 cells expressing HBV induced by tetracycline and HepAD38 cells not expressing HBV induced by tetracycline


title: RNA-seq results in the liver of male hepatocyte-specific GAPDH knockout mice in fasting supplemented with or without serine

city: Wuhan

first: Yihao

last: Zhou

state: Hubei

address: Luojiashan Road

department: College of Life Sciences

institute: Wuhan university

summary: Our study represents the first detailed analysis of transcriptomes in liver of  hepatocyte-specific GAPDH knockout mice to understand key pathways and regulatory genes, with biologic replicates, generated by RNA-seq technology

overall_design: Each sample consists of a pool of RNAs extracted from liver tissue of 2 different 12-week-old male mice. For each experimental group, two biological replicates of RNA samples were sent for RNA-seq.

first: Kun

last: Huang

first: Ling

last: Zheng


title: N6-methyladenosine modulate second cell fate decision by enhancing mRNA stability of pivotal transcription factors during primitive endoderm development

first: Li

last: Tang

address: No.5 Yiheyuan Road Haidian District


title: Mitochondria-lysosome coupling contributes to lysosome acidification and aging

city: Novato

first: Qingqing

last: Liu

state: California

address: 8001 Redwood Blvd

institute: Buck Institute - Novato, CA

summary: Nearly all cellular processes are pH dependent. The acidic pH inside the lysosome (vacuole in yeast) is essential for cellular content degradation, signaling, and autophagy. Defect in lysosome/vacuole acidification is a conserved hallmark of aging and age-related diseases. Traditionally, lysosome/vacuole is thought to import free protons (H⁺) from the surrounding neutral cytosol. In this study, we uncovered a previously unrecognized, conserved lysosome/vacuole acidification mechanism, involving lysosomal/vacuolar uptake of H+ pumped out by mitochondrial electron transport chain through membrane contacts between mitochondria and lysosomes/vacuoles. Aging/senescence-associated disruption of mitochondria-lysosome/vacuole contacts causes lysosomal/vacuolar de-acidification, which can be reversed by expressing a linker to connect these organelles and through an asymmetry-dependent rejuvenation process in daughter cells. Preserving lysosomal acidification in senescent human cells prevents the induction of major senescence-associated secretory phenotype factors and enhances autophagic flux. These findings reshape our current understanding of the mechanisms underlying lysosomal/vacuolar (de-)acidification in both young and aged/senescent cells.

overall_design: Senescent IMR90 cells with and without an engineered Mito-Lyso linker are induced by exposing to a single dose of 10 Gray ionizing radiation (IR) for 9 minutes according to the treatment condition in Campisi lab. After IR, placed cells back to the incubator and changed medium every 48 hrs. Assays for senescent cells were done 10 days after IR. Pre-senescent IMR90 cells are with less than 15-17 passages.

first: Sungmin

last: Yoo

first: Alice

last: Zhang

first: Liying

last: Li

first: Hetian

last: Su

first: Lingraj

last: Vannur

first: Chuankai


title: BRD8 is a therapeutic vulnerability for overcoming resistance to dual ER/HER2 blockade therapy in HR+/HER2+ breast cancer [scRNA-seq]

city: Madison

first: Huy

last: Dinh

state: WI

address: 1111 Highland Avenue

institute: University of Wisconsin-Madison

summary: Hormone receptor (HR)-positive, HER2-positive breast cancers are resistant to endocrine and anti-HER2 therapies due to crosstalk between estrogen receptor (ER) and HER2. However, how anti-HER2 agents activate ER as a mechanism of resistance remains unknown. Using single-cell RNA sequencing, we identified Bromodomain Containing Protein 8 (BRD8) as a major mediator of ER activation in response to neratinib, a HER2 tyrosine kinase inhibitor. BRD8 expression was rapidly induced by various anti-HER2 agents (neratinib, lapatinib and trastuzumab) and its expression positively correlates with ER. BRD8 regulates both ER-dependent and -independent growth promoting pathways. Moreover, BRD8 ablation re-sensitizes fulvestrant- or neratinib-resistant HR+/HER2+ cells to neratinib, suggesting that combinatorial targeting of BRD8 and HER2 attenuates signal crosstalk and possibly overcomes treatment resistance to dual anti-ER/HER2 blockade therapy. This work identifies BRD8 as not only a central hub for ER signaling activation upon anti-HER2 treatment, but also a druggable vulnerability for treating HR+/HER2+ breast cancer.

overall_design: Single-cell RNA-seq expression profiling of BT474 breast cancer cell lines with different treatments.

first: Ang

last: Gao

first: Parth

last: Khatri

middle: Q

first: Wei

last: Xu


title: Gene expression profile at single cell level of monocytes sorted from C57BL/6J mouse ear tissue at 3 weeks post Leishmania major infection [scRNA-seq]

city: Magdeburg

first: Yan

last: Fu

state: Saxony-Anhalt

address: Haus 26, Institut für Molekulare und Klinische Immunologie, Otto-von-Guericke-Universität Magdeburg, Leipziger Str. 44

institute: Otto-von-Guericke-Universität Magdeburg

summary: Monocytes are considered as a homogeneous population when encountering intracellular pathogen infections. We used signle cell RNA sequencing (scRNA-seq) to analyze the diversity of monocytes and the interaction with pathogen proliferation status at the infection site of leishmania major.

overall_design: Monocytes from the mKikumeGreen&Red Leishmania major-infected C57BL/6J mouse ears  were sorted by Fluorescence-activated cell sorting (FACS) according to the presence or absence of mKikumeGreen & Red signal and analyzed using scRNAseq. Specifically, there were 70 cells infected with high-proliferating parasites (mKikumeGreen+), 75 cells infected with low-proliferating parasites (mKikumeRed+), and 73 cells are uninfected (mKikume-negative).

first: Christiphe

last: Toussaint

first: Ina

last: Sauerland

first: Ehsan

last: Vafadarnejad

first: Lars

last: Philipsen

first: Patricia

last: Gintschel

first: Laura

last: Knop

first: Jonas

last: Amore

first: Nouria

last: Jantz-Naeem

first: Iris

last: Baars

first: Juliane

last: Mohr

first: Julia

last: Prein

first: Nicole

last: Gröger

first: Andreas

last: Braun

first: Thomas

last: Tüting

first: Sascha

last: Kahlfuß

first: Burkhart

last: Schraven

first: Robert

last: Geffers

last: Joller

first: Antoine-Emmanuel

last: Saliba

last: Schüler

middle: J

last: Müller


title: Landscape of mitochondrial RNA binding proteome [RNAseq_siDHX30_siTFam]

city: Guangzhou

first: Jinshuai

last: Sun

address: Guangzhou Institutes of Biomedicine and Health,Chinese Academy of Sciences

institute: Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences

summary: We combine RNA 5-ethynyluridine labelling, optimized mitochondria isolation, with click chemistry-based affinity purification to systematically capture the RBPs interacting with mitochondria-located RNAs. This methodology, termed mitochondrial RNA interactome using click chemistry (mitoRICK), unexpectedly revealed a rapid turnover of mitochondria-encoded RNAs compared to their nuclear-encoded counterparts. MitoRICK has identified 122 high-confidence and 25 lower-confidence mitochondrial RNA binding proteins, which expanded the repertoire of mitochondrial RNA binding proteome, and provided a new avenue for understanding mitochondrial RNA metabolism or inherited diseases. We found that MOV10, amongst many newly-identified RNA helicases, is a novel mitochondria-associated RBP. Functionally, MOV10 is important for MT-RNR1 expression or unwinding the mitochondrial double-strand RNAs. As a proof-of-concept, we applied mitoRICK to naïve-to-primed pluripotency transition for studying mitochondrial RBP dynamics, suggesting that intact mitochondrial RNA metabolism is essential for the cell fate transition. MitoRICK is broadly applicable to understand dynamics of mitochondrial RNA binding proteome in various cell contexts.

overall_design: RNA profiles of knockdown of Dhx30 and Tfam by siRNA in the Mouse embryonic stem cells.

first: Yingying

last: Lin

first: Xiwei

last: Wangh

first: Xichen

last: Bao


title: Landscape of mitochondrial RNA binding proteome [mitoRICK]

overall_design: RNA signals of the initial proteinase- and nuclease-treated mitochondria (Input) and of samples captured by mitoRICK (Pull-down) without or with EU treatment (Control or MitoRICK).


title: PAF1 N-terminal extension-generated PAF1c-SKIP-CDKB module regulates plant-specific cell cycle in the plant kingdom [RNA-seq]

status: Public on Mar 19 2026

first: Yan

last: Li

address: Haidian

institute: Capital Normal University

summary: Through systematic biochemical analyses, we identified a novel protein-protein interaction between PAF1 and SKIP in Chlamydomonas reinhardtii.  To investigate the functional significance of this interaction, we generated targeted mutants using CRISPR-Cas9 genome editing technology.  Phenotypic characterization revealed that both PAF1 and SKIP mutants exhibit coordinated regulation of the cell cycle progression.  To elucidate the molecular mechanism underlying PAF1-SKIP mediated cell cycle regulation, we employed an integrated multi-omics approach.  RNA sequencing (RNA-seq) analysis was performed to identify differentially expressed genes, while chromatin immunoprecipitation sequencing (ChIP-seq) was utilized to map the genomic loci directly bound by the PAF1-SKIP protein complex.  This comprehensive strategy enabled us to delineate the transcriptional network governed by the PAF1-SKIP regulatory module.

overall_design: Wild-type Chlamydomonas reinhardtii, crpaf1-1, and crskip-1 mutants were cultured in TAP liquid medium under a 14 h light/10 h dark cycle at 22°C for two generations and harvested at late G1 phase for RNA extraction.

first: Renshen

last: Yuan


title: p53 safeguards chemical reprogramming of human somatic cells towards pluripotency.

first: Jingxiao

last: Cao

overall_design: Refer to individual Series


title: Human-induced pluripotent Stem Cell-Derived Hepatic Stellate Cells in Liver Organoids: Maturation Promotion and Essential Role in Vascularization

city: Tokyo

first: Yunzhong

last: Nie

state: Tokyo

address: Room 330, 3F west, 1st  Bldg, Shirokanedai 4-6-1, Minato

department: Department of Computational Biology and Medical Sciences

country: Japan

institute: The University of Tokyo

summary: Human-induced pluripotent stem cell (hiPSC)-derived liver organoids (LOs) are valuable for studying human liver organogenesis but face challenges in faithfully recapitulating certain processes, like vasculogenesis, due to the lack of specific cell components. Hepatic stellate cells (HSCs), which are liver-specific pericytes and might be crucial for liver vasculogenesis, remain underutilized in developmental studies because of their disease-related "activated" status and inefficient generation process. Here, we present an efficient method for generating hiPSC-derived HSCs (hiPSC-HSCs) resembling non-activated HSCs in the healthy human liver. These hiPSC-HSCs exhibit exceptional expandability (>105-fold) while maintaining essential cellular features. Additionally, in entirely hiPSC-derived LOs consisting of HSCs, hepatic endoderm, and endothelial cells, hiPSC-HSCs play a vital role in LO maturation and vascularization, both in vitro and in vivo. This work represents a significant advancement in understanding HSC roles in human liver development, and LOs containing non-activated hiPSC-HSCs hold potential in modeling congenital human liver diseases.

overall_design: mRNA expression profiles of  hiPSC, hiPSC derived septum transversum mesenchyme (hiPSC-STM), hepatic stellate cell (hiPSC-HSC) (with passage), hepatic endoderm (hiPSC-HE), endothelial cells (hiPSC-EC), and human primary HSC (pHSC)

first: Xia

last: Yang

first: Yun-Zhong

last: NIE

first: Hideki

last: Taniguchi


title: Phase Separation of DUX Family Proteins Drives Totipotent-like State via 3D Genome Reorganization and Retrotransposon Activation [CUT&Tag]

city: Shaanxi

first: Qifeng

last: Gao

state: Shaanxi

address: No.3 Taicheng Road, Yangling, Shaanxi, China

institute: Northwest A&F University

summary: The acquisition of totipotency requires transcriptional activation of endogenous retroviruses (MERVL/HERVL) and zygotic genome activation (ZGA) related genes, yet the molecular mechanisms linking chromatin architecture to this process remain elusive. Here, we demonstrate that mouse Dux and human DUX4, double homeobox transcription factors essential for totipotency, form liquid-liquid phase-separated (LLPS) condensates through conserved arginine residues within intrinsically disordered regions (IDRs) in the Homeobox domain. These condensates recruit CBP/p300 and CTCF to establish super-enhancers (SEs) at MERVL/MT2 loci, enabling H3K27ac deposition and chromatin accessibility. Hi-C analysis revealed that DUX-driven phase separation facilitates 3D genome reorganization, including de novo formation of enhancer-promoter loops and TAD boundary shifts. Disruption of LLPS (DUXR70A) abolished SE assembly, transcriptional activation, and embryonic chimerism. Strikingly, human DUX4 required phase separation for both myotoxic gene activation and cytotoxicity in facioscapulohumeral muscular dystrophy (FSHD) models. Our study establishes a paradigm wherein phase separation integrates transcriptional control with 3D genome remodeling to license totipotency, with direct implications for developmental biology and disease therapy.

overall_design: To investigate the transcriptomic consequences of DUX-mediated phase separation and its necessity for the induction of a totipotent-like state, we performed RNA-seq profiling across various experimental conditions. In this study, RNA-seq was used as a primary readout to evaluate how the global gene expression program, particularly the activation of 2C-specific genes and retrotransposons like MERVL, was altered following the overexpression of wild-type DUX. To evaluate the role of Dux across different totipotency induction systems, we performed RNA-seq on Dux-knockdown (KD) cells subjected to RA-induced, PlaB-induced, or Zscan4-induced totipotency programs. This component of the study was designed to assess whether these alternative pathways for inducing a totipotent-like state could bypass the loss of Dux. Furthermore, to evaluate the functional requirement of DUX-induced super-enhancers (SEs) in gene activation, we conducted RNA-seq on Tdpoz-SE knockout (KO) cells during DUX-mediated induction. By comparing these results with Dux-induced control samples, we sought to test whether SEs are indispensable for the activation of key totipotency genes. Finally, to define the specific contribution of liquid-liquid phase separation (LLPS), we performed RNA-seq on cells expressing the phase-separation-deficient mutant (DUXR70A). This design allowed us to measure whether the loss of DUX nuclear condensates would abolish the activation of the totipotent transcriptome, even when the mutant’s DNA-binding capacity remained intact. Collectively, we utilized these datasets to assess how DUX coordinates distal gene expression through SE-mediated architectural reorganization.

first: Leilei

last: Hai

first: Ziqiang

last: Wu

first: Penghui

first: Han

last: Kang

first: Xiaohui

last: Song

first: Jinlian

last: Hua

first: Shiqiang

last: Zhang

first: Gang

last: Ren

first: Jihong

first: Leqian

last: Yu

first: Yulei

last: Wei

first: Junjun

last: Ding

first: Fan


title: CarS2 mediated cysteine catabolism drives brown fat development and thermogenesis through persulfidating EBF2 [AKO and wt Acute cold]

first: Xuyun

last: Zhao

overall_design:
To investigate the role of CarS2 in thermogenic function of brown adipose tissue, we established the CarS2 condition  knock out mice in adipose tissue by cross with adiponectin-cre mice.
AKO mice and wild type littermates were maintained in an 4℃ incubater for 4 hours,then were euthanasia and brown adipose tissue were harvest and frozed immediately with liquid nitrogen.
We performed gene expression profile using data obtained from RNA-seq of BAT from wild type and AKO mice.

first: Hongyong

first: Xin

last: Peng


title: BRD8 is a therapeutic vulnerability for overcoming resistance to dual ER/HER2 blockade therapy in HR+/HER2+ breast cancer [bulkRNA-seq]

first: Huy

last: Dinh

overall_design: RNA-seq expression profiling of BT474 breast cancer cell lines with different treatments.

first: Ang

first: Parth

last: Khatri

first: Wei

last: Xu


title: Phase Separation of DUX Family Proteins Drives Totipotent-like State via 3D Genome Reorganization and Retrotransposon Activation [RNA-Seq]

first: Qifeng

first: Leilei

first: Ziqiang

first: Penghui

last: Li

first: Han

last: Yang


title: p53 safeguards chemical reprogramming of human somatic cells towards pluripotency [bulk RNA-seq]

first: Jingxiao

last: Cao

overall_design:
To characterize samples during  human chemical reprogramming by RNA-seq.
C, 6, N, A, J are abbreviations for five small chemical molecules. Their full names are CHIR, 616452, TTNPB, Akt, and JNKIN8, respectively.

(bulkRNA_4)  To characterize samples during human chemical reprogramming by RNA-seq.


title: Cell-intrinsic effect of IL-7R signaling of monocytes within C57BL/6J mouse ears infected with Leishmania major [bulk RNA-seq]

first: Yan

last: Fu

department: Institut für Molekulare und Klinische Immunologie

summary: To determine the cell-intrinsic effect of IL-7R signaling, we sorted il7r-/- (CD45.2+) and WT (CD45.1+) monocytes as internal controls from the same Leishmania major (Lm) infected ears of 50:50 WT:il7r-/ -bone marrow chimera  and performed population RNAseq of pools of 50-100 cells per population and infection site using the SMARTseq2 protocol.

overall_design:
Monocytes from the mKikumeGreen Leishmania major-infected CD45.1+ C57BL/6J mouse ears  were sorted by Fluorescence-activated cell sorting (FACS) according to CD45.2 (il7r-/-) and CD45.1+ (wt) and analyzed using scRNAseq. Specifically, leishmania fluorescence mKikumeGreen and cell surface markers Ly6c, CD11c,and CCR2 were used to identify subpopulations of monocytes: Hi1 (mKikumeGreen+CD11chiLy6hiCCR2lo-int), Lo3 (mKikumeGreen+CD11cloLy6cloCCR2lo), U1 (mKikume-negCD11chiLy6clowCCR2low), and U2 (mKikume-negativeCD11cloLy6chiCCR2hi). Comparative gene expression profiling analysis of RNA-seq data for il7r-/- and wt monocytes within each subpopulations were performed.
Please note that 'BMC' in the sample titles represents 'bone marrow chimeric'.

first: Christiphe

last: Toussaint

first: Ina

last: Sauerland

first: Ehsan

last: Vafadarnejad

first: Lars

last: Philipsen

first: Patricia

last: Gintschel

first: Laura

last: Knop

first: Jonas

last: Amore

first: Nouria

last: Jantz-Naeem

first: Iris

last: Baars

first: Juliane

last: Mohr

first: Julia

last: Prein

first: Nicole

last: Gröger

first: Andreas

last: Braun

first: Thomas

last: Tüting

first: Sascha

last: Kahlfuß

first: Burkhart

last: Schraven

first: Robert

last: Geffers

last: Joller

first: Antoine-Emmanuel

last: Saliba

last: Schüler

last: Müller


title: IL-7 mediated immunoregulatory feedback loop promotes dampening of immune effector functions by hypoinflammatory monocytes.

first: Yan

last: Fu

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


title: Potential of RAD51C as a hub gene and therapeutic target in bladder cancer

first: Ying

last: Xiao

address: 197 Ruijin 2nd Road, Huangpu District, Shanghai, China

institute: Ruijin Hospital

summary: Abstract Background The global incidence of tumors is rising sharply. Immune checkpoint inhibitors (ICIs) represent the mainstream of immunotherapeutic approaches. Despite the remarkable clinical benefits of immunotherapy in various tumors, ICIs resistance remains an urgent challenge to be addressed. The transcription factors cAMP response element-binding protein 1(CREB1) and c-Jun converge to regulate oncogenesis and immune modulation. Methods Based on the 'CREBP1CJUN_01' gene set from the Molecular Signatures Database and single-cell RNA sequencing data, we identified 116 CREB–c-Jun transcription factor target-related differential genes (CR.Sig) and refined to 20 CREB–c-Jun transcription factor target-related important feature genes (Hub-CR.Sig) through multi-algorithm machine learning. These Hub-CR.Sig enabled prognostic risk modeling, molecular subtyping of bladder cancer, and somatic mutation stratification. Results Among these 20 signatures, RAD51C emerged as a top-ranked driver that was correlated with advanced T/N/M stages, associated with poor survival outcomes and is highly expressed in bladder cancer tissues. Functional assessment revealed that RAD51C knockdown suppressed bladder cancer cell proliferation, invasion, migration, and DNA repair capacity in vitro, while also reducing tumor formation in vivo. RNA sequencing analysis implicated the enrichment of JAK-STAT signaling pathways. Meanwhile, we identified an additional type of RAD51C+ fibroblast that was significantly associated with the depth of tumor invasion. Conclusions The study establishes the Hub-CR.Sig as an immunotherapy response classifier and nominates RAD51C targeting as a promising therapeutic strategy for bladder cancer.

overall_design: Comparativegene expression profilinganalysis of RNA-seq data for 5637 cells and its KD derivatives (shRAD51C)


title: CarS2 mediated cysteine catabolism drives brown fat development and thermogenesis through persulfidating EBF2 [BAC and L1 differ]

first: Xuyun

last: Zhao

address: South Chongqing Road

summary: Targeting transcriptional regulatory complex is a promising strategy to activate thermogenic fat and treat  obesity. In this study, we focus on identifying critical and special metabolic pathways enriched during brown adipocyte differentiation. By analyzing metabolomics and RNA sequencing results from precursor and mature brown and white adipocytes, we revealed cysteine catabolism pathway is a unique metabolic process that is exclusively increased in the brown adipocyte differentiation. Then, we demonstrated cysteine persulfidation synthase CarS2 is directly induced by EBF2 and mediates cysteine catabolism to produce cysteine persulfide and triggers brown fat protein persulfidation in vitro and in vivo. Loss of CarS2 in thermogenic fat blocks brown and beige fat formation and reduced thermogenesis and energy expenditure in mice. More importantly, CarS2 generates cysteine persulfide and its derivative H2S cell autonomously induces brown adipogenesis and enhances uncoupled respiration by persulfidating EBF2. Mechanistically, the persulfidated EBF2 facilitates its interaction with PPARγ and enhanced the recruitment of the EBF2-PPARγ complex on browning gene promoter, thus driving brown fat development and boosting thermogenic function. Furthermore, the treatment of CarS2 coenzyme PLP or H2S donor elevates brown adipocyte function and ameliorates obesity progression in mice under HFD feeding, indicating a novel metabolite-based strategy for the obesity treatment.

overall_design:
To investigate the  gene expression profile of BAC cells before and after differentiation, we extracted the RNA of BAC at day 0 and day 4 during the differentiation.we performed gene expression profile using data obtained from RNA-seq before and after BAC differentiation with duplicate.
To investigate the  gene expression profile of 3T3-L1 cells before and after differentiation, we extracted the RNA of 3T3-L1 at day 0 and day 6 during the differentiation.we performed gene expression profile using data obtained from RNA-seq before and after 3T3-L1 differentiation with duplicate.

first: Hongyong

last: Song

first: Xin

last: Peng


title: Differences in microRNA expression in rat liver following single- and combined administration of caffeine and dextromethorphan

city: hirakata

first: Yuko

last: Emoto

institute: Kansai Medical University

type: Non-coding RNA profiling by high throughput sequencing

summary: Caffeine and dextromethorphan are among the most commonly abused substances by young people in Japan. However, reports on interactions between caffeine and dextromethorphan and comprehensive miRNA analyses following the administration of these drugs are limited. We aimed to determine whether miRNA expression differs in rats administered caffeine and dextromethorphan alone or in combination. We performed a comprehensive analysis of miRNAs in the livers of rats treated with caffeine, dextromethorphan, or both drugs.

overall_design: Six-week-old male Sprague-Dawley rats (Jcl:SD).After the acclimatization period, the 7-week-old rats were divided into four groups: control, caffeine, dextromethorphan, and caffeine plus dextromethorphan (n = 5 per group). The rats were orally administered the drugs for 5 consecutive days, as described below. The caffeine group (CAF) received 120 mg/kg caffeine, the dextromethorphan group (DEX) received 100 mg/kg dextromethorphan hydrobromide monohydrate, and the combination group (CAF + DEX) received 120 mg/kg caffeine and 100 mg/kg dextromethorphan hydrobromide monohydrate. All drugs were dissolved in sterile distilled water. Age-matched control groups (control) were treated with vehicle (sterile distilled water) only. The liver was collected from the abdominal vena cava 1 h after the final administration.


title: Transcriptional and chromatin accessibility profiling of melanoma cells during BRAF inhibitor–induced drug tolerance

city: Nashville

first: Blake

last: Baleami

state: TN

address: 1161 21st Ave S

department: Human Genetic PhD Program

institute: Vanderbilt University

summary: Drug tolerance can emerge rapidly in melanoma following treatment with BRAF inhibitors. This transition has been associated with transcriptional and chromatin state remodeling. To investigate the molecular features of this process, we profiled melanoma cells before and during exposure to a BRAF inhibitor. We generated single-cell RNA sequencing (scRNA-seq), bulk RNA sequencing (bulk RNA-seq), and bulk ATAC-seq datasets from SKMEL5 melanoma cells. Cells were analyzed under untreated conditions and following treatment with 8 µM BRAF inhibitor (PLX4720) for defined time periods corresponding to drug-tolerant states. Bulk RNA-seq was performed on multiple SKMEL5 subclones (SC01, SC07, SC10) to characterize transcriptional responses across biological replicates. scRNA-seq and ATAC-seq were used to profile transcriptional heterogeneity and chromatin accessibility changes associated with drug tolerance.

overall_design: SKMEL5 melanoma cells were cultured under standard conditions and treated with the BRAF inhibitor PLX4720 (8 µM) to induce drug tolerance. Samples were collected under untreated conditions and after defined treatment durations corresponding to drug-tolerant states. In this study, cells treated with PLX4720 for 8 days are referred to as idling cells. Single-cell RNA sequencing (scRNA-seq) and bulk ATAC-seq were performed on untreated cells and on idling cells treated with PLX4720 for 8 days. Bulk RNA sequencing was performed on three independent SKMEL5 subclones (SC01, SC07, SC10) under untreated conditions and following PLX4720 treatment for 3 days and 8 days, with three biological replicates per condition. These datasets were generated to characterize transcriptional and chromatin accessibility changes associated with the emergence of drug tolerance in melanoma cells.

first: Corey

last: Hayford

first: Philip

last: Stauffer

first: B

last: Bishal Paudel

first: Aziz

last: Al'Khafaji

first: Amy

last: Brock

first: Vito

last: Quaranta

first: Daren

middle: R

last: Tyson

first: Leonard

last: Harris


title: Phase Separation of DUX Family Proteins Drives Totipotent-like State via 3D Genome Reorganization and Retrotransposon Activation

first: Qifeng

last: Gao

first: Leilei

last: Hai

first: Ziqiang

last: Wu

first: Penghui

last: Li

first: Han

last: Kang

first: Xiaohui

first: Jinlian

last: Hua

first: Shiqiang

last: Zhang

first: Gang

last: Ren

first: Jihong

last: Yang

first: Leqian

last: Yu

first: Yulei

last: Wei

first: Junjun

last: Ding

first: Fan


title: Genetically modifying the RNA-binding protein Regnase-1 reveals its critical role in Treg homeostasis and functions in vivo

city: Houston

first: Ge

last: Deng

state: Texas

address: 6670 Bertner Avenue, R7-217

institute: Houston Methodist Hospital and Houston Methodist Research Institute

summary: Regnase-1 is a CCCH-type RNA-binding protein that degrades mRNA species via its ribonuclease activities to inhibit immune responses. But how and by what mechanisms Regnase-1 controls Foxp3+ Tregs in vivo remains poorly studied. Here we generated a conditional knock-in mouse in which a degradation-resistant Regnase-1 mutant (R111A) was selectively expressed in Foxp3+ Tregs as a transgene (Foxp3YFP-CreLSL-R111Afl/fl) to investigate the roles of Regnase-1 in Tregs. We found that all male Foxp3YFP-CreLSL-R111Afl/fl mice died of fatal autoimmune diseases by 4 weeks of age, which was accompanied by massive activation of T effector cells in vivo. We also found that the Foxp3+ Tregs in male Foxp3YFP-CreLSL-R111Afl/fl mice were depleted in the periphery, but not Tregs in the thymus. Further analyses revealed that the Foxp3+ Tregs expressing the Regnase-1 mutant R111A displayed an intrinsic survival defect and that transferring wild type Foxp3+ Tregs into the young male Foxp3YFP-CreLSL-R111Afl/fl mice could rescue the lethal autoimmune phenotype. Mechanistically, we demonstrated that the Regnase-1 readily degraded Bcl2l1 mRNA (encoding Bcl-xL) via binding to its stem loop structure, thus preventing Bcl-xL expression in Tregs. Consequently, Tregs expressing the Regnase-1 mutant R111A exhibited not only survival defects but also failure to respond to IL-2 in vivo, thus leading to their eventual demise. Together, our studies suggest that Regnase-1 can have a profound impact on Foxp3+ Tregs and that modulating Regnase-1 in Tregs may have important therapeutic implications.

overall_design: RNA profiling of FACS-sorted WT and R111A CD4+ T cells  with anti-CD3 plus CD28 (5 μg/mL+1 μg/mL) activation for 12 hours.

first: Xiaojun

last: Su

first: Xiang

last: Xiao

first: Si

last: Sun

first: Mou

last: Wen

first: Ashton

last: Connor

first: Zhuyun

last: Mao

first: Rafik

middle: M

last: Ghobrial

first: Wenhao

last: Chen

first: Xian


title: Landscape of mitochondrial RNA binding proteome

first: Jinshuai

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus


title: Genetically modifying the RNA-binding protein Regnase-1 reveals its critical role in Treg homeostasis and functions in vivo [HyperTRIBE]

summary: In HyperTRIBE analysis, a hyperactive mutant of the catalytic domain of ADAR enzyme is fused to an RNA-binding protein of interest in order to identify RNAs that are bound by the protein interest through RNA-sequencing (Rahman et al Nat Protoc. 2018). By discovery of RNAs where adenosines have been edited to iosines, the RNAs that were bound by the protein of interest and then edited by ADAR are identified genome-wide. We conducted this analyis for Regnase-1, an RNA binding protein with multiple described cellular functions, including mRNA degradation and T cell activation. Because Regnase-1 can degrade the mRNA quickly, we used the nuclease-dead mutant Reg1-D141N to conduct this assay in activated CD4+ T cells from wild-type mice.

overall_design: CD4+ T cells overexpressing Reg1-D141N-ADAR, Ctrl-ADAR or control empty vector for 24 hours were FACS-sorted for RNA-seq to look for A-G(I) editing events

last: Li


title: Optineurin modulates mTORC2/AKT/STAT3 signaling to control MHC II expression and adaptive immunity against HSV-1

city: Chicago

first: Leonid

last: Feferman

state: Illinois

address: 900 S. Ashland Ave

department: Research Resources Center

institute: University of Illinois at Chicago

summary: Herpes simplex virus 1 (HSV-1) infection contributes to immunopathogenic diseases and lacks an effective vaccine. Improving antigen presentation is key to better vaccine strategies and more robust immune responses. Here, we show that OPTN, an autophagy receptor traditionally involved in protein recycling, unexpectedly stabilizes RICTOR (mTORC2), a crucial step in enhancing MHC-II surface expression in dendritic cells. OPTN regulates the AKT/mTOR/STAT3 pathway, with AKT2 isoform playing a central role. Using scRNA-seq and transgenic mouse models, we identify the mechanistic details of this pathway. Dysregulation impairs antigen presentation, weakening immunity and vaccine efficacy. Our findings uncover a novel function for OPTN and highlight its role in coordinating innate and adaptive immune defenses, with implications for vaccine development and immune response modulation in HSV-1 and other viral and bacterial diseases.

overall_design: HSV-1 infection was performed by corneal scarification method.Their corneas were scarified in a 3 x 3 grid using a 30-gauge needle followed by the application of HSV-1 McKrae at  5 × 105  PFU.    At 8dpi animals were sacrificed and cLN were harvested from 4 animals in each group and pooled into one sample. cLNs were manually harvested using 5 ml syringe and isolated immune cells were further subjected to DC isolation using Miltenyi kit. Isolated DCs were mixed in a specific ratio with lymphocytes from the same tissue to enrich the DC population.

first: Rashmi

last: Kadam

first: Deepak

last: Shukla

first: Mark

last: Maienschein-Cline

first: George

last: Chlipala


title: Sox8 and Sox9 influence differentiation and nuclear positioning of retinal Muller glia.

city: Pittsburgh

first: Clayton

middle: Pio

last: Santiago

state: Pennsylvania

address: 1622 Locust St.

department: Ophthalmology

institute: University of Pittsburgh

summary: Temporal patterning of retinal progenitor cells governs the sequential generation of retinal cell types, with gliogenesis occurring late in development. Sox8 and Sox9, members of the SoxE transcription factor family, are highly expressed in late-stage retinal progenitor cells and mature Müller glia (MG), yet their functional roles remain incompletely defined. Here we employed gain- and loss-of-function approaches, single-cell multiomic profiling, and injury models to investigate Sox8/9 function. Overexpression of Sox8 and/or Sox9 in early-stage RPCs suppressed early-born cell fates and promoted photoreceptor generation, consistent with a role in late-stage temporal identity. Conversely, conditional deletion of Sox8 and/or Sox9 in late-stage progenitors did not impair MG specification, but caused radial displacement of MG nuclei into the outer retina and modest changes in glial gene expression. Loss of Sox8/9 in mature MG modestly increased proliferation post-injury without inducing neurogenic competence. These findings suggest that Sox8/9 are dispensable for gliogenesis and repression of neurogenic competence, but are essential for proper laminar positioning and maturation of retinal Müller glia.

overall_design: Single cell RNA-seq or Single cell ATAC-seq was performed on control or Sox8/9 knockout retinal cells using 10x Genomics Chromium Single Cell 3’ Reagents Kit v3.1, Single Cell 3’ Reagents Kit v4 or Multiome Reagent Kit

first: Nicole

last: Pannullo

first: Debarpita

last: Datta

first: Jared

last: Tangeman

first: Lizhi

last: Jiang

first: Leighton

middle: H

last: Duncan

first: Seth

last: Blackshaw


title: Palmitoleic acid promoted by BMPR2 signaling primes CD169 macrophages and alleviates liver fibrosis [PM_POA_PA]

city: Shanghai

first: Jinkun

last: Zhong

address: No. 130, Dong'an Road, Xuhui District, Shanghai

institute: Fudan University

summary: Bone morphogenetic proteins (BMPs) participate in the energy metabolism. BMP receptor type 2 (BMPR2) is expressed in the liver. Whether BMPR2 is involved in the pathophysiology of nonalcoholic fatty liver disease (NAFLD) has not been studied. In this study, we analyzed the RNA-sequence data of several patient cohorts and found that BMPR2 expression decreases at the stages of advanced fibrosis. A mouse model featuring BMPR2 knockout demonstrated that BMPR2 knockout in the liver produced profibrotic effect upon long-term high-fat and high-fructose (HFF) diet challenge. BMP signaling promoted biosynthesis of unsaturated fatty acids, which was repressed under BMPR2 knockout condition. The most affected unsaturated fatty acid, palmitoleic acid (POA), was found to activate CD169 macrophages. Moreover, CD169 macrophages were revealed to have high levels of lysosomal enzyme along with matrix metalloproteinase 8, which degraded collagen and alleviated fibrosis. Our study demonstrated that BMPR2 influences over the progression of liver fibrosis induced by the HFF diet through fatty acid-regulated CD169 macrophages. POA-primed macrophages might be a potential therapeutic strategy for the treatment of liver fibrosis.

overall_design: RNA-seq profling of wildtype peritoneal macrophages treated with palmitoleic acid(POA) and palmitic acid(PA).

first: Hong

last: Ma

first: Qilin

last: Peng

first: Mengyuan

last: Dai

first: Linyuan

last: Wang

first: Jianan

first: Danchun

last: Huang

first: Yang

last: Liu

first: Yan

last: Tang

first: Wei

last: Xu

first: Qi-qun

first: Shuwen

last: Qian


title: snRNA-Seq analysis of islets from WT mice treated with or without 0.5mM Glucosamine

city: Jerusalem

first: Yael

last: Riahi

address: Kyriat Hadassah

country: Israel

institute: The Hebrew University

summary: In diabetes, hyperglycemia drives the progression of b-cell failure and multi-organ complications. Prolonged exposure to high glucose stimulates mTORC1, resulting in impaired glucose metabolism and exacerbation of ER stress and inflammation. However, the precise mechanism by which glucose regulates mTORC1 activity in diabetes remains unclear. We performed metabolic labeling with 13C6- glucose in diabetic animals treated with or without the SGLT2 inhibitor (SGLT2i) dapagliflozin or insulin, followed by targeted metabolomics and metabolic flux analysis. We found that tissue glucose concentrations strongly correlate with glucosamine, rather than with other glucose or amino acid metabolites. Plasma glucosamine levels are increased in patients with impaired fasting glucose (IFG) and type 2 diabetes (T2D) and are inversely correlated with b-cell function. Glucosamine is synthesized by amine group transfer from glutamine to glucose, which enhances O-GlcNAcylation (O-GlcN) of mTORC1- regulating proteins, including Raptor, via O-GlcNAc transferase (OGT), subsequently activating mTORC1. We propose that the glucosamine-mTORC1 pathway is a key mediator of glucose toxicity in diabetes.  We studied the role of glucosamine in mediating beta-cell glucotoxicity.

overall_design:
To investigate the molecular mechanisms underlying glucotoxicity in beta cells, we performed snRNA-seq analysis. Pancreatic islets were isolated from three WT mice, using a split-sample design: islets from each mouse were divided equally into two groups, receiving either 0.5 mM Glucosamine or vehicle control for 24 hours.
This yield three paired biological replicates per condition.
Following treatment, islets were dissociated into single-cell suspension and nuclei were subsequently isolated.

first: Erez

last: Dror

first: Ziv

last: Teselpapa

first: Gil

last: Leibowitz


title: Landscape of mitochondrial RNA binding proteome [Par_clip]

first: Jinshuai

last: Sun

overall_design: RNA signals of the mitochondria purified from HeLa cells with 4sU labelling (PAR-CLIP). Only the Reverse reads were used in the data analysis and uploaded. Bowtie was applied to map sequencing reads against GRCh38 genome with up to two mismatches allowed.

first: Yingying

last: Lin

first: Xiwei

last: Wangh

first: Xichen

last: Bao


title: Integrating epigenome-wide Mendelian randomization and multi-omics data validation to identify epigenetic drivers of idiopathic pulmonary fibrosis

first: Xingyu

last: Lv

address: No. 107, West Jiangxi Road

department: Sun Yat-Sen Memorial Hospital

institute: Sun Yat-sen University

summary: Idiopathic pulmonary fibrosis (IPF) is a complex disease without clear etiology or effective therapy. While DNA methylation has been implicated in IPF pathogenesis, the tissue-specific causal effects of the epigenetic factors on IPF remain undetermined. Here, we performed epigenome-wide Mendelian randomization using blood-based methylation quantitative trait loci of 420,509 CpG sites and genome-wide association study for IPF to elucidate the causal effects of the CpG sites on IPF. Totally, 452 CpG sites has shown putative causal effects on IPF risk after Bonferroni correction. Among them, 13 CpG sites have shown strong colocalization evidence with genetic factors associated with IPF. Specifically, DNA methylation at CpG sites within MAN2A2 and TRIM27 shows significant differences between IPF lungs and controls, correlating with altered mRNA expressions of these genes in lung tissues. The CpG site in MAN2A2 is a binding site of ZNF384 according to transcription factor databases. RNA sequencing in the TGFβ1-induced alveolar epithelia confirmed significantly reduced expression of MAN2A2 and ZNF384 comparing to the controls. Collectively, our study suggests a putative causal link between DNA methylation within MAN2A2 and IPF risk, wherein lung-specific DNA methylation in MAN2A2 may perturb the interaction between ZNF384 and MAN2A2, revealing novel roles for these genes in IPF pathogenesis.

overall_design: RNA samples derived from TGFβ1-induced A549 cells and control cells were sequenced on the NovaSeq6000 platform (Illumina; USA). Details of RNA sequencing are described in the Supplementary Methods. RNA expression levels were quantified by fragments per kilobase million (FPKM). We compared the mRNA expression levels of target genes and TFs using unpaired two-tailed Student’s t-tests.

first: Zhaoqi

last: Li

first: Luna

last: Hong

first: Shuangyu

last: Yang

first: Fengzhan

last: Ye

first: Linjie

first: Shanping

last: Jiang

first: Huiying

last: Zhao


title: Distinct Transcriptomes for Cardiac Repair following multidose intramyocardial administration of Umbilical Cord Blood Mononuclear cells(UCB-MNCs)

city: Rochester

first: Nkechi

last: Odogwu

state: MN

address: 200 First Street SW

department: Cardiovascular Medicine

institute: Mayo Clinic

summary: Cardiac regeneration remains a major therapeutic challenge, and cell-based strategies offer a promising avenue. In this study, we investigated the transcriptional responses to single versus multidose intramyocardial administration of umbilical cord blood–derived mononuclear cells (UCB-MNCs) in a porcine model of right ventricular pressure overload. Transcriptome profiling revealed distinct gene expression patterns associated with repeated dosing, including activation of pathways related to cardiomyocyte proliferation, angiogenesis, and extracellular matrix remodeling. Taken together, these data provide a molecular framework for understanding how dosing strategy modulates gene expression programs during cardiac repair and identify novel targets for enhancing the efficacy of regenerative cell therapies.

overall_design: Porcine right ventricular myocardium samples were collected from a preclinical model of right ventricular pressure overload to study the transcriptional effects of umbilical cord blood–derived mononuclear cell (UCB-MNC) therapy. Animals were randomized into three treatment groups: multidose UCB-MNC, single-dose UCB-MNC, and vehicle control. Tissue was harvested at defined post-treatment timepoints, flash-frozen, and stored at −80 °C until RNA extraction. Total RNA was isolated from homogenized myocardial tissue using standardized protocols, and only samples with RNA integrity number (RIN) ≥ 7 were included. RNA libraries were prepared and sequenced on an Illumina platform, targeting sufficient depth per sample to enable differential expression analysis. Biological replicates correspond to individual animals per group and timepoint; no technical replicates were sequenced. The primary variables under investigation are dosing strategy (multidose vs. single dose vs. control) and secondary covariates such as sex, weight, and batch recorded and accounted for in analysis. This design allows direct comparison of how dosing regimens influence gene expression programs associated with cardiac repair.

first: Timothy

last: Nelson


title: Chromatin occupancy by Tox3 in mouse cerebellum

city: SAN FRANCISCO

first: Xiaoxin

last: Chen

address: 555 mission bay blvd. south, RM382A

department: Cardiovascular Research Institute

institute: University of California, San Francisco

summary: Organ development relies on precise transcriptional control, yet how lineage-defining factors like Atoh1/Math1 drive robust gene expression despite weak intrinsic transactivation activity remains unclear. Here, we present a comprehensive atlas profiling 1,904 transcription regulators across organs, uncovering TOX3 as a potential co-activator of Atoh1 in cerebellar granule neuron progenitors. Tox3-deficient mice display severe ataxia and cerebellar hypoplasia, driven by depletion of granule neuron progenitors, diminished Atoh1 expression, and impaired primary cilia. Single-nucleus RNA-seq analyses reveals compromised maintenance of the progenitor pool. TOX3 is also highly expressed in subsets of medulloblastoma, and its deletion reduces cerebellar neoplasia and prolongs survival in a mouse model. Mechanistically, Tox3 and Atoh1 co-occupy and synergistically activate E-boxes in shared target genes by up to 120-fold, including an ultra-conserved E-box downstream of Atoh1. These findings establish Tox3 as a critical Atoh1 co-activator in cerebellar development, tumorigenesis and evolution, and provides a resource for exploring novel transcriptional regulators in progenitor maintenance and organogenesis.

overall_design: ChIP-seq of TOX3 in control and Tox3 neuronal conditional knockout mice cerebella at postnatal day 1.

first: Chen

last: Xiaoxin

first: Huang

middle: N

last: Guo


title: Phase Separation of DUX Family Proteins Drives Totipotent-like State via 3D Genome Reorganization and Retrotransposon Activation [ATAC-Seq]

first: Qifeng

last: Gao

overall_design: To investigate the transcriptomic consequences of DUX-mediated phase separation and its necessity for the induction of a totipotent-like state, we performed RNA-seq profiling across various experimental conditions. In this study, RNA-seq was used as a primary readout to evaluate how the global gene expression program, particularly the activation of 2C-specific genes and retrotransposons like MERVL, was altered following the overexpression of wild-type DUX. To evaluate the role of Dux across different totipotency induction systems, we performed RNA-seq on Dux-knockdown (KD) cells subjected to RA-induced, PlaB-induced, or Zscan4-induced totipotency programs. This component of the study was designed to assess whether these alternative pathways for inducing a totipotent-like state could bypass the loss of Dux. Furthermore, to evaluate the functional requirement of DUX-induced super-enhancers (SEs) in gene activation, we conducted RNA-seq on Tdpoz-SE knockout (KO) cells during DUX-mediated induction. By comparing these results with Dux-induced control samples, we sought to test whether SEs are indispensable for the activation of key totipotency genes. Finally, to define the specific contribution of liquid-liquid phase separation (LLPS), we performed RNA-seq on cells expressing the phase-separation-deficient mutant (DUXR70A). This design allowed us to measure whether the loss of DUX nuclear condensates would abolish the activation of the totipotent transcriptome, even when the mutant’s DNA-binding capacity remained intact. Collectively, we utilized these datasets to assess how DUX coordinates distal gene expression through SE-mediated architectural reorganization.

first: Leilei

last: Hai

first: Ziqiang

last: Wu

first: Penghui

first: Han

last: Kang

first: Xiaohui

last: Song

first: Jinlian

last: Hua

first: Shiqiang

last: Zhang

first: Gang

last: Ren

first: Jihong

first: Leqian

last: Yu

first: Yulei

last: Wei

first: Junjun

last: Ding

first: Fan


title: RNAseq analysis of Tox3 dependent gene expression in the mouse cerebellum

first: Xiaoxin

overall_design: RNA-seq profiling of control and Tox3 neuronal conditional knockout mice cerebellum at day1 after birth.


title: Single-nucleus RNA-seq analysis of Tox3 dependent gene expression in the mouse cerebellum

overall_design: single-nucleus RNA-seq profiling of control and Tox3 neuronal conditional knockout mice cerebella at postnatal day 1.

first: Chen


title: m6Am methyltransferase PCIF1 regulates osteogenesis via WNT signaling

city: New Orleans

first: Hong-Wen

last: Deng

state: LA

address: 1440 Canal St, suite 1619G

institute: Tulane University

summary: Skeletal development depends on the self-renewal and differentiation of MSCs. Emerging research underscores the regulatory functions of RNA methylation on bone homeostasis. Here, we show PCIF1, the N6,2′-O-dimethyladenosine (m6Am) methyltransferase, is essential for maintaining bone mass and promoting osteogenic differentiation of MSCs. Multiple complementary analyses—including GWAS, TWAS, and single-cell transcriptomics—collectively point to PCIF1 as a regulator of human bone mineral traits and a marker of early mesenchymal stem cells. Global or MSC-specific Pcif1 deletion elicits osteoporotic pathology in mice, although myeloid cell-specific Pcif1 knockout does not induce femur bone alternation alterations. Mechanistically, Pcif1 knockout decreases m6Am signals of Wnt-related genes (Wnt11, Fzd4, Fgfr2) and accelerates mRNA degradation. This down-regulates active β-Catenin protein, and thus impairs osteogenic function of MSCs. Additionally, the WNT agonist attenuates the osteoporosis-like phenotype induced by Pcif1 deletion. These findings highlight the crucial role of PCIF1-mediated m6Am modification in regulating osteogenesis and suggest potential therapeutic implications for bone disorders.

overall_design: Human femoral head specimens were obtained from patients undergoing total hip arthroplasty. Bone tissues were enzymatically digested to generate single-cell suspensions. Hematopoietic and immune cells were depleted using magnetic-activated cell sorting to enrich osteolineage cell populations. Single-cell RNA sequencing libraries were prepared using the 10x Genomics Chromium Single Cell 3′ platform. Libraries were sequenced on an Illumina NovaSeq 6000 instrument using paired-end sequencing.

first: Wei

first: Kuan-Jui

last: Su

first: Zhehui

last: Pan

first: Zhong

first: Qi

last: Yin

first: Weimin

last: Lin

first: Linfeng

last: Liu

first: Yun

last: Gong

first: Bocheng

last: Liang

first: Yimeng

last: Cai

first: Qiwen

last: Li

first: Hui

last: Shen

first: Quan

last: Yuan


title: Polycomb Repressive-Deubiquitinase Complex Safeguards Oocyte Epigenome and Female Fertility by Restraining Polycomb Activity

city: Davis

first: Satoshi

last: Namekawa

address: One Shields Avenue

institute: UC Davis

summary: Mouse oocytes exhibit a unique chromatin landscape characterized by broad H3K27ac and H3K27me3 domains, demarcating euchromatin and facultative heterochromatin, respectively. However, the mechanisms underlying this non-canonical landscape remain elusive. Here we report BAP1, a core component of the Polycomb Repressive-Deubiquitinase (PR-DUB) complex, as a key negative regulator of Polycomb activity during oogenesis. BAP1 restricts pervasive H2AK119ub1 accumulation in oocytes and protects oocyte-specific broad H3K27ac, particularly within gene-poor regions, from ectopic H3K27me3 deposition. While PR-DUB has been linked to gene repression, in oocytes BAP1 primarily promotes transcription and contributes minimally to Polycomb-mediated silencing. BAP1-dependent transcriptional activation during oogenesis is essential for oocyte developmental competence, maternal-to-zygotic transition, and female fertility. Notably, the aberrant H3K27me3 landscapes established in BAP1-deficient oocytes, but not H3K27ac alterations, persist in preimplantation embryos. The ectopic H3K27me3 domains nonetheless are resolved by early post-implantation stages, and loss of maternal BAP1 does not impair either canonical or non-canonical genomic imprinting. Together, these findings reveal a critical role for PR-DUB in safeguarding the oocyte epigenome by protecting euchromatin from ectopic Polycomb activity, rather than enforcing transcriptional repression.

overall_design: Chromatin profiling analysis of quantitative CUT&Tag data for WT P1 oocytes.

first: Mengwen

last: Hu


title: Closed form of ezrin binds RNA to maintain a metastatic phenotype in osteosarcoma

city: Washington

first: Aykut

last: Uren

state: DC

address: 3970 Reservoir Road, N.W.

department: Department of Oncology

institute: Georgetown University

summary: Ezrin is a cytoplasmic protein that can exist in multiple conformational forms that are regulated by phosphorylation at the threonine 567 residue. Phosphorylated open form of ezrin was generally accepted as the active form because it can translocate to the plasma membrane whereas unphosphorylated closed ezrin, sequestered in the cytoplasm, have been considered inactive. Here, we identified closed ezrin as a novel RNA binding protein with biological activity. Our previous work established that ezrin directly interacts with cytoplasmic RNA binding proteins (RBPs). Here we show that ezrin expression correlates with RBPs in human osteosarcoma samples. Recombinant ezrin in its closed conformation directly bound RNA with 6.1 nM affinity. Ezrin binding specificity was highest for the guanine-rich sequences and RNA G-quadruplexes (G4 RNA).  Reintroduction of closed ezrin expression rescued the transcriptome and proteome profiles in an ezrin-null osteosarcoma cell line. Closed ezrin also bound endogenous mRNAs that were enriched for pathways related to RNA processing and splicing, DNA maintenance, and cellular metabolism. Closed ezrin also rescued the metastatic capability of osteosarcoma xenografts in zebrafish. Taken together, our data established that the closed conformation of ezrin, previously thought to be inactive, can directly bind RNA and contribute to the metastatic phenotype.

overall_design: We first knocked out the endogenous ezrin expression in K7M2 mouse osteosarcoma cell line by using CRISPR/Cas9 gene editing at the EZRIN gene locus. We then stably rescued the ezrin expression by stably transfecting them with a T567A mutant EZRIN cDNA carrying vector. The resulting cell lines were referred here as COL3A.

first: Emre

last: Deniz


title: Single-Cell RNA Sequencing of PBMCs from Clinically Stable Connective Tissue Disease-associated Interstitial Lung Disease (CTD-ILD) Patients after Lung Transplantation

city: Beijing

first: Qingya

last: Shi

address: 30 Shuangqing Road, Haidian District

department: Tsinghua Medicine

institute: Tsinghua University

summary: Connective tissue disease–associated interstitial lung disease (CTD-ILD) is a major indication for lung transplantation in patients with advanced disease. Here, we performed single-cell RNA sequencing (scRNA-seq) of peripheral blood mononuclear cells (PBMCs) from two clinically stable CTD-ILD patients after lung transplantation to characterize circulating immune cell populations and their transcriptional profiles. This dataset provides a single-cell transcriptomic resource for investigating immune heterogeneity and systemic immune status in CTD-ILD patients following lung transplantation.

overall_design: Expression profiling by high throughput sequencing

first: Shiyao

last: Wang

first: Huaping

last: Dai

first: Wenhui

last: Chen


title: Transcriptome sequencing analysis of culm brittle phenotypes in rice  TAC4/TAC4L double knockout mutants

city: Yangzhou

first: Chongyuan

last: Zhong

address: Hanjiang

institute: Yangzhou University

summary: In rice, simultaneous knockout of TAC4L (LOC_Os04g19140) and TAC4 (LOC_Os02g25230) resulted in a brittle culm and leaf phenotype, whereas single knockout of either TAC4L or TAC4 did not produce such a brittle phenotype. To elucidate the regulatory pathway underlying the brittle phenotype mediated by TAC4L and TAC4 in rice, we performed transcriptome sequencing using the penultimate culm at the heading stage.

overall_design: RNA-seq profiling of culms in wild-type japonica rice Zhonghua 11 (ZH11) and its knockout lines (TAC4-KO, TAC4L-KO, and TAC4L/TAC4-KO) at the heading stage.

first: Shengyuan

last: Sun

first: Changjie

last: Yan


title: CapMux: a Snakemake pipeline for early demultiplexing of split-pool scRNA-seq data into sample-resolved outputs

city: Vilnius

first: Denis

last: Baronas

address: Saulėtekio al. 7

country: Lithuania

institute: Vilnius University

summary: Single-cell RNA sequencing methods based on split-pool combinatorial barcoding enable high-throughput profiling, yet sample identity is often encoded during early barcoding steps rather than through the library index. Consequently, reads from multiple biological samples remain pooled, complicating per-sample analysis and selective extraction of samples of interest. Here, I present CapMux, a Snakemake-based pipeline for processing split-pool scRNA-seq data from raw sequencing files to sample-resolved outputs. CapMux supports workflows starting from either BCL files or FASTQ files and reconstructs sample identity by integrating sub-library index information with the experiment-specific barcoding plate layout. The pipeline was developed for the CapSeq method but is configurable for related scRNA-seq combinatorial barcoding designs through specification of barcode positions and experimental layout. CapMux resolves pooled data into outputs for each sample, enabling independent quality control summaries, mapping statistics, count matrices, and downstream visualizations. Runtime benchmarking indicated that secondary demultiplexing step added only a modest computational overhead. Together, these results show that CapMux provides a practical and adaptable framework for recovering sample-level resolution from split-pool scRNA-seq data.

overall_design: CapSeq scRNA-seq experiment performed on Neuro-2a, NIH/3T3, and mESC cell lines. Each cell line was distributed across four BC1 plate columns: Neuro-2a cells in columns 1–4, NIH/3T3 cells in columns 5–8, and mESCs in columns 9–12.


title: Decoding a combinatorial RNA polymerase II CTD grammar that specifies protein-coding gene transcription and mRNA export [TT-seq]

city: Austin

first: Qian

last: Zhang

state: TX

address: 2515 Speedway

institute: The Unibersity of Texas at Austin

summary: The C-terminal domain (CTD) of RNA polymerase II orchestrates transcription via specific post-translational modifications (PTMs), notably phosphorylation. The combinatorial phosphorylation of CTD can potentially encode the precise recruitment of transcription machinery but lacks robust evidence. Addressing the challenge of detecting combinatorial CTD phosphorylation in cells without specific antibodies, we developed a novel genome-wide approach exploiting antibody masking and selective interference removal to identify dual phosphorylation patterns. Using this method, we uncovered pT4pS5 phosphosites occurring exclusively during protein-coding gene transcription but excluded in non-coding genes. Using multiple sequencing methods, the role of T4 phosphorylation in elongation is highlighted. Particularly, pT4pS5 marks recruit the Transcription and Export complex to facilitate mRNA export. These findings provide robust evidence that CTD operates as a combinatorial, gene-specific coding system, encoding precise transcriptional instructions. Our approach offers a versatile framework for mapping all combinatorial PTMs on RNA polymerase II, advancing molecular insights into gene-specific transcriptional regulation.

overall_design: RPB1 degron HEK293 cells were transiently transfected with RPB1 constructs containing either the 52× wild-type CTD sequence or a T4A mutation in every CTD repeat. 48 hours after transfection, auxin was added for an additional 48 hours to induce degradation of endogenous RPB1. Nascent RNA was then labeled with 4-thiouridine (4-SU) for 20 min and collected for TT-seq analysis.

first: Svetlana

middle: B

last: Panina

first: Kangsan

last: Kim

first: Jonathan

last: Smailys

first: Haley

last: Hardtke

first: Seema

last: Irani

first: Brendan

last: Floyd

first: Edward

last: Marcotte

first: Yan


title: Epigenetic evolution of IDHwt glioblastomas

city: Rotterdam

first: Bo

last: Deng

address: Dr. Molewaterplein 40

country: Netherlands

institute: Erasmus MC

summary: Although the genetic evolution of IDH-wildtype glioblastomas has extensively been investigated, limited studies have addressed the epigenetic evolution. Understanding the epigenetic evolution is particularly relevant as demethylation of the MGMT promoter may form a means to treatment resistance.  Methods  We generated whole genome DNA methylation data of 64 matched primary-recurrent samples from IDH-wildtype glioblastoma patients. Data were combined with three publicly available datasets into a final cohort consisting of 418 samples. In the combined dataset, we determined change in genome-wide DNA-methylation. MGMT promoter methylation was defined using the MGMT-STP27 algorithm. CoxPH regression was used to investigate the impact of identified changes on survival.   Results  Our analysis demonstrates that the methylome of IDH-wildtype glioblastomas is highly stable. Changes that do occur can mostly be allocated to differences in tumor purity. Conversion from a MGMT promoter methylated to unmethylated status at progression occurred infrequently, but significantly more often than the converse. Conversion was associated with poorer overall and progression-free survival compared to those that remained MGMT methylated. Similar to previously reported, MGMT promoter methylation status was significantly associated with overall-, progression-free and post-recurrence survival. Despite this large survival difference, very few CpGs were co-methylated with the MGMT methylation status. Of these, the vast majority lied within the MGMT gene body and were inversely correlated with MGMT promoter methylation status.    Conclusions   The methylome of IDH-wildtype glioblastomas is highly stable at tumor progression. This stability is also present in the MGMT promoter.

overall_design: Genome-wide DNA methylation data was generated from 64 samples of patients included in the GSAM (Glioblastoma-Stability of Actionable Mutations) study.DNA methylation profiling for all GSAM samples was performed using the Illumina EPIC (850K) methylation array. For this, tumor DNA was extracted from FFPE tissue using the QIAamp DNA FFPE Tissue Kit (Qiagen), and the quantity of isolated DNA was assessed with a Qubit 3.0 fluorometer (Invitrogen). Genome-wide DNA methylation profiling was done using the Illumina HumanMethylationEPIC (850K) beadchip.

first: Pim

last: French


title: CD5L Promotes Efferocytosis and Resolution of Retinal Ischemic Injury

city: Little Rock

first: Rami

middle: Ahmad Nawaf

last: Shahror

state: AR

address: APT 277, 11800 Pleasent Ridge Rd.

institute: University of Arkansas for Medical Sciences

summary: Ischemia-induced retinopathy is a defining characteristic of prevalent ocular conditions such as diabetic retinopathy and central retinal artery and vein occlusions. Current therapeutic interventions for ischemic retinopathies have shown limited efficacy, highlighting the need for a better understanding of the underlying mechanisms. Histone deacetylase 3 (HDAC3) is a member of the class I histone deacetylase enzyme family, which participates in regulating gene expression by removing acetyl groups from histone proteins. We have recently uncovered the role of myeloid HDAC3 as a potential key inhibitor of efferocytosis following retinal ischemia, the process by which myeloid cells clear dead cells to facilitate tissue repair. Here, we investigated the underlying mechanism by which HDAC3 inhibits efferocytosis through regulating the CD5 antigen-like (CD5L)/cluster of differentiation 36 (CD36) axis, which was identified through RNA-sequencing. We utilized myeloid-specific HDAC3 knockout (KO) mice in our in vivo studies and in vitro macrophage efferocytosis assays. To assess the role of CD5L in retinal injury, we subjected CD5L KO mice to ischemia-reperfusion (IR) and treated wild-type (WT) mice with recombinant CD5L. We evaluated retinal injury outcomes through retinal imaging, neurovascular assessments, and functional tests conducted in vivo. Additionally, we conducted in vitro studies to investigate the underlying mechanisms. RNA sequencing data identified CD5L as a pro-efferocytic molecule that is upregulated in HDAC3 knockout macrophages after exposure to apoptotic cells in vitro. Similarly, CD5L is also upregulated in the retinas of myeloid HDAC3 knockout mice following IR injury, where it colocalized with myeloid cells. HDAC3 suppresses CD5L transcription in an LXR-dependent manner. Experiments conducted with CD5L KO mice and treatment involving recombinant CD5L protein demonstrated a protective role for CD5L against retinal ischemic injury. Additionally, we found that CD36, a receptor for CD5L that facilitates the clearance of apoptotic cells, is upregulated in the retina myeloid cells following IR injury. In vitro experiments indicated that CD5L mediates efferocytosis via CD36.

overall_design: Gene expression profiling analysis of RNA-seq data from HDAC3 knockout and control macrophages after exposure to apoptotic cells, and non apoptotic cells, or no treatment. The expermintes were done in  biological triplcates.

first: Bushra

last: Zaman

first: Piyanan

last: Chuesiang

first: Melissa

last: Wild

first: Esraa

last: Shosha

first: Yuet-Kin

last: Leung

first: Shengyu

last: Mu

first: Nancy

last: Rusch

first: Abdelrahman

middle: Y

last: Fouda


title: Pan-cancer single-cell atlas of mouse and human tumor-associated dendritic cells

city: Brussels

first: Daliya

last: Kancheva

address: Laarbeeklaan 103, Building E

country: Belgium

institute: Vrije Universiteit Brussel - VIB

summary: Dendritic cells (DCs) are critical inducers of anti-tumor immunity. Yet, a comprehensive mapping of mouse and human DC subsets and states in a cancer context is lacking. Aiming to address this knowledge gap, we have generated pan-cancer mouse and human tumor-associated DC (TADC) scRNA-seq atlases, encompassing 15 mouse tumor models and 10 human cancer types, within which we identified several lineage-defined DC subsets along with functional states. We show that TADCs acquire an inflammatory profile with tumor progression and that tumor-mediated reprogramming occurs within the DCs from lymph nodes of tumor-bearing mice. Importantly, we demonstrate that TADCs are conserved across mice and human, and that gene signatures of different TADC subsets/states correlate with patient outcomes. Overall, we provide an in-depth characterization of the TADC compartment in mouse and human cancers, which can improve our understanding of the tumor microenvironment and contribute to the development of new anti-cancer therapies.

overall_design: E0771 scRNAseq kinetic dataset: CD45+7-AAD- MHC-IIlow-hiCD11c-/+ cells from tumors of orthotopic E0771 breast cancer mouse model, 9, 14 or 21 days post tumor inoculation. B16-F10 scRNAseq dataset: CD45+7-AAD- cells from tumors of B16-F10 melanoma mouse model. LLC-OVA dataset: CD45+7-AAD- cells (tumors and tumor-draining lymph nodes, scRNAseq) or unsorted splenic cells (scRNA/BCR-seq) of a subcutaneous LLC-OVA lung cancer mouse model. KP kinetic CITEseq dataset: CD45+7-AAD- cells from tumors of a spontaneous KrasLSL-G12D/+; Trp53fl/fl  (KP) lung cancer mouse model,  5 weeks  (early stage) or 8 weeks (late stage)  post Cre-induction. PyMT_spontaneous scRNAseq dataset: CD45+Viobility 488/520- cells from tumors of spontaneous MMTV-PyMT breast cancer mouse model. KPM scRNAseq dataset: unsorted cells from the tumors of a spontaneous KrasLSL-G12D/+;p53fl/fl;Msh2fl/fl (KPM) lung cancer mouse model.

first: Caro

last: Aarushi

first: Laoui

last: Damya


title: Bulk RNA sequencing of CD36⁺ and CD36⁻ cells isolated from hiPSC-derived airway epithelial cells

city: Kyoto city

first: Shimpei

last: Gotoh

state: Kyoto prefecture

address: 53 Kawahara-cho, Shogoin, Sakyo-ku

department: Department of clinical application

institute: Kyoto University CiRA

summary: To investigate whether CD36 can serve as a reliable surface marker for isolating deuterosomal cells from human induced pluripotent stem cell (hiPSC)-derived airway epithelial cells, we sorted CD36-positive and CD36-negative populations at an early stage of differentiation and performed bulk RNA sequencing. CD36⁺ cells were highly enriched for deuterosomal and multiciliated cell markers

overall_design: CD36-positive and -negative cells were isolated by flow cytometry from air–liquid interface (ALI) cultures of airway epithelial cells differentiated from the B2-3 SFTPC-GFP reporter hiPSC line　(Gotoh et al.,2014). This reporter line was originally established from the 201B7 hiPSC line, which was derived from a healthy donor (Takahashi et al., 2007). Total RNA of CD36-positive and CD36-negative cells was isolated using the RNeasy Micro Kit (Qiagen) following the manufacturer’s instructions. RNA integrity for each sample was assessed using the Agilent 2100 BioAnalyzer, with all samples showing an RNA integrity number (RIN) exceeding 9.5.

first: Haruka

last: Yamaki

first: Satoshi

last: Konishi


title: Palmitoleic acid promoted by BMPR2 signaling primes CD169 macrophages and alleviates liver fibrosis [albcre_bmpr2]

first: Jinkun

last: Zhong

overall_design: RNA-seq profling of livers from liver-specific knockout of BMPR2 mice(LKO) and wildtype mice(WT).

first: Hong

last: Ma

first: Qilin

last: Peng

first: Mengyuan

last: Dai

first: Linyuan

last: Wang

first: Jianan

first: Danchun

last: Huang

first: Yang

last: Liu

first: Yan

last: Tang

first: Wei

last: Xu

first: Qi-qun

first: Shuwen

last: Qian


title: Epigenetic approaches by analysis of circulating miRNAs and global DNA methylation in veterinarians occupationally exposed to volatile anesthetics

city: Botucatu

first: Tony

middle: Fernando

last: Grassi

state: São Paulo/SP

address: Av. Professor Mário Rubens G. Montenegro, s/nº

country: Brazil

institute: 1São Paulo State University (UNESP), Medical School

summary: Occupational exposure to waste anesthetic gases (WAGs) in veterinary operating rooms represents a potential health concern due to limited environmental monitoring. In this study, plasma samples from veterinarians chronically exposed to isoflurane and sevoflurane and from matched unexposed controls were analyzed to investigate molecular alterations associated with WAG exposure. Circulating microRNA expression was profiled using the NanoString nCounter® Human v3 miRNA Expression Assay. Differential expression analysis was performed to identify miRNAs associated with occupational exposure. Additional bioinformatic analyses were conducted to explore the potential biological pathways related to the identified miRNAs.

overall_design: This study investigated the impact of occupational exposure to waste anesthetic gases (WAGs) in veterinarians working in veterinary operating rooms. Plasma samples from 57 individuals were analyzed, including 29 exposed subjects and 28 matched unexposed controls. Circulating microRNA expression was profiled using the NanoString nCounter® Human v3 miRNA Expression Assay. Differential expression analysis was performed to identify miRNAs associated with WAG exposure.

last: Fernando Grassi

first: Mariane

last: Aparecida Pereira Silva

first: Maria

last: Vitória Destro

first: Iael

last: Weissberg Minutentag

first: Patrícia

last: Pintor dos Reis

first: Bruno

last: Spinosa De Martinis

first: Mónica

last: Cappetta

first: Leandro

last: Gobbo Braz

first: Mariana


title: Cholesterol-rich diet exacerbates steatohepatitis in the STAM mouse model

city: Nuthetal

first: Heike

last: Vogel

state: BRB

address: Arthur-Scheunert-Allee 114-116

department: Department of Experimental Diabetology

institute: German Institute of Human Nutrition Potsdam-Rehbruecke

summary: Metabolic dysfunction-associated steatotic liver disease (MASLD) is a widespread chronic liver disorder spanning simple steatosis to metabolic dysfunction-associated steatohepatitis (MASH) and fibrosis. Suitable animal models are crucial for therapeutic development, yet many fail to reproduce progression to fibrosing MASH. Here, we employed the STAM model in male C57BL/6J mice to simulate MASLD and investigated whether a high-fat, high-cholesterol (HFHC) diet would intensify disease progression compared with the standard high-fat (HF) diet. HFHC feeding in the STAM model accelerated hepatic lipid accumulation, crown-like structure formation, inflammation, and fibrosis, as demonstrated by histological analyses. Transcriptomic profiling and pathway enrichment confirmed activation of lipid and cholesterol metabolism, regulation of TNF production, and inflammatory signaling. Reactome analysis further indicated alterations in extracellular matrix formation and immune system pathways, consistent with advanced fibrotic progression. In summary, integration of HFHC feeding with the STAM model produced a more severe liver phenotype that better mirrors the pathological spectrum of MASLD.

overall_design: At age of two days male mice were injected with single subcutaneous dose of 200 µg streptocotozin dissolved in 0.1 M citrate buffer (pH 4.5) (STZ, S).  At 4 weeks of age mice were switched to ad libitum feeding of a high-fat diet (HF; Ssniff) or a cholesterol-enriched high-fat diet (HFHC; Altromin). Mice were sacrificed after a 6-hour fast at 8 or 12 weeks of age.

first: Wenke

last: Jonas