title: miRNA expression profiling of human atherosclerotic plaques

first: Yuanyuan

last: Wei

summary: To characterize transcriptional and post-transcriptional changes during human atherosclerosis development, we then performed gene expression profiling analysis using data obtained from RNA-seq of 3 different stages of human carotid atherosclerotic plaques.

overall_design:
Comparative miRNA expression profiling analysis of RNA-seq data for early fibroatheromas (EFAs), late fibroatheromas (LFAs) with thick ﬁbrous caps, and thin-cap fibroatheromas (TCFAs).
>>>Raw data for human samples not available due to patient privacy concerns<<<

first: Yuhuai

last: Xie

first: Zhen

last: Zhao


title: Antibody Mediated Diversification of Primary and Secondary Humoral Immune Responses

status: Public on Apr 02 2026

city: New York

first: Gabriela

middle: S

last: Silva Santos

state: NY

address: 1230 York Avenue

country: USA

institute: The Rockefeller University

type: Expression profiling by high throughput sequencing

summary: Humoral immune responses diversify over time but whether secreted antibodies influence this process is unknown. Using antibody secretion-deficient mice this study shows a profound impact of secreted antibodies on the evolution of B cell diversity after vaccination.

overall_design: WT, M-only, mM-only mice were immunized using RBD, Spike or HIV proteins as indicated. Draining lymph nodes have been harvested at D14 after primary or secondary immunization for RBD and Spike immunization, and at D12 for HIV protein immunization. B cell populations have been sorted and analyzed using 10X genomics scRNAsequencing for BCR sequences and hashtags.

first: Dennis

last: Schaefer-Babajew

first: Laurine

last: Binet

first: Chiara

last: Ruprecht

first: Lachlan

middle: P

last: Deimel

first: Mohamed

middle: A

last: ElTanbouly

first: Dounia

last: Gharrassi

first: Gabriella

last: Lima dos Reis

first: Clara

last: Uhe

first: Kai-Hui

last: Yao

first: Brianna

last: Hernandez

first: Parul

last: Agrawal

first: Anna

last: Gazumyan

first: Leonidas

last: Stamatatos

first: Harald

last: Hartweger

first: Michel

middle: C

last: Nussenzweig


title: Zinc-dependent RNA-Binding Protein Controls Hepatocyte Senescence and Recovery from Alcohol-related Liver Failure

city: Durham

first: Rajesh Kumar

last: Dutta

state: NC

address: 1049 Umstead Grove Way

department: Medicine

institute: Duke University

summary: Why alcohol-induced liver disease (ALD) resolves after abstinence in most people but progresses to liver failure in others remains poorly understood. Experimental models show that increased exposure to pro-inflammatory cytokines exacerbates ALD, yet clinical trials targeting these cytokines have failed.  The TNFα-inducible zinc finger protein 36 (ZFP 36) family of RNA binding proteins controls the outcomes of TNFα exposure by destabilizing suites of mRNAs that execute the pleiotropic downstream actions of TNFα. To investigate the role of RNA binding protein ZFP36L1 in regulating hepatocyte fate and its contribution in the progression of ALD.We selectively deleted ZFP36L1 in  mouse hepatocytes (ZFP36L1HEPKO) to assess its impact on ALD progression, transcriptional reprogramming, senescence and direct mRNA targets. In parallel, we analyzed human liver explants to evaluate ZFP36L1 in relation to hepatocyte senescence, disease severity, and zinc-dependent regulation.Deletion of ZFP36L1 exacerbated ALD and activated transcriptional programs driving ductal trans-differentiation, inflammation and senescence.  Mechanistically, ZFP36L1 directly destabilized Cdkn1a (p21) and Jag1 mRNAs, thereby suppressing hepatocyte senescence and JAG1-NOTCH signaling. In human liver explants, ZFP36L1 activity declined in parallel with increasing hepatocyte senescence and ALD severity, and was closely associated with impaired zinc-dependent signaling. Manipulation of zinc availability altered ZFP36L1 activity and expression of its direct targets.  These findings uncover a zinc-dependent ZFP36L1-regulon that governs hepatocyte fate by repressing p21- and JAG1-driven senescence and NOTCH activation, and highlight ZFP36L1as a promising therapeutic target in ALD.

overall_design: RNA-seq of livers from liver-specific ZFP36L1 knockout and fl/fl control adult male mice

first: Rajesh

middle: K

first: Perry

last: Blackshear

first: AnnaMae

last: Diehl


title: Spatial transcriptomic analysis of mouse liver in a high-fat diet model with kimchi supplementation

city: Gwangju

first: Wooje

last: Lee

address: 86, Kimchiro, Namgu

department: Kimchi Healthcare Research Group

country: South Korea

institute: World Institute of Kimchi

summary: This study generated 10x Genomics Visium spatial transcriptomic profiles from mouse liver sections to investigate the effects of high-fat diet and kimchi supplementation on hepatic gene expression organization. Samples were obtained from male C57BL/6J mice assigned to normal diet (ND), high-fat diet (HFD), high-fat diet supplemented with spontaneously fermented kimchi (S-K), and high-fat diet supplemented with starter kimchi fermented with Leuconostoc mesenteroides KCKM0828 (LMS-K). The dataset was designed to assess diet-associated transcriptional changes across the liver lobule and to evaluate how kimchi intervention modulates zonation-related hepatic programs along the portal-to-central axis. Raw FASTQ files were prepared for SRA submission through GEO, and processed outputs include filtered gene-barcode matrices, HDF5 matrices, spatial coordinate files, scale factor files, tissue images, and summary reports.

overall_design: Male C57BL/6J mouse liver samples from dietary intervention groups were analyzed using 10x Genomics Visium Spatial Gene Expression. Each sample represents one liver section profiled as an independent Visium library. The four samples correspond to the following conditions: Liv_N-3, ND; Liv_H-4, HFD; Liv_K-4, S-K; and Liv_S2-7, LMS-K. The study was designed to compare spatially resolved hepatic transcriptional patterns among dietary groups, with emphasis on zonation-associated differences across the portal-to-central axis.

first: Ha-Rin

last: Moon


title: The genomic landscape of HSF1 binding in beige adipocytes.

status: Public on Apr 03 2026

city: Shanghai

first: Yu

last: Li

address: Minhang Dongchuan Road No.500

country: China

institute: East China Normal University

type: Genome binding/occupancy profiling by high throughput sequencing

summary: To understand the mechanism of HSF1 in beige adipocytes we analysis the HSF1 binding genomic landscape.

overall_design: The genome-wide binding pattern of HSF1 was examined in C3H10T1/2-derived beige adipoctyes.


title: Mitochondrial L-2-hydroxyglutarate is a physiologic signaling metabolite [ATAC-seq]

city: Chicago

first: Navdeep

middle: S.

last: Chandel

state: IL

address: 303 E. Superior Street

department: Department of Medicine & Biochemistry and Molecular Genetics

institute: Northwestern University

summary: L-2-hydroxyglutarate (L-2-HG) is a low-abundance metabolite in mammals due to the mitochondrial enzyme L-2-HG dehydrogenase (L2HGDH), which oxidizes L-2-HG to 2-oxoglutarate (2-OG) to prevent its accumulation. In humans lacking L2HGDH activity, L-2-HG builds up, leading to L-2-hydroxyglutaric aciduria, a rare childhood neurometabolic disorder. Thus, L-2-HG is often classified as a toxic metabolite. Furthermore, L-2-HG is produced in response to hypoxia, acidic pH, and electron transport chain (ETC) impairment. However, whether L-2-HG has a physiological function is unclear. Here, we investigated whether L-2-HG qualifies as a physiologic signaling metabolite by testing three criteria: regulated levels, defined molecular targets, and a measurable physiological function. We report that an elevated mitochondrial NADH/NAD⁺ ratio drives malate dehydrogenase 2 (MDH2) to reduce 2-OG into L-2-HG, while L2HGDH oxidizes L-2-HG back to 2-OG within the mitochondrial matrix without requiring a functional ETC. Proteome integral solubility alteration (PISA) assays revealed the KDM4 family of H3K9 demethylases as L-2-HG-responsive targets. Consistent with this, elevated L-2-HG repressed nascent transcription of specific gene sets and promoted accumulation of the repressive histone mark H3K9me3 at those loci. In vivo, ubiquitous early embryonic L2HGDH overexpression lowered L-2-HG, reduced postnatal growth, and caused postnatal mortality. Despite systemic lowering of L-2-HG levels, kidneys exhibited a unique vulnerability, displaying functional impairments and histologic alterations. Mechanistically, L-2-HG depletion in the postnatal kidney resulted in the loss of H3K9me3 specifically at L1MdTf retrotransposons, leading to their derepression and coinciding with activation of the integrated stress response (ISR) and inflammation pathways. Our findings establish mitochondrial L-2-HG as a physiologic signaling metabolite that couples mitochondrial redox state to chromatin regulation, which is essential for postnatal kidney function and survival. These findings suggest that metabolites previously regarded as toxic may also serve critical physiological functions.

overall_design: ATAC-seq was performed on mouse embryonic stem cells (mESCs) transduced with empty vector (EV) or rat L2HGDH cDNA and treated with 500 nM myxothiazol for 16 hours.

first: Ram

last: Chakrabarty

first: Qiushi

last: Jin

first: Dongmei

last: Wang

first: Rogan

last: Grant

first: Feng

last: Yue


title: Mitochondrial L-2-hydroxyglutarate is a physiologic signaling metabolite [mRRBS]

type: Methylation profiling by high throughput sequencing

overall_design: mRRBS was performed on mouse embryonic stem cells (mESCs) transduced with empty vector (EV) or rat L2HGDH cDNA and treated with 500 nM myxothiazol for 16 hours.

first: Kathryn

last: Helmin

first: Benjamin D

last: Singer


title: Effects of LPS-induced neutrophil senescence on gene expression

city: Chongqing

first: Yaqian

last: Duan

address: 76 Linjiang Road, Yuzhong District

institute: The Second Affiliated Hospital of Chongqing Medical University

summary: Neutrophils are the most abundant leukocytes in human peripheral blood but exhibit considerable heterogeneity, with distinct subsets characterized by diverse functional states. Lipopolysaccharide (LPS) has been shown to drive neutrophil senescence, a process distinct from apoptosis. However, the global transcriptional landscape of LPS-induced aged neutrophils remains largely unexplored. Here, we performed whole-transcriptome RNA sequencing (RNA-seq) on primary human neutrophils isolated from healthy donors. Neutrophils were treated with LPS to induce senescence, with untreated cells serving as controls. Our comparative transcriptomic analysis revealed that senescent neutrophils display significant upregulation of multiple genes associated with neutrophil extracellular trap (NET) formation compared to non-aged neutrophils. This dataset provides a comprehensive resource for investigating the molecular signatures underlying neutrophil senescence and its potential implications in inflammation and host defense.

overall_design: Freshly isolated human peripheral blood neutrophils were stimulated with lipopolysaccharide (LPS, 10 ng/mL) to induce senescence, and RNA sequencing analysis was performed to compare the transcriptomic profiles between LPS-treated aged neutrophils and untreated non-aged neutrophils.

first: Shan

last: Yu

first: Xuanyi

first: Rong

last: Wu

first: Yuqing

last: Zhao

first: Xuehua

last: Kong

first: Liang


title: ONX-0914 Ameliorates Hepatic Steatosis in MASLD by Stabilizing HDAC2 and Inhibiting SREBP1 Acetylation via PSMB8 Inhibition

city: Wuhan

first: Ye

last: zhou

address: 1037 Luoyu Road, Hongshan District, Wuhan, Hubei, 430074, P.R. China

institute: Huazhong University of Science and Technology Tongji Medical College

summary: To investigate the mechanism by which ONX‑0914 ameliorates MASLD, HFD‑fed mice were randomized to receive the selective PSMB8 inhibitor ONX‑0914 (10 mg/kg in 10% captisol, s.c., every other day) or vehicle control for an additional 6 weeks while remaining on the HFD. At the experimental endpoint, liver tissues were collected for RNA sequencing.

overall_design: Liver tissues were collected from two groups of male C57BL/6J mice: (1) HFD‑fed + vehicle, (2) HFD‑fed + ONX‑0914 (10 mg/kg, s.c., every other day, 6 weeks). Each group contained 2 biological replicates. Total RNA was extracted and subjected to Illumina NovaSeq 6000 sequencing (PE150).


title: SAM68 is a crucial post-transcriptional regulator of cardiogenesis

city: Barcelona

first: Elias

last: Bechara

address: Dr Aiguader 88

country: Spain

institute: CRG

summary: RNA-binding proteins (RBPs) of the STAR family play important roles in mammalian development, yet their precise contributions to lineage specification remain incompletely understood. Here, using CRISPR–Cas9 knockout models combined with multi-omics approaches, we investigate the roles of two STAR proteins, SAM68 and QKI, in mouse embryonic stem cells (mESCs). Both RBPs contribute to mESC proliferation, self-renewal, and efficient differentiation into cardiomyocytes. Our data support a role for QKI in the regulation of cardiac differentiation programs. In addition, our analyses reveal a previously underappreciated contribution of SAM68 to cardiomyocyte specification, acting through multiple post-transcriptional mechanisms. Despite belonging to the same protein family, SAM68 and QKI regulate largely distinct post-transcriptional programs during differentiation. Specifically, SAM68 modulates alternative splicing and promotes the biogenesis of a subset of cardiac-enriched circular RNAs through binding to intronic regions flanking back-splice sites and association with NF90/110. In addition, SAM68 associates with untranslated regions of key differentiation-related transcripts, including Gata4 mRNA, a key transcription factor regulating cardiogenesis, and likely functions in complex with other RNA-binding proteins to regulate these transcripts. Together, these findings identify SAM68 as a regulator of multiple RNA processing pathways contributing to cardiomyocyte differentiation and provide insight into how STAR proteins shape post-transcriptional gene regulatory networks during early development.

overall_design: Sam68 iCLIP2 experiment using E14 cells at Day0, Day3 and Day10 of differentiation. The experiment was performed as 5 replicates per time point.

first: Laura

last: Broglia

first: Sarah

last: Wolf

first: Melina

last: Klostermann

first: Kathi

last: Zarnack

first: Gian Gaetano

last: Tartaglia


title: A novel platform for parallel T cell receptor cloning and testing enables anti-neoantigen tumor immunotherapy

city: Pittsburgh

first: Warren

last: Shlomchik

state: PA

address: 5051 Centre Ave

institute: University of Pittsburgh

summary: Tumor-infiltrating CD8 cells recognize neoantigens created by tumor-specific mutations. Nonetheless, even after checkpoint inhibitor therapy, most patients progress. A deeper understanding of anti-tumor responses could facilitate development of better therapies. To enable such studies, we applied TCXpress, a high throughput platform that clones fully expressible TCRs from single cells into retro- or lenti- viral vectors without sequencing or gene synthesis, to study TCRs from CD8 cells infiltrating mouse MC38 tumors. We expressed cloned TCRs in reporter cells and interrogated TCR specificity by coculturing them with B6WT3 cells transduced with tandem minigenes encoding predicted neoantigens. We isolated TCRs reactive against epitopes from mutant Rpl18, ADPGK, PSMD2 and Zc3h7b along with self-reactive TCRs that recognize normal B6 and MC38 cells. Importantly, we successfully treated MC38-bearing mice with T cells transduced with anti-Rpl18 TCRs. These results establish a system that could be used to study many types of T cell responses and validates a therapeutic approach that could be tested in the clinic.

overall_design: Whole genome sequencing of MC38-Leiden cells was performed to identify potential neoantigens compared to reference B6 germline.

first: Alexander

middle: M

last: Rowe

first: Smriti

last: Chaurasia

first: Wenzhong

last: Wei

first: Lau

last: Garcia-Dieguez

first: Katherine

last: Querry

first: Jonathon

last: Schiebel

first: Christy

last: Smolak

last: Muralles

first: Daniel

last: Wikenheiser

first: Kevin

last: Quann

first: Collin

last: Pirner

first: Kentin

last: Codispot

first: Mark

middle: J

middle: D


title: Biallelic inactivating variants in the chromatin remodeler DMAP1 cause a syndromic neurodevelopmental disorder

city: Philadelphia

first: Dong

last: Li

address: 3615 Civic Center BLVD

institute: Children's Hospital of Philadelphia

summary: Chromatin remodeling is a dynamic epigenetic process that alters chromatin structure to gauge gene accessibility, enabling precise spatiotemporal gene expression, with disruptions often underlying neurodevelopmental disorders (NDDs), although the mechanistic underpinning remains incompletely understood. Despite essential roles in chromatin remodeling processes such as DNA methylation, and histone acetylation and deposition, DMAP1 has not been implicated in human disease. We identified 20 individuals from 16 families with a syndromic NDD carrying homozygous or compound heterozygous variants in DMAP1. Neural-specific knockdown of its Drosophila ortholog, dDMAP1, caused pupal lethality, structural defects in the mushroom body (MB), decreased dendrite length, abnormal social behavior and mechanical-induced seizures. Wildtype human DMAP1 could largely compensate for the loss of dDMAP1 in knockdown flies, whereas patient variants failed to restore or differentially rescued the phenotypes, confirming their pathogenicity with differing severity. Transcriptome profiling of dDMAP1 knockdown fly brains nominated Cbl and SF1 as downstream targets. Their overexpression rescued the aforementioned lethality and MB defects. Finally, a DNA methylation episignature was identified, leading to the molecular diagnosis of an additional patient. Our findings demonstrate that biallelic inactivating variants in DMAP1 cause a novel syndromic NDD, expanding the short list of recessive disease-causing genes within the epigenetic machinery.

overall_design:
fly brain tissues harvested at the 3rd instar larval stage from WT controls and elav-Gal4 driving knockdown flies for DMAP1

***************************************************************
The table below lists GEO accessions reused/reanalyzed for this study.
***************************************************************


title: A study for intratumoral heterogeneity of glioblastoma [scRNA-seq]

city: Shenyang

first: Sun

middle: Xiang

last: Hao

state: Liaoning

address: puhe road 77th

institute: CMU

summary: Intratumoral heterogeneity (ITH) is a key driver of therapy resistance in glioblastoma (GBM). This study established a five-gene signature-based gITH classifier through multi-omics analysis, demonstrating robust prognostic predictive value. High-gITH tumors exhibited enhanced molecular complexity, with PDLIM4 identified as the central regulator showing strong correlations with stem-like properties and poor clinical outcomes. Functional validation confirmed that PDLIM4 knockdown suppressed ITH and tumor progression. Our work not only establishes a transcriptome-based quantification framework for GBM heterogeneity, but also reveals PDLIM4 as a promising therapeutic target, offering novel precision medicine strategies.

overall_design: we conducted an integrated study combining multi-region transcriptomics and machine learning. A novel computational framework (gITH score) was developed to objectively quantify transcriptional heterogeneity in GBM. Furthermore, a clinically validated machine learning classifier, derived from heterogeneity levels, was established to stratify patients and predict survival outcomes. Through systematic functional validation, one gene was identified as a central regulator of stemness and developmental plasticity, mechanistically linking it to heterogeneity-driven tumor progression.


title: Phosphorylation of serines 287/288 in DEK regulates cell-type-specific chromatin occupancy and compaction [ATAC-seq]

city: Suzhou

first: Gongjie

last: Wu

address: Renai Road

institute: Xi'an Jiaotong-Liverpool University

summary: The conserved multifunctional chromatin modulator and oncogene DEK exhibits context-dependent genomic binding and function, but how these activities are regulated in cancer remains poorly understood. Using multi-omics and biochemical approaches, we find that while DEK predominantly occupies promoter-proximal regions in HeLa cells and primary melanocytes, its chromatin binding is dramatically reduced in melanoma cell lines—despite DEK overexpression. We attributed this to CK2-mediated phosphorylation, which governs DEK chromatin association and transcriptional output in a cell-type-specific manner. Phosphoproteomics identified 34 phosphorylation sites, including S287 and S288 within the DEK C-terminal DNA-binding domain. Strikingly, CK2 inhibition and concomitant loss of phosphorylation at S287/S288 triggered DEK redistribution to promoter regions, coinciding with transcriptional repression of oncogenic pathways and global chromatin compaction. Melanoma subtypes showed divergent responses: NRAS-mutant cells displayed dynamic, phosphorylation-dependent DEK redistribution, whereas BRAF-mutant cells lacked detectable DEK binding. Our work establishes DEK as a phosphorylation-sensitive regulator of chromatin states, with CK2-mediated modification orchestrating its tumor-specific regulatory functions. These findings nominate phospho-DEK as a potential biomarker and therapeutic target in melanoma and possibly other cancers.

overall_design:
This submission includes chromatin accessibility, chromatin occupancy, and gene expression profiling from human cancer and non-cancer cell models under defined genetic and pharmacological perturbations.
ChIP-seq samples were generated from HeLa S3 cells, primary human melanocytes, and SK-Mel-103 melanoma cells to profile chromatin binding under control conditions and following treatment with the CK2 inhibitor CX-4945.
Gene expression microarray samples were obtained from parental HeLa S3 cells and DEK knockout (DEK KO, clone 1E7) HeLa S3 cells, allowing comparison of transcriptomic changes associated with loss of DEK expression.
ATAC-seq samples were generated from HeLa S3 cells, DEK KO HeLa S3 cells (clone 1E7), and SK-Mel-103 cells to assess chromatin accessibility changes associated with genetic depletion of DEK and pharmacological CK2 inhibition. SK-Mel-103 samples include control conditions as well as CX-4945 treatment and DEK siRNA-mediated knockdown.

first: Ferdinand

last: Kappes


title: Mitochondrial L-2-hydroxyglutarate is a physiologic signaling metabolite [m6A-seq]

first: Navdeep

last: Chandel

overall_design: m6A-seq was performed on untreated or myxothiazol-treated (500 nM, 16 hours) mouse embryonic stem cells (mESCs) transduced with empty vector (EV) or rat L2HGDH cDNA .

first: Ram

last: Chakrabarty

first: Michael

last: Kahl

first: Zhaofa

last: Xu

first: Rogan

last: Grant

first: Yongchao

last: Ma


title: Bimodal Imaging-Guided Temporal Optimization of Combined Chemotherapy and DC Vaccine Immunotherapy

city: Beijing

first: Xingru

last: Tan

address: No. 1 Shuaifuyuan, Dongcheng District, Beijing

institute: peking union medical college

summary: Dendritic cell (DC) vaccines represent a clinically promising strategy for cancer treatment using a "prime-boost" approach. Achieving synergistic antitumor efficacy with chemotherapy requires precise temporal coordination, yet the timing of post-chemotherapy vaccination remains unstandardized, hindering its clinical translation. To address this, we designed a nanoprobe (Fe@O-DT(Cy7)) and applied bimodal imaging (FMI/MPI) to track DC vaccine migration and apoptosis after oxaliplatin treatment in a colorectal cancer model. We identified an optimal therapeutic window on day 7 post-chemotherapy, coinciding with peak immunogenic cell death (ICD). DC vaccination within this 7-d window ensured optimal lymph node homing, minimal apoptosis, and enhanced antitumor efficacy, characterized by immune cell infiltration and tertiary lymphoid structure (TLS) formation. Mechanistically, this window correlated with NF-κB pathway activation and CCL19/CCL21 upregulation. This study elucidates the role of the LTα1β2-LTβR axis in ICD-modulated DC vaccination and reveals the function of TLS in DC vaccine efficacy.

overall_design: 1、Tumor and lymph node tissues from different treatment groups, including control, DC, OXA, OXA+DC Day 3, OXA+DC Day 7, and OXA+DC Day 10 groups. 2、Lymph node tissues from different drug stimulation groups, including control, TNF-α, DC, and OXA groups. 3、For the combination therapy of oxaliplatin and DC vaccine, lymph node tissues were collected from eight groups with varying time intervals between oxaliplatin administration and subsequent DC vaccine treatment: 0.5, 1, 2, 3, 5, 7, 10, and 14 days. 4、Lymph node tissues from groups receiving two consecutive DC vaccine treatments with different intervals, including 3, 7, and 10 days. Lymph node tissues include popliteal and inguinal lymph nodes.Tissues were immediately snap-frozen in liquid nitrogen after removal.

first: Juncheng

first: Yafei

last: Qi


title: De novo transcriptome assembly of the Korean minipig (Sus scrofa) [II]

city: Seoul

first: Suyeon

last: Wy

address: 20, Neungdong-ro, Gwangjin-gu

department: Department of Biomedical Science and Engineering

institute: Konkuk University

summary: Minipigs are animal models widely used in biomedical studies due to their physiological and anatomical similarities to humans. However, a comprehensive resource for the Korean minipig (Sus scrofa) transcriptome remains unavailable. In this study, we constructed a de novo transcriptome of the Korean minipig using RNA-seq data obtained from ten tissues across ten samples. The final assembly comprised 57,085 coding transcripts with an average length of 3,075 nucleotides and an N50 of 4,258 nucleotides. In total, 65.4% of the transcripts were annotated, and biological functions were assigned. Transcript expression profiling and principal component analysis showed that samples clustered by tissue type, reflecting transcriptomic features shared across tissues. Comparative analysis demonstrated that the novel transcriptome assembly had contiguity and completeness comparable to those available for pig and minipig breeds. Overall, this study provides a comprehensive transcriptomic resource for the Korean minipig, facilitating further functional analyses.

overall_design: Analyzing transcriptomic features of two different types of Korean minipigs, ET- and L-type minipigs, using RNA-seq data for ten tissues.

first: Woncheoul

last: Park

first: Jaebum

last: Kim


title: Cell size-dependent mRNA transcription drives proteome remodeling [RNA-seq]

city: Stanford

first: Dong Shin

last: You

address: 327 Campus Drive

institute: Stanford University

summary: Increasing cell size drives proteomic changes that impact cell physiology. However, the molecular basis of size-dependent proteome remodeling has remained unclear. Here, we develop an inducible Cyclin D1 expression system in human cells to generate proliferating cells spanning over a two-fold size range. We use this system to make comprehensive genome-wide measurements of mRNA and protein concentrations and stability. We find that protein and mRNA turnover rates are weakly related to cell size, but that mRNA concentrations are strongly size-dependent. This establishes that transcriptional regulation is the basis of proteome remodeling. Live-cell imaging of nascent mRNAs using the MS2 system is used to measure how transcriptional dynamics change with cell size. Larger cells prolong transcriptional bursts and shorten inactive periods between bursts but maintain similar burst amplitudes to achieve transcriptional scaling. Together, our results show how transcription is modulated by cell size to remodel the proteome and alter cell physiology.

overall_design: RNA-seq profiling of CCND1 overexpression or knockdown hTERT-RPE-1 lines with and without induction by doxycycline or DMSO at day 4 and day 5 of treatment. Two replicates are provided

first: Christopher

middle: H

last: Bohrer

first: Purva

last: Rumde

first: Ioannis

last: Sanidas

first: Matthew

last: Swaffer

first: Daniel

middle: R

last: Larson

first: Josh

middle: E

last: Elias

last: Lanz

first: Jan

last: Skotheim


title: Dynamic Suspension Culture System Reveals HIF-1α-Driven Stemness Maintenance via Dual Suppression of the p53 Pathway in Cancer Stem Cells

city: Chengdu

first: Xiaoli

last: Mu

address: No. 37 Guoxue Lane

institute: West China Hospital of Sichuan University

summary: Cancer stem cells (CSCs) drive tumor recurrence and therapeutic resistance; however, their investigation remains limited by inefficient enrichment strategies and an incomplete understanding of stemness regulation. In this study, we developed an optimized dynamic suspension culture system that efficiently enriches CSCs. Compared with conventional static culture, this system generated spheroids with increased expression of stemness markers, enhanced proliferative capacity, and improved stability. Transcriptomic analyses further demonstrated that the system mimics fluid shear stress within the tumor microenvironment (TME), thereby activating the HIF-1 signaling pathway. Mechanistically, HIF-1α sustains stem cell activity through dual suppression of the p53 pathway: it directly represses p21 transcription by binding to its promoter, while simultaneously upregulating MDM2 to drive the subsequent degradation of p53. Based on this mechanism, combining the HIF-1α inhibitor PX-478 with the p53 activator doxorubicin synergistically suppressed CSC properties in vitro and significantly inhibited tumor growth in a subcutaneous xenograft model. Collectively, these findings establish an effective CSC enrichment strategy and identify the HIF-1α/p53 axis as a critical regulator of cancer stem cell properties and a promising therapeutic target.

overall_design: This study employed different methods to enrich B16F10 cancer stem cells (CSCs): conventional three-dimensional culture and dynamic suspension culture (the methods used in this study). In addition, HIF-1α knockdown, overexpression, and control cells were also collected. Each experiment was repeated three times and then pooled. After enrichment, all cell types were lysed using Trizol reagent and thoroughly homogenized to ensure complete cell disruption before total RNA was extracted.

first: Yongxin

last: Yu


title: Comparative single-cell multiomic analysis reveals evolutionarily conserved and species-specific cellular mechanisms mediating natural retinal aging [Zebrafish multiome_ATAC]

city: Baltimore

first: Pin

last: Lyu

address: 400 N Broadway

institute: JHU

summary: Biological age is a major risk factor in the development of common degenerative retinal diseases such as age-related macular degeneration and glaucoma. To systematically characterize molecular mechanisms underlying retinal aging, we performed integrated single- cell RNA- and ATAC-Seq analyses of the retina and retinal pigment epithelium (RPE) across the natural lifespan in zebrafish, mice, and humans. By profiling gene expression and chromatin accessibility, we identified extensive cell type- and species-specific aging-dependent changes, with a much smaller number of broadly expressed and conserved genes that include regulators of inflammation and autophagy. We constructed predictive aging clocks for retinal cell types and observed dynamic, reversible shifts in cellular age following acute injury. Spatial transcriptomic analysis revealed region-specific aging signatures and proximity effects, with Müller glia exhibiting pro-rejuvenating influences on neighboring neurons. Targeted Müller glia-specific induction of Yamanaka factors reduced molecular age in rod photoreceptors and bipolar cells without altering glial age. Our findings define conserved and divergent regulatory and signaling pathways mediating retinal aging, highlighting Müller glia as potential therapeutic targets for combating age-associated retinal dystrophies.

overall_design: To comprehensively profile aging-dependent changes in gene expression and regulation at the cellular level, we performed combined snRNA- and ATAC-Seq at multiple timepoints across the full natural lifespan of zebrafish and mice. In zebrafish, we profiled retina and RPE from 1, 3, 6, 12, 18, 22, 24, 30, 36, and 48 months of age, with 8-10 samples pooled in each of 1-2 experimental replicates (Fig. 1A). For mice, we profiled retina and RPE from 5, 12, 17, 32, 49, 68, 91, 106, and 120 weeks of age analyzing a pool of 2 male and 2 females in 2 experimental replicates. Additional RPE enriched samples from mice were collected at 5, 49, and 91 weeks of age.

first: Isabella

last: Palazzo

first: Yang

last: Jin

first: Leah J.

last: Campbell

first: Clayton P.

last: Santiago

first: Rogger P.

last: Carmen-Orozco

first: Thanh

last: Hoang

first: Jared A.

last: Tangeman

first: Alexander D.

first: Shawn H.

last: Yang

first: Jianming

last: Shao

first: Rui

last: Chen

first: David

last: Hyde

first: Jiang

last: Qian

first: Seth

last: Blackshaw


title: HIRA-mediated H3.3 deposition preserves hepatocyte cell identity during non-proliferative liver aging [ATAC-seq]

city: La Jolla

first: Xue

last: Lei

state: CA

address: 10901 N Torrey Pines Rd

institute: SBP medical discovery institute

summary: Age-associated functional decline is partly driven by progressive chromatin degeneration. Conversely, maintenance of chromatin integrity preserves cell identity and promotes healthy aging, albeit through different mechanisms in proliferating and non-proliferating cells. However, whether tissue regeneration with its associated cell proliferation can rescue defects in epigenetic maintenance in otherwise non-proliferative cells remains unclear. The histone chaperone HIRA deposits the histone variant H3.3 in a DNA replication–independent manner, leading to its accumulation in aging, non-proliferating cells. Here, we show that hepatocyte-specific loss of HIRA causes loss of cell identity, metabolic dysfunction, and accelerated fibrotic pathology with age. Transcriptomic and epigenomic analyses indicate that HIRA–H3.3 preserves chromatin integrity and sustains transcription of highly expressed genes. Partial hepatectomy, associated with induced proliferation, restores cell identity with compensatory deposition of canonical histones H3.1/2. Together, these results demonstrate that HIRA-mediated H3.3 deposition is essential for safeguarding cell identity and tissue function during aging of non-proliferating cells, but this function can be rescued by tissue regeneration with associated cell proliferation.

overall_design: ATAC sequencing was used to study the chromatin accessibility changes of HIRA KO livers in mice. We used female wildtype mice injected with AAV8 Tbg-Cre (control)  and HIRA fl/fl C57BL/6 mice injected with AAV8 Tbg-Cre (KO mice). Mice were injected at 6 months and then aged for an additional 12 months. Chromatin accessibility analysis was performed using data obtained from bulk-ATAC seq of 4 samples (2 independent replicates per experimental group).

first: Rouven

last: Arnold

first: Rabi

last: Murad

first: Peter

last: Adams


title: Genome-wide histone acetylation underscores tumor intrinsic immune signaling induced by photothermal therapy in ovarian cancer

city: Washington

first: Reddick

last: Walker

state: DC

address: 800 22nd St NW SEH 8960

department: MITM

institute: The George Washington University

summary: Ovarian cancer remains the most lethal gynecological malignancy with few durable responses to standard of care platinum-based chemotherapies and emerging immunotherapies. This poor response is attributed to the immunosuppressive tumor microenvironment which lacks effector lymphocytes necessary for tumor clearance. Therefore, increasing tumor intrinsic immune signaling remains a therapeutic goal for OC. One strategy for improved tumor intrinsic immune signaling relies on induction of tumor cell immunogenic cell death (ICD) features including tumor cell surface expression of Calreticulin and Hsp70 with concurrent release of immunostimulatory signals that activate innate and adaptive immune cells. The induction of ICD through a nanoparticle-based heat shock treatment, termed Prussian blue nanoparticle photothermal therapy (PBNP-PTT), has shown strong recruitment of antigen-specific lymphocytes to the tumor microenvironment in glioblastoma and neuroblastoma preclinical models. While the molecular signatures of ICD are well described, the epigenetic reprogramming during treatment-induced tumor intrinsic immune signaling remains unknown. Therefore, this study sought to determine the efficacy of PBNP-PTT heat shock in ovarian cancer and characterize epigenetic responses that accompany its induced tumor intrinsic immune signaling. We first demonstrate that PBNP-PTT activates tumor intrinsic immune signaling in ovarian cancer and leads to improved monocyte activation through tumor cell proinflammatory cytokine release and presentation of Calreticulin and Hsp70. Accompanying this intrinsic immunostimulatory effect, PBNP-PTT induces genome-wide chromatin accessibility via H3K9ac, which associates with chromatin remodeling and heat shock response genes. Lastly, we show targeting histone acetylation via histone deacetylase inhibition (Panobinostat) improves the tumor intrinsic immune signaling potential of PBNP-PTT. These findings indicate that PBNP-PTT activates the release of immunostimulatory factors in ovarian cancer cells through H3K9ac rewiring and suggests the potential of combination with HDACi to improve tumor intrinsic immune signaling in ovarian cancer.

overall_design: These data include H3K9ac-targetd CUT&RUN-sequencing files post treatment with Prussian blue nanoparticle photothermal therapy at 10 minutes or 6 hours post treatment. A prussian blue only control (POB, no heat) is used to normalize the signatures gained from the photothermal treatment.

last: Walker III

first: Melissa

last: Hadley

first: Jose

last: Colina

first: Kevin

last: Nestler

first: Abby

last: Lee

first: Elizabeth

last: Sweeney

first: Rohan

last: Fernandes

first: Katherine

middle: B

last: Chiappinelli


title: Comparative single-cell multiomic analysis reveals evolutionarily conserved and species-specific cellular mechanisms mediating natural retinal aging [Mouse_multiome_scRNA]

first: Pin

last: Lyu

first: Isabella

last: Palazzo

first: Yang

last: Jin

first: Leah J.

last: Campbell

first: Clayton P.

last: Santiago

first: Rogger P.

last: Carmen-Orozco

first: Thanh

last: Hoang

first: Jared A.

last: Tangeman

first: Alexander D.

last: Park

first: Shawn H.

last: Yang

first: Jianming

last: Shao

last: Chen


title: Mitochondrial L-2-hydroxyglutarate is a physiologic signaling metabolite [RNA-seq}

first: Navdeep

last: Chandel

overall_design: RNA-seq was performed on mouse embryonic stem cells (mESCs) transduced with empty vector (EV) or rat L2HGDH cDNA, as well as on Wild-Type and L2HGDH-Overexpressing (homozygous) mouse kidneys and livers.

first: Ram

last: Chakrabarty

first: Rogan

last: Grant


title: Comparative single-cell multiomic analysis reveals evolutionarily conserved and species-specific cellular mechanisms mediating natural retinal aging [Zebrafish_multiome RNA]

first: Rui

first: David

last: Hyde

first: Jiang

last: Qian

first: Seth

last: Blackshaw


title: CAR T therapy against the MiTF-driven protein GPNMB [Multiome-Seq]

city: Calgary

first: Sorana

last: Morrissy

state: Alberta

address: 3330 Hospital Dr NW

department: Department of Biochemistry and Molecular Biology

country: Canada

institute: University of Calgary

summary: CAR T therapy for solid tumors is constrained by the scarcity of safe, uniformly expressed cell-surface targets. Here we identify GPNMB, a MiT fusion-driven protein, as highly, homogeneously, and stably expressed in primary and relapsed translocation-positive alveolar soft part sarcoma (ASPS) and renal cell carcinoma (tRCC). We develop a GPNMB-directed CAR T cell product, GCAR1, which demonstrates potent activity against patient-matched cells, organoids and xenograft models. In a first-in-human treatment of two patients with relapsed/refractory, metastatic ASPS, GCAR1 is well tolerated and induces stable disease for up to 6 months, accompanied by resolution of many non-target lesions. GCAR1 T cells expand in peripheral blood as a polyclonal population and remain detectable for up to 6 months. Spatial transcriptomics identify multiple immunosuppressive niches adjacent to T cells infiltrating treatment-resistant lesions; biological PDL1 blockade or PD1 and TIGIT double knockout overcomes this resistance and shows synergistic activity with GCAR1 in xenograft models. Taken together, these data provide clinical proof-of-concept for treating solid tumors with CAR T cells targeting a surface antigen driven by an oncogenic gene fusion.

overall_design: Multiome-seq: Single cells were extracted from the single-patient's peripheral blood and tumour biopsy samples that were collected before and after GCAR1 treatment. Joint scRNA-and-scTCR libraries were preared using the 10x Genomics platform (10x Genomics Chromium GEM-X Single Cell V(D)J + 5′ Gene Expression (v3) kit).


title: Comparative single-cell multiomic analysis reveals evolutionarily conserved and species-specific cellular mechanisms mediating natural retinal aging [Mouse_multiome_scATAC]

first: Pin

last: Lyu

first: Isabella

last: Palazzo

first: Yang

last: Jin

first: Leah

first: Clayton

first: Rogger

first: Jared

first: Alexander

first: Shawn


title: HIRA-mediated H3.3 deposition preserves hepatocyte cell identity during non-proliferative liver aging [RNA-Seq 2]

first: Xue

last: Lei

overall_design: RNA sequencing was used to study the transcriptomic changes of HIRA KO livers across a timecourse in mice. We used female wildtype C57BL/6 mice injected with AAV8-Tbg-saCas9-sgRNA-Rosa26 (control) and female wildtype C57BL/6 mice injected with AAV8-Tbg-saCas9-sgRNA-HIRA (KO). Mice were injected at 6 months of age and livers were harvested at 3 weeks and 6 months post-injection. Gene expression analysis was performed using data obtained from bulk RNA-seq of 20 samples (5 independent replicates per experimental group).

first: Rouven

last: Arnold

first: Peter

last: Adams


title: HIRA-mediated H3.3 deposition preserves hepatocyte cell identity during non-proliferative liver aging [RNA-Seq]

overall_design: RNA sequencing was used to assess the effect of liver regeneration on the transcriptomic defects caused by HIRA loss in the livers of mice. HIRA was deleted using two independent approaches: Cre-mediated deletion in HIRAfl/fl C57BL/6 mice injected with AAV8-Tbg-Cre (with wildtype C57BL/6 mice injected with AAV8-Tbg-Cre as controls), and CRISPR-mediated deletion in wildtype C57BL/6 mice injected with AAV8-Tbg-saCas9-sgRNA-HIRA (with wildtype C57BL/6 mice injected with AAV8-Tbg-saCas9-sgRNA-Rosa26 as controls). Male mice were injected at 6 months of age, and partial hepatectomy was performed 4 weeks post-injection to induce liver regeneration. In a paired design, the resected liver tissue served as the pre-regeneration sample and the remaining liver was collected 5 days post-resection as the post-regeneration sample, such that each mouse served as its own control. Gene expression analysis was performed using data obtained from bulk RNA-seq of 16 samples (4 groups: control before regeneration, control after regeneration, HIRA KO before regeneration, HIRA KO after regeneration; 4 independent replicates per group, 2 from each deletion approach).


title: LINGO4 coordinates intrinsic IL-22 production and microbiota-dependent ILC3 homeostasis to regulate intestinal immunity

city: Saint Louis

first: Shitong

last: Wu

state: MO

address: 660 S Euclid Avenue

institute: Washington University in St. Louis

summary: LINGO4 is a leucine-rich repeat and immunoglobulin-like domain-containing transmembrane protein encoded immediately adjacent to Rorc, the gene for RORγt, raising the possibility that it contributes to the biology of RORγt⁺ lymphocytes. However, its impact on these cells and resistance to enteric infections has remained unknown. Here, we identify LINGO4 as a critical regulator of group 3 innate lymphoid cells (ILC3s). Lingo4–/– ILC3s exhibit a profound, cell-intrinsic defect in IL-22 production linked to impaired STAT3 activation, mitochondrial dysfunction, elevated reactive oxygen species, and increased apoptosis. In vivo, Lingo4 deficiency also drives a dysbiotic gut microbiota, resulting in an additional, microbiota-dependent loss of ILC3s. These combined defects increase susceptibility to C. difficile and C. rodentium, while Lingo4–/– driven IL-22 reduction confers protection against S. Typhimurium. Immunoprecipitation of tagged LINGO4 reveals interaction networks enriched in mitochondrial pathways, providing mechanistic insight into its role in ILC3 metabolic fitness and intestinal immunity.

overall_design: Bulk RNAsequencing of sort-purified lamina propria ILC3s from WT and Lingo4-/- KO mice

first: Jose

middle: L

last: Fachi

first: Tihana

last: Trsan

first: Susan

last: Gilfillan

first: Marina

last: Cella

first: Steven

last: Van Dyken

first: Marco

last: Colonna


title: HIRA-mediated H3.3 deposition preserves hepatocyte cell identity during non-proliferative liver aging [RNA-Seq HIRAKO_1Y]

overall_design: RNA sequencing was used to study the transcriptomic changes of HIRA KO livers in mice. We used male and female wildtype mice injected with AAV8 Tbg-Cre (control 1), HIRA fl/fl C57BL/6 mice injected with AAV8 Tbg-MCS (control 2) and HIRA fl/fl C57BL/6 mice injected with AAV8 Tbg-Cre (KO mice). Mice were injected at 6 months and then aged for an additional 12 months. Gene expression analysis was performed using data obtained from bulk-RNA seq of 18 samples (3 independent replicates per experimental group).


title: GPNMB as a viable CAR Target for MITF pathognomonic fusion-driven cancers: A first-in-person GCAR1 Trial [ASPS-02_VisiumHD]

first: Sorana

summary: CAR T therapy for solid tumours is limited by a lack of safe and uniformly expressed cell-surface targets. We identify the MITF-driven gene GPNMB as being highly, homogeneously and stably expressed from primary and relapsed translocation-positive alveolar soft part sarcoma (ASPS) and renal cell carcinoma (tRCC). We developed a 2nd generation GPNMB-targeting CAR T therapy (GCAR1) that shows activity against cells, organoids and PDX models. First-in-person GCAR1 treatment of a patient with metastatic ASPS was well tolerated and generated stable disease for 6 months, with the majority of non-target lung lesions resolving post-treatment. A diverse population of GCAR1 cells expanded in blood, peaking at 14 days post-treatment. Spatial transcriptomics revealed immunosuppression, including PDL1 expression, in close proximity to T cells infiltrating a treatment-resistant lung lesion, and PDL1 blockade showed synergy with GCAR1 in a PDX model. Our data provide a proof-of-concept for targeting oncogenic translocation-driven genes with CAR T therapy.

overall_design: A total of four samples were used in this analysis, three of which came from a rapidly progressing stage IV metastatic alveolar soft part sarcoma (ASPS) patient and a single sample from a patient-derived xenograft (PDX) model of ASPS treated with GCAR1. For the prior, the three samples included, one section from the untreated primary thigh tumor and two sections from GCAR1 treatment resistant lung needle biopsies. The PDX model was developed from a separate patient, and the model, but not the patient, was treated with GCAR1. In all cases, GCAR1 treatment was done at a dose of 1e6 GCAR1 CAR T-cells / kg. All of these samples were analyzed using Visium HD spatial transcriptomics.

first: Gill

last: Gurveer

first: Morrissy

last: Sorana


title: HIRA-mediated H3.3 deposition preserves hepatocyte cell identity during non-proliferative liver aging [RNA-Seq H33KO_3wks]

overall_design: RNA sequencing was used to study the transcriptomic changes of H3.3 KO livers in mice. We used male wildtype mice injected with AAV8 Tbg-Cre (control 1), H3f3afl/fl_H3f3bfl/fl C57BL/6 mice injected with AAV8 Tbg-MCS (control 2) and H3f3afl/fl_H3f3bfl/fl C57BL/6 mice injected with AAV8 Tbg-Cre (KO mice). Mice were injected at 6 months and then aged for an additional 3 weeks. Gene expression analysis was performed using data obtained from bulk-RNA seq of 10 samples (n = 3 for both of the controls, n = 4 for the KO).


title: HIRA-mediated H3.3 deposition preserves hepatocyte cell identity during non-proliferative liver aging [CosMx]

overall_design: CosMx Spatial Molecular Imager (SMI) was used to profile spatial gene expression in HIRA hepatocyte-specific knockout (KO) and control mouse livers. HIRA fl/fl C57BL/6 mice were injected with AAV8-TBG-Cre at 6 months of age to achieve hepatocyte-specific HIRA deletion (KO, n=3). Wildtype C57BL/6 mice injected with AAV8-TBG-Cre served as controls (n=3). Mice were aged for an additional 12 months post-injection and livers were collected at 18 months of age. Spatial transcriptomic profiling was performed on FFPE liver sections using the NanoString CosMx SMI platform (6 samples total; 3 biological replicates per group).

first: Christina

last: Shi


title: Comprehensive analysis of phenotypes and transcriptome characteristics reveal  important contribution of chronic stress and  immune response in the pathogenesis of acne vulgaris

city: Kunming

first: Shuyun

last: Yang

address: Dianmian Street Num374

department: Department of Dermatology

institute: The Second Affiliated Hospital of Kunming Medical University

summary: To investigate the molecular and physiological mechanisms by which chronic stress exacerbates acne pathogenesis. we used a murine model of stress-induced acne to perform comprehensive transcriptomic

overall_design: Comparative gene expression profiling analysis of RNA-seq data for Acne vs. Control, Acne_Pressure vs. Control, and Acne_Pressure vs. Acne

first: Xiaolan

last: Li


title: HIRA-mediated H3.3 deposition preserves hepatocyte cell identity during liver aging

type: Methylation profiling by genome tiling array

summary: Age-associated functional decline is partly driven by progressive chromatin degeneration. Maintenance of chromatin integrity preserves cell identity and promotes healthy aging, but through different mechanisms in proliferating and non-proliferating cells. However, specific mechanisms of chromatin maintenance and their compensatory capacity in proliferating and non-proliferating cells are undefined. The histone chaperone HIRA deposits the histone variant H3.3 in a DNA replication-independent manner, leading to its accumulation in aging, non-proliferating cells. Here, we show that hepatocyte-specific loss of HIRA causes loss of cell identity, metabolic dysfunction, and accelerated fibrotic pathology with age. Transcriptomic and epigenomic analyses indicate that HIRA-H3.3 preserves chromatin integrity and sustains transcription of highly expressed genes, including cell identity genes. Partial hepatectomy, associated with induced proliferation, restores identity of HIRA knockout livers with compensatory deposition of canonical histones H3.1/2. Together, these results demonstrate that HIRA-mediated H3.3 deposition is essential for safeguarding cell identity and tissue function during aging of non-proliferating cells, but this function can be rescued by tissue regeneration and associated cell proliferation.

overall_design: DNA methylation profiling was used to study the epigenetic changes of HIRA KO livers in mice. We used female wildtype C57BL/6 mice injected with AAV8-Tbg-Cre (control) and female HIRAfl/fl C57BL/6 mice injected with AAV8-Tbg-Cre (KO). Mice were injected at 6 months and then aged for an additional 12 months. DNA methylation analysis was performed using the Illumina Infinium Mouse Methylation BeadChip on 8 samples (4 independent replicates per experimental group).


title: HIRA-mediated H3.3 deposition preserves hepatocyte cell identity during non-proliferative liver aging

overall_design: CutnTag for H3.3 was performed to validate antibody specificity. We used a H3f3afl/fl; H3f3bfl/fl C57BL/6 mouse injected with AAV8-Tbg-Cre (H3.3 double KO) and a wildtype C57BL/6 control injected with AAV8-Tbg-Cre. Mice were injected when they were 6 months old and aged for an additional 3 weeks before tissue collection. Gene regulatory analysis was performed using data obtained from CutnTag of 2 samples (n = 1).


title: HIRA-mediated H3.3 deposition preserves hepatocyte cell identity during non-proliferative liver aging [CUT&Tag]

overall_design: CutnTag was used to study the epigenetic changes of HIRA KO livers in mice. We used female wildtype mice injected with AAV8 Tbg-Cre (control 1),  and HIRA fl/fl C57BL/6 mice injected with AAV8 Tbg-Cre (KO mice). Mice were injected at 6 months and then aged for an additional 12 months. Histone modification distribution analysis was performed for H3.3, H3K27ac, and H3K36me3 using data obtained from of 6 samples (3 independent replicates per experimental group).


title: C-type natriuretic peptide pre-maturation culture affects cytoplasmic maturation in bovine oocytes

city: Baton Rouge

first: Hao

last: Ming

state: Louisiana

address: 39 Forestry Ln

department: School of Animal Science

institute: Louisiana State University

summary: In vitro oocyte maturation (IVM) is a vital step for many assisted reproductive technologies including in vitro embryo production. Modulation of ribonucleic acid (RNA) transcripts, lipids, mitochondria, and other molecules during maturation influences development significantly. To improve current maturation systems, inclusion of pre-maturation culture has been investigated to delay resumption of meiosis and allow appropriate progression of cytoplasmic maturation using C-type natriuretic peptide (CNP). In this study, the transcriptome and molecular fingerprint of germinal vesicle, conventional IVM, and CNP-IVM oocytes were compared to identify transcripts and associated pathways altered by CNP treatment. There were similar pathways associated with upregulated transcripts in both types of mature oocytes involved with mitochondrial function and translation. Oocytes that were matured under conventional conditions contained transcripts associated with messenger RNA surveillance, Polycomb repressive complex, and reactive oxygen species. Oocytes matured with CNP treatment contained transcripts associated with cofactor biosynthesis, autophagy, apoptosis, arginine and proline metabolism, and mechanistic target of rapamycin (mTOR). Compared to immature oocytes, all mitochondrial-derived protein encoding genes were upregulated after conventional IVM but not after CNP-IVM. The molecular fingerprint of lipid droplets was also significantly altered in CNP-treated oocytes. Overall, results of these experiments have demonstrated measurable transcriptomic and molecular differences in CNP treated oocytes that may enhance oocyte and embryo quality.

overall_design: Cumulus-oocyte complexes were collected via transvaginal ultrasound guided follicle aspiration from donor cattle treated with follicle stimulating hormone (FSH) and either placed directly into maturation media (conventional IVM) or into a 6-hour pre-maturation culture containing 200 nM CNP followed by maturation (CNP-IVM). After 22 hours of maturation, oocytes were either fertilized and cultured to the blastocyst stage for quality assessment or stripped of cumulus cells and used for single oocyte analyses including RNA sequencing and molecular fingerprinting. Germinal vesicle stage oocytes were also denuded upon collection and compared with conventionally matured and CNP-treated metaphase II (MII) oocytes.

first: Emily

last: Girka

first: Zongliang

last: Jiang

first: Kenneth

last: Bondioli


title: Single-cell RNA sequencing of primary colorectal cancer, matched adjacent normal tissues, and paired liver metastases

city: Nanjing

first: Yueming

last: Sun

state: Prof.

address: 140 Hanzhong Road, Gulou District, Nanjing, Jiangsu

department: Department of General Surgery

institute: The First Affiliated Hospital of Nanjing Medical University

summary: This study will perform single-cell RNA sequencing on primary tumor tissues (T) and matched adjacent non-tumor tissues (N) from six colorectal cancer patients, along with paired liver metastatic lesions (M) from three of these patients, to construct an integrated single-cell atlas spanning primary tumors, adjacent normal tissues, and distant metastases. This multi-region, patient-matched design enables systematic dissection of cellular heterogeneity, dynamic immune responses, and key molecular mechanisms driving metastasis in colorectal cancer.

overall_design:
This study employs a multi-region, patient-matched design, performing single-cell RNA sequencing on primary tumor tissues (T) and matched adjacent non-tumor tissues (N) from six colorectal cancer patients. For three of these patients who also have liver metastases, paired metastatic liver lesions (M) are additionally included. All samples are processed immediately after surgical resection, dissociated into high-viability single-cell suspensions using a standardized enzymatic protocol, and subjected to single-cell capture and library preparation on the 10x Genomics platform.

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

first: Junwei

last: Tang

first: Yifei

last: Feng

first: Xiaowei

last: Wang

first: Sheng


title: “Zipper” Grammar of CTD Governs the Spatial Programing of the Transcription Cycle of RNA Polymerase II [TT-seq]

city: Austin

first: Yan Jessie

last: Zhang

state: TX

address: 100 E 24th St

department: Molecular Biosciences

institute: The University of Texas at Austin

summary: RNA polymerase II transcribes all protein-coding genes in eukaryotes, with its C-terminal domain (CTD) acting as a regulatory platform throughout the transcription cycle. Despite its simple sequence, the conserved heptad repeats encode a regulatory grammar critical for transcription initiation, elongation, and termination. Using structural analysis, single-molecule imaging, and synthetic CTD-engineered Pol II constructs, we defined the minimal sequence requirements across transcriptional stages. We show that while SP motifs are essential phosphorylation sites, flanking residues tolerate variability. In contrast, the periodic positioning of tyrosine residues is indispensable for pre-initiation complex (PIC) assembly through interactions with the Mediator, which are disrupted upon Ser5 phosphorylation triggering promoter escape. This supports a “zipper” model where sequential tyrosine engagement stabilizes PIC assembly and progressive phosphorylation propels Pol II escape. Site-specific Ser2/5 phosphorylation orchestrates 3’-end processing factor recruitment. These findings redefine the functional grammar of the Pol II CTD and explain how a low-complexity sequence achieves regulatory specificity.

overall_design: TT-seq profiling of HEK293 degron cells that were tranfected with an RPB1 with a 26X wildtype CTD or a 26X Y1T and T4Y CTD

first: Qian

first: Haley

last: Hardtke


title: Cell type-specific ablation of siRNAs by Arabidopsis RTL1 reveals a role of phloem companion cells in systemic post-transcriptional gene silencing

city: Versailles

first: Hervé

last: Vaucheret

country: France

institute: INRAE

type: Non-coding RNA profiling by high throughput sequencing

summary: Small RNAs regulate numerous biological processes, including intercellular communication and inter-kingdom relationships, yet their tissue-specific functions remain poorly understood. Dissecting these roles requires tools that selectively eliminate small RNAs in defined cell types. Virus-encoded proteins such as P19 have been widely used to neutralize small RNAs owing to their sequestration capacities. However, their restricted binding spectrum to 21-22-nt small RNAs and lack of discrimination between miRNAs and siRNAs limit their utility. Here we show that constitutive expression of RNASE THREE-LIKE1 (RTL1) in Arabidopsis thaliana, previously shown to degrade siRNA precursors but not miRNA precursors, also degrades siRNA duplexes, irrespective of their size, sequence, or sub-cellular localization. Grafting experiments combined with small RNA sequencing revealed that shoot-derived siRNAs accumulate in dcl2 dcl3 dcl4 mutant roots, but are eliminated upon ectopic RTL1 expression, demonstrating that RTL1 efficiently degrades mobile siRNAs in vivo. Because the Arabidopsis endogenous RTL1 is epigenetically silenced in wild-type plants, its ectopic expression under cell type specific promoters enables targeted depletion of siRNAs in selected tissues. As a proof of concept, we show that expression of RTL1 in phloem companion cells delayed, but did not abolish, systemic post-transcriptional gene silencing (PTGS), thus indicating that mobile siRNAs transit through these cells while also using alternative routes. Together, these results establish RTL1 as a powerful tool to dissect siRNA mobility pathways and tissue-specific functions.

overall_design: Small RNA-seq profiling of siRNA moving in Arabidopsis thaliana from scions to rootstocks in a grafting experiment were WT scions expressing a silent p35S:GUS transgene (L1 line) were grafted onto rootstocks of the dcl2 dcl3 dcl4 triple mutant or of the dcl2 dcl3 dcl4 triple mutant ectopically expressing the RTL1 gene under the control of the UBQ10 promoter.

first: Martin

last: Lacroix

first: Agnès

last: Yu


title: MELK inhibition promotes megakaryopoiesis and platelet recovery in immune thrombocytopenia through ERK–cofilin signaling

first: Jiaqian

last: Qi

state: Jiangsu

address: No.188 Shizi Road

institute: The First Affiliated Hospital of Soochow University

summary: Maternal embryonic leucine zipper kinase (MELK) has been implicated in cell-cycle regulation, yet its transcriptional programs in megakaryocytic cells remain incompletely defined. To profile MELK-dependent gene expression changes, we performed bulk RNA-seq on the human megakaryoblastic cell line MEG-01 transduced with a control pLKO.1 vector or a MELK-targeting shRNA (sh2-MELK), with three independent biological replicates per condition. Paired-end sequencing reads (raw FASTQ files) and processed gene-level expression quantifications are provided to support differential expression and pathway analyses of MELK knockdown in megakaryocytic cells.

overall_design: This study uses a two-group comparative RNA-seq design to evaluate transcriptional changes following MELK knockdown in MEG-01 cells. MEG-01 cells were transduced with either control pLKO.1 or MELK-targeting shRNA (sh2-MELK). Three independent biological replicates were generated for each condition (n = 3 per group). Total RNA was extracted from each replicate, sequencing libraries were prepared, and samples were sequenced in paired-end mode. Differential gene expression was assessed by comparing sh2-MELK versus control to identify MELK-associated pathways in megakaryocytic cells.

first: Xueqian

first: Depei

last: Wu

first: Yue

last: Han


title: Mitochondrial L-2-hydroxyglutarate is a physiologic signaling metabolite [CUT&Run]

city: Chicago

first: Yuki

last: Aoi

address: 303 E Superior St. SQBRC 7-300

department: Biochemistry & Molecular Genetics

institute: Northwestern University

summary: L-2-hydroxyglutarate (L-2-HG) is a low-abundance metabolite in mammals due to the mitochondrial enzyme L-2-HG dehydrogenase (L2HGDH), which oxidizes L-2-HG to 2-oxoglutarate (2-OG) to prevent its accumulation1. In humans lacking L2HGDH activity, L-2-HG builds up, leading to L-2-HG aciduria, a rare childhood neurometabolic disorder2. Thus, L-2-HG is often classified as a toxic metabolite2–5. Furthermore, L-2-HG is produced in response to hypoxia, acidic pH, and electron transport chain (ETC) impairment6–10. However, whether L-2-HG has a physiological function is unclear. Here, we investigated whether L-2-HG qualifies as a physiological signaling metabolite by testing three criteria: regulated levels, defined molecular targets, and a measurable physiological function. We report that elevated mitochondrial NADH/NAD⁺ ratio drives malate dehydrogenase 2 (MDH2) to reduce 2-oxoglutarate (2-OG) into L-2-HG, while L-2-hydroxyglutarate dehydrogenase (L2HGDH) reversibly oxidizes L-2-HG to 2-OG within the mitochondrial matrix, independently of the ETC. Proteome integral solubility alteration (PISA) assays revealed KDM4 family of H3K9 demethylases as L-2-HG-responsive targets. Consistent with the targets, elevated L-2-HG repressed nascent transcription of specific gene sets and promoted accumulation of the repressive histone mark H3K9me3 at those loci. In vivo, early embryonic L2HGDH overexpression lowered L-2-HG broadly, reduced postnatal growth, and caused postnatal mortality. Despite systemic lowering, kidneys displayed a unique vulnerability, developing fibrosis and functional decline. Mechanistically, L-2-HG depletion in the postnatal kidney resulted in a specific loss of H3K9me3 at L1MdTf retrotransposons, leading to their de-repression and coinciding with activation of the integrated stress response and inflammation pathways. Our findings establish mitochondrial L-2-HG as a physiological signaling metabolite that couples mitochondrial redox to chromatin regulation and is essential for postnatal kidney development and survival. These findings suggest that metabolites previously regarded as toxic may also serve critical physiological functions.

overall_design: To study the impact of L2HGDH overexpression on epigenetic modifications and transcription in mouse kidney, CUT&RUN assays for H3K9me3 and RNA Polymerase II were performed.

first: Ram

last: Chakrabarty

first: Navdeep

last: Chandel


title: HDAC inhibition unlocks tumor plasticity and enhances immunotherapy response in Myc- Driven Small Cell Lung Cancer [ATAC-Seq]

first: Linying

last: Zhu

state: Maryland

address: 20 Penn St

department: School of Medicine

institute: University of Maryland, Baltimore

summary: Small Cell Lung Cancer (SCLC) is a highly aggressive malignancy, accounting for approximately 15% of all lung cancer cases. Characterized by low immunogenicity, SCLC may utilize epigenetic mechanisms to evade immune detection. Here, we demonstrate that entinostat, a class I histone deacetylase inhibitor (HDACi) upregulates immune-related genes in human SCLC cells. In vivo, we confirmed entinostat treatment increased expression of immunecheckpoint ligands and antigen presentation machinery in Myc-driven tumors in a Rb1/Trp53/MycT58A (RPM) SCLC mouse model, while shifting tumors from a neuroendocrine(NE)-high to a NE-low phenotype. Notably, combining entinostat with anti-PD-1 immunotherapy significantly enhances T-cell infiltration, suppresses tumor growth, and prolongs survival in RPM allograft models. These findings underscore the potential of entinostat to reprogram the immunological landscape and NE status of SCLC, enhance immune checkpoint blockade efficacy, and improve therapeutic outcomes.

overall_design: ATAC-seq profiling of Rb1/Trp53/MycT58A (RPM) induced lung tunor and normal lung tissue. And the induced tumors treated with vehicle, IgG, anti-PD-1 antibody, entinostat and the combination of anti-PD-1 and entinostat.

first: Azam

last: Ghafoor

first: Lorinc

last: Prongor

first: Haoxuan

last: Ying


title: Elevated Labile Iron Contributes to Membrane Repair Deficits in FSHD

city: Washington,DC

first: Chen

last: Yi Wen

state: Washington,DC,

address: Sheikh Zayed Campus,111 Michigan Avenue,

department: Center for Precision Medicine and Genomics Research

institute: Childrens National Hospital

summary: Facioscapulohumeral muscular dystrophy (FSHD) is an autosomal dominant muscle disease caused by expression of the transcription factor DUX4, which we previously found to be associated with plasma membrane (PM) repair deficits. However, the molecular mechanisms associated with poor membrane repair are not clear. Here, we show PM injury itself results in mild but significant increase in DUX4 mRNA expression, as well dysregulation of genes enriched in pathways critical for membrane repair. We find these dysregulated genes to be sensitive classifiers of FSHD status in human muscle biopsies, indicating their potential utility as novel biomarkers. In addition, FSHD myoblasts show dysregulation of genes in the ferroptosis cell death signaling pathway in response to membrane damage. Subsequent experiments indicated FSHD myoblasts present with hallmark features of ferroptotic stress, including elevated labile ferrous (Fe2+) iron, and lipid peroxidation at baseline and post-injury. Furthermore, we demonstrate that the expression of ferroptosis biomarkers is elevated in FSHD muscle biopsies, which is predictive of the presence of inflammation and positively correlates with the degree of fatty infiltration in FSHD skeletal muscle.  Increasing labile iron and lipid peroxidation worsens membrane repair in FSHD myoblasts, while treatment with the iron chelator 2,2′-Bipyridyl and ferroptosis inhibitor ferrostatin-1 improve repair. These data are the first to identify signs of ferroptotic stress in FSHD myoblasts, and demonstrate the potential therapeutic benefit of iron chelation and ferroptosis inhibition to improve membrane repair capacity in FSHD.

overall_design:
To investigate the transcriptional response to plasma membrane injury, we  performed single cell RNAseq of immortalized human myoblast cell lines from 2 donors with FSHD, and 2 healthy siblings
2 myoblast cell lines from individuals with FSHD and two myoblast cell lines from Healthy individuals were collected prioor to membrane injury (baseline), and at 6-hours post-injury. A total of  8 samples were obtained

first: Adam

last: Bittel

first: Surajit

last: Bhattacharya

first: Lulya

last: Okubamariam

first: Yi-Wen

last: Chen


title: “Zipper” Grammar of CTD Governs the Spatial Programing of the Transcription Cycle of RNA Polymerase II [ChIP-Seq]

first: Yan Jessie

last: Zhang

overall_design: ChIP-seq of GFP-tagged RPB1 with engineered C-terminal domain (CTD) in HEK293T cells.

first: Qian

first: Haley

last: Hardtke


title: AATF supports proliferation of glioblastoma cells by sustaining mitochondrial respiration through an NRF-1-dependent mechanism [ChIP-seq]

city: Rome

first: Stefano

last: Di Giovenale

address: Via Elio Chianesi,  53

country: Italy

institute: IRCCS National Cancer Institute Regina Elena

summary: The ability of cancer cells to promote cellular proliferation by preferentially using glycolysis as primary source of energy, has long been considered a hallmark of tumour metabolism. However, emerging evidence suggests a more complex situation with many tumours exhibiting a pronounced dependence on mitochondrial respiration through oxidative phosphorylation (OXPHOS) for their development and maintenance. In line with this, numerous studies have reported an upregulation of mitochondrial genes and OXPHOS components across multiple cancer types. Glioblastoma (GBM) is the most frequent and malignant brain tumour in adults, characterized by rapid proliferation, resistance to therapy and ability to recur. In addition to a profound genetic and molecular heterogeneity, GBM also exhibits strong metabolic heterogeneity with different grade of dependence from mitochondrial activity. Notably, the transcription factor Nuclear Respiratory Factor 1 (NRF-1), a key regulator of OXPHOS gene expression and mitochondrial functions, has recently been linked to GBM progression and poor prognosis. Che-1/Apoptosis Antagonizing Transcription Factor (AATF) is a transcriptional regulator with a crucial role in several cancer types where it contributes to tumorigenesis by promoting cell cycle arrest and apoptosis, as well as resistance to therapy. Here, we show that AATF expression is correlated with clinical outcome in GBM patients. Moreover, we demonstrate that its depletion leads to cell cycle arrest, impaired mitochondrial respiration, and disrupted mitochondrial architecture in GBM cells. In particular, we provide evidence that AATF interacts with NRF-1 and it is essential for NRF-1-mediated transcription of the OXPHOS genes by affecting RNA polymerase II recruitment and chromatin structure. Overall, our findings highlight a previously unrecognized role of AATF in GBM proliferation and mitochondrial metabolism and suggest it as a potential target for therapeutic intervention.

overall_design: ChIPseq experiment targeting NRF1 in U138 cell in control and interfired AATF.

first: Cristina

last: Sorino

first: Italia

last: Falcone

first: Francesca Romana

last: Auciello

first: Claudio

last: Pulito

first: Federica

last: Lo Sardo

last: Scalera

first: Francesca

last: De Nicola

first: Valeria

last: Catena

first: Ludovica

last: Ciuffreda

first: Brindusa Ana Maria

last: Arteni

last: Giuliani

first: Bruno

last: Amadio

first: Giovanni

last: Blandino

first: Maurizio

last: Fanciulli

first: Simona

last: Iezzi


title: HDAC inhibition unlocks tumor plasticity and enhances immunotherapy response in Myc- Driven Small Cell Lung Cancer

first: Linying

last: Zhu

overall_design: RNA-seq profiling of Rb1/Trp53/MycT58A (RPM) induced lung tumor and normal lung tissue.

first: Azam

last: Ghafoor

first: Lorinc

last: Prongor

first: Haoxuan

last: Ying


title: HDAC inhibition unlocks tumor plasticity and enhances immunotherapy response in Myc- Driven Small Cell Lung Cancer.

overall_design: RNA-seq profiling of Rb1/Trp53/MycT58A (RPM) induced lung tunor and normal lung tissue (GSE309192). The induced tumors treated with vehicle, IgG, anti-PD-1 antibody, entinostat and the combination of anti-PD-1 and entinostat.


title: MK2/p38/p53 negatively regulate basal IL-1β and noncanonical NF-κB signaling

city: Hannover

first: Sarah

middle: Maria

last: Herr

address: Carl-Neuberg-Str. 1

country: Germany

institute: Medizinische Hochschule Hannover

summary: Interleukin (IL)-1β is an endogenous pro-inflammatory pyrogen that affects both the innate and adaptive immune systems. Uncontrolled release of IL-1β is associated with sterile inflammatory diseases, such as arthritis, gout, and atherosclerosis as well as the development of tumors. In an initial investigation of the role of the MAPKAP kinase MK2 in IL-1β processing in macrophages, we found that Il1b mRNA and IL-1β protein levels were increased in unstimulated MK2-knockout (KO) macrophages. Additionally, we observed elevated basal IL-1β concentrations in the serum of MK2/3 double KO mice. Further analysis revealed an activation of the noncanonical NF-κB pathway in the absence of MK2 or its interacting activator p38α. Rescue by overexpression of MK2, its kinase-inactive mutant MK2K79R or p38α reduced activation of the noncanonical NF-κB pathway components as well as Il1b mRNA expression. Interestingly, the basal protein level of the tumor suppressor p53 is reduced in the absence of MK2 or p38α.We demonstrate that p53 interacts with mitochondrial caspase-3, which cleaves RelB and therefore inhibits the noncanonical NF-κB pathway, affecting subsequent Il1b and p53 expression. These results not only explain the increased basal levels in MK2-knockout macrophages, but also uncover a new autoregulatory mechanism of p53 expression. Additionally, they reveal a new mechanism that contributes to the long-discussed link between cancer and inflammation, in which the tumor suppressor p53 represses cytokine expression.

overall_design: RNA-seq profiling of untreated or IL-1α treated immortalized MK2-KO macrophages rescued with either MK2 (iMK2-KO+MK2) or an empty vector (iMK2-KO+GFP)

first: Sarah Maria

first: Matthias

last: Gaestel


title: Circulating cell populations as response predictors and targets to improve immunotherapy in metastatic lung cancer

city: Sao Paulo

first: Kenneth

last: Gollob

address: Albert Einstein Ave, 627/701

country: Brazil

institute: Hospital Israelita Albert Einstein

summary: The effectiveness of immunotherapy varies among patients with advanced non-small cell lung cancer (NSCLC). Here we investigated the baseline immune status in stage IV NSCLC patients treated with anti-PD-1 plus chemotherapy to understand the immune mechanisms and unveil systemic markers associated with treatment response. Responders had elevated frequencies of circulating T cells expressing CD69, TCF-1 and CXCR-3. In contrast, non-responders presented increased frequencies of CTLA-4, CD161 and IL-10 expressing CD4+ and CD8+ T cells. These systemic T cell immune profiles were mirrored in the tumor microenvironment of an independent cohort. Concurrent CTLA-4 and PD-1 blockade was able to reactivate an anti-tumor profile in T cells from non-responder patients, emphasizing the pivotal role of CTLA-4 in contributing to an immunosuppressive environment that hinders effective treatment in NSCLC. This work supports the implementation of personalized immunotherapies based on systemic immune biomarkers, offering a promising approach to enhance treatment outcomes in advanced NSCLC.

overall_design: We included patients of both sexes aged 18 years or over, histologically confirmed diagnosed of stage IV NSCLC that were treated with the combination of pembrolizumab + chemotherapy. Patients with disease progression were included in the “non-responder” group and patients with complete or partial response or stable disease were included in the “responder” group. Responses were assessed 9 weeks or 6 months after starting treatment. Peripheral blood was collected in EDTA tubes at the time of diagnosis, before starting the treatment, and PBMC were purified using a Ficoll gradient and cryopreserved in FBS / DMSO.

first: Amanda

last: Figueiredo


title: Discovery of kidney disease targets using multimodal human podocyte injury models

city: Durham

last: Barreto

address: 1427 Fitzpatrick Center

institute: Duke University

summary: Podocyte injury is a hallmark of chronic kidney disease (CKD), but whether different injury signals perturb unified or distinct molecular targets remains unclear. Using human induced pluripotent stem cell (hiPSC)-derived podocytes, we modeled cellular injury via exposure to diabetic, inflammatory, chemical toxin, biomechanical, and infectious stressors. Transcriptomic analysis revealed both shared and unique changes in gene expression across injury modes. While drug-induced injuries triggered broader transcriptional responses, conserved pathways related to lysosome function, RNA metabolism, and immune activation were identified across models. Importantly, we discovered that NEU1, CD82, ABI3BP, and ADAM17 as targets of human podocyte injury. We confirmed enrichment of these targets in kidney disease patient biopsies, underscoring their in vivo relevance and potential as therapeutic targets. These findings highlight the value of human-relevant experimental models and provide insight into podocyte injury responses, offering a framework for future precision medicine approaches.

overall_design: Human iPSC lines were cultured under standard conditions and used for podocyte differentiation. hiPSCs were grown on a Matrigel matrix in mTeSR1 medium and induced into podocytes through a stepwise differentiation protocol involving mesoderm induction followed by intermediate mesoderm and podocyte induction. After differentiation, the podocytes were exposed to various injury conditions, including high glucose, high glucose with inflammatory cytokines, Adriamycin, cytochalasin D, and lipopolysaccharide. RNA was isolated from three biological replicates per condition, and RNA quality was assessed. RNA sequencing was performed per sample for differential gene expression analysis.

first: Bowen

last: Jiang

first: Morgan

last: Burt

first: Nikolaos

last: Dimitrakakis

first: Samira

last: Musah


title: Increased constitutive interferon-β levels and altered CD4 T cell homeostasis induced by expression of a viral glycoprotein

city: Freiburg

first: Thomas

last: Boehm

address: Stuebeweg 51

department: Developmental Immunology

institute: Max Planck Institute for Immunobiology and Epigenetics

summary: Besides the strong type I interferon (IFN-I) response induced by infections, IFN-I is also produced constitutively at lower levels. Constitutive IFN-I production is often regulated by resident microbiota and is essential for induction of efficient immune responses. Here, we demonstrate in transgenic (tg) mice that expression of a single viral envelope gene, the glycoprotein (GP) of lymphocytic choriomeningitis virus (LCMV), increased constitutive IFN-β levels and induced numerous IFN-stimulated genes (ISGs). In addition, self-antigen-driven CD44highCD62Llow memory-phenotype and regulatory CD4 T cell populations were enlarged in these LCMV GP-tg mice but bona fide antigen-specific memory CD4 T cells were unchanged. In conclusion, our study shows that increased constitutive IFN-β levels induced by expression of a single viral envelope gene and accompanied by upregulated ISGs significantly altered CD4 T cell homeostasis. Furthermore, our data imply that a pathway capable of upregulating constitutive IFN-β levels can sense LCMV GP expression in vivo.

overall_design: Total RNA was extracted from splenic lymphocytes (CD4+, CD8+, CD19+) of MHCI.GP+ and MHCI.GP- mice and subjected to bulk RNAseq.

first: Hanspeter

last: Pircher

first: Oliver

last: Thomas

first: Anne

last: Häfke

first: Annette

last: Oxenius


title: Tumor suppressor genotype influences the extent and mode of immunosurveillance in lung cancer

city: Stanford

first: Haiqing

last: Xu

address: 327 Campus Drive

department: Biology

institute: Stanford University

summary: The impact of cancer driving mutations in regulating immunosurveillance throughout tumor development remains poorly understood. To better understand the contribution of tumor genotype to immunosurveillance, We generated and validated lentiviral vectors that establish an epi-allelic series of increasingly immunogenic neoantigens, integrating them with autochthonous Cre-regulated cancer models, CRISPR/Cas9-mediated somatic genome editing, and tumor barcoding into a single system. Using this sytem, we show that in the context of KRAS-driven lung cancer and strong neoantigen expression, tumor suppressor genotype dictates the degree of immune cell recruitment, positive selection of tumors with neoantigen silencing, and tumor outgrowth.

sample_organism:
Escherichia coli
Mus musculus

platform_organism:
Escherichia coli
Mus musculus

overall_design: We use Tuba-seq to precisely quantify the effects of tumor suppressor genotype under varying levels of neoantigen expression in the context of KRAS-driven lung cancer.

first: Keren

last: Adler

first: Amy

last: Gladstein

first: Valerie

last: Irizarry-Negron

first: Maggie

last: Robertson

first: Katherine

last: Doerig

first: Dmitri

last: Petrov

first: Monte

last: Winslow

first: David

last: Feldser


title: Effects of Human Plasma-Like Medium on hPSC-derived cardiomyocytes in vitro

city: Madison

first: Xiaotian

last: Zhang

state: WI

address: 1415 Engineering Hall

department: Chemical and Biological Engineering

institute: University of Wisconsin-Madison

summary: Maturing hPSC-CMs in vitro is critical for advancing drug discovery and cardiotoxicity screening. However, widely used basal medium such as RPMI contain non-physiological compositions that limit their relevance to human cardiac physiology. In this study we introduced Human Plasma-Like Medium (HPLM) as a better basal medium supplemented with B27, a physiologically tailored alternative designed to systematically replicate the salt concentrations and polar metabolite profiles of adult human plasma. Starting with Day16 hPSC-CMs, we cultured cells for two weeks in HPLM/B27 or control media (standard and low-glucose [5 mM] RPMI/B27) and assessed maturation outcomes at Day30. Compared to standard and low-glucose RPMI/B27, HPLM/B27 significantly enhanced hPSC-CM maturity across transcriptomic, structural, functional, and metabolic aspects. These improvements included accomplished myosin heavy chain isoform switching (MYH6 to MYH7), accelerated ventricular-specific myosin light chain isoform switching (MLC2a to MLC2v), elongated sarcomeres (~2.07 µm), enhanced calcium transient kinetics, and coordinated activation of oxidative and glycolytic metabolism. Collectively, these findings establish HPLM as a better basal medium than low glucose RPMI for driving cardiomyocyte maturation in vitro.

overall_design: High purity of hPSC-CMs (>80%, WTC-11 line) generated by GiWi protocol was randomized into three different media conditions on Day16, including standard RPMI/B27, low-glucose (5mM) RPMI/B27, and HPLM/B27 (n=3 for each media condition with each sample comes from 1 well of a 12-well plates). Those three experimental groups were maintained in each medium for additional two week with media change every 3 days. Day 16 hPSC-CMs cultured in standard RPMI/B27 was collected on Day16 with hPSC-CMs in other three different media conditions were collected on Day30.

first: Aaron

last: Simmons

first: Sean

last: Palecek


title: Single-nucleus RNA sequencing of human subcutaneous adipose tissue biopsies from 59 Finnish participants.

city: Los Angeles

first: Kyla

last: Gelev

email: kylagelev@gmail.com, kzg@g.ucla.edu

address: 695 Charles E. Young Drive South

institute: University of California, Los Angeles

summary: We performed single-nucleus RNA-sequencing on subcutaneous adipose tissue biopsies from 59 Finnish donors from the Kuopio OBesity Surgery study (KOBS) cohort.

overall_design: Frozen subcutaneous adipose tissue (SAT) biopsies (n=59) were pooled and batched into 8 batches with 7-8 SAT samples per batch. We randomly assigned samples to batches to minimize correlations between phenotypes of interest and batch. We took 100mg from each sample prior to pooling the samples together for nuclei isolation and applied the 10x Genomics nuclei isolation protocol. Library construction was performed using the Single Cell 3’ v3.1 chemistry. All libraries were equally distributed across two lanes and sequenced on the Novaseq X Plus 25B to target 70,000 reads per nucleus.

middle: Z

first: Päivi

last: Pajukanta

first: Jussi

last: Pihlajamäki


title: AATF supports proliferation of glioblastoma cells by sustaining mitochondrial respiration through an NRF-1-dependent mechanism [RNA-seq]

first: Stefano

last: Di Giovenale

overall_design: RNAseq experiment of U138 cell lines with control and interfirence of AATF conducted in triplicates.

first: Cristina

last: Sorino

first: Italia

last: Falcone

first: Francesca Romana

last: Auciello

first: Claudio

last: Pulito

first: Federica

last: Lo Sardo

last: Scalera

first: Francesca

last: De Nicola

first: Valeria

last: Catena

first: Ludovica

last: Ciuffreda

first: Brindusa Ana Maria

last: Arteni

last: Giuliani

first: Bruno

last: Amadio

first: Giovanni

last: Blandino

first: Maurizio

last: Fanciulli

first: Simona

last: Iezzi


title: Preservation of Myogenic Tone in Kidney Microvasculature of Dahl Salt-Sensitive Rats. Role of C-C motif chemokine ligand 2

city: Birmingham

first: Kelly

middle: Anne

last: Hyndman

state: Al

address: 1918 University BLVD

institute: University of Alabama at Birmingham

summary: Kidney injury mechanisms that develop in salt-sensitive (SS) hypertension are incompletely understood.  Using the Dahl SS rat, which is a prominent model of salt-sensitive hypertension and associated kidney injury, loss of function of the pro-inflammatory transcription factor, ETS-1, corrects kidney microvascular autoregulatory dysfunction and injury.  C-C motif chemokine ligand 2 (CCL2) is a direct target of ETS-1, so SS rats lacking Ccl2  (termed (SSCcl2 -/-) were therefore examined. Unlike SS rats, SSCcl2 -/- rats did not develop salt-sensitive hypertension, loss of autoregulation of afferent arterioles, or kidney injury.  cDNA libraries from renal microvasculature of these rats was sequenced.

overall_design: Male Dahl salt sensitive (SS) rats and SS Ccl2 null (SS Ccl2-/-) rats were placed on 0.3% NaCl or 4.0% NaCl diets for 2 weeks.  Kidneys were excised and the microvasculature dissected. Total RNA was extracted from the kidney microvasculature and cDNA libraries generated.

first: Wenguang

last: Feng

first: Zhengrong

last: Guan

first: Wei-Zhong

last: Ying

first: Conrad

first: Kelly A.

first: Paul W.

last: Sanders


title: Mosquito- and tick-borne orthoflavivirus infections dysregulate microcephaly-associated genes in human neural progenitor cells

city: Atlanta

first: Emilio

middle: E.

last: Espínola

state: Georgia

address: 145 Piedmont Ave SE

institute: Georgia State University

summary: Human neural progenitor cell (hNPC) transcriptomic responses to infections with Zika virus (ZIKV), Powassan virus (POWV), or two strains of West Nile virus were analyzed at three times after infection in a single batch of hNPCs differentiated from induced pluripotent stem cells. The WNV infections displayed the fastest replication kinetics but the POWV infection produced the highest viral yield. All four infections upregulated common and unique innate immune and antiviral response genes by 12 hpi and induced upregulation of stress pathway genes by 24 hpi. The WNV infections downregulated cell cycle genes earlier than the ZIKV or POWV infections. Transcription factors regulating gene expression changes were predicted. All four orthoflaviviruses dysregulated the expression of 262 microcephaly-associated genes, with virus-specific differences in the extent of dysregulation. The data suggest that fetal hNPC infection in utero by each of the four orthoflaviviruses studied could induce neurodevelopmental disorders.

overall_design: RNA-seq profiling of human neural progenitor cells (hNPCs) infected with Zika virus (ZIKV) PRVABC59, Powassan virus (POWV) LB, West Nile virus (WNV) MAD78, or WNV NY99, at 12, 24, and 48 hours post-infection.


title: CUX1 Manifests Species-Dependent Regulation in Controlling Adipocyte Differentiation [ChIP-Seq]

city: Philadelphia

first: Ryan

middle: Patrick

last: Calhoun

address: 3400 Civic Center Boulevard

institute: University of Pennsylvania

summary: Adipocyte differentiation, also known as adipogenesis, is a crucial process for maintaining the adipocyte pool and plays a key role in insulin sensitivity and systemic metabolism. Adipogenesis is governed by master transcription factors, such as C/EBPβ. However, there remains an unmet need to identify novel transcriptional factors within the C/EBPβ regulatory network in the differentiation of primary human adipose-derived precursor cells (hAPCs). Chromatin immunoprecipitation and selective isolation of chromatin-associated proteins (ChIP-SICAP) and CRISPR/cas9 knockout (KO) screening approach was applied in hAPCs. RNA-seq and ChIP-seq were also utilized for pathway and gene enrichment analyses. We also constructed a APCs-specific KO mouse model. Deletion of CUX1 in hAPCs impairs adipogenesis, whereas CUX1 overexpression (OE) increases human adipogenesis. RNA-seq and ChIP-seq analyses reveal that CUX1 promotes the expression of key adipogenic genes, including PPARG, a master regulator of adipocyte fate determination. Interestingly, Cux1 deletion enhances, while CUX1-OE suppresses adipogenesis in mouse APCs (mAPCs). ChIP-seq analysis of CUX1 in mAPCs further supports its contrasting role in mice. Additionally, in vivo tracing of mAPCs shows that Cux1 deletion promotes adipocyte differentiation. Collectively, these findings highlight that CUX1 exerts species-specific regulation of adipogenesis, acting as a differential regulator of fat development in humans and mice.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

first: Chen

last: Yang

first: Liu

last: Lin

first: Calhoun

last: Ryan

first: Cheng

last: Lan

first: Steger

last: David

first: Seale

last: Patrick


title: Campylobacter jejuni regulates cell cycle progression to potentiate host cell invasion

city: PULLMAN

first: Eric

middle: Aaron

last: Shelden

state: WA

address: 1770 NE Stadium Way

department: School of Molecular Biosciences

institute: Washington State University

summary: To assess the effects of C. jejuni infection on cell cycle progression, INT407 cells were synchronized, then infected with either wild-type C. jejuni, or a mutant lacking Ciac. Uninfected cells were used as controls. Cells were harvested and processed for single cell RNA sequencing analysis 20 hours after infection, and differentially expressed genes analyzed to determine effects of infection on gene expression and cell signaling.

overall_design: INT407 cells were cultured in EMEM 10%-FBS in 100 x 20 mm tissue culture dishes overnight and then treated with RO-3306 for 24 hours. Following incubation, the cells were rinsed with MEM containing 1% FBS. The cells were infected with the C. jejuni 81-176 wild-type strain and ∆ciaC mutant for 20 hours. Cells were also incubated with EMEM 1%-FBS (control). The samples were processed as outlined in Sample Preparation Demonstrated Protocol, as instructed (10xgenomics.com).

first: Prabhat

last: Talukdar

first: Megan

middle: C

last: Dines

first: Amruta

middle: S

last: Kale

last: Driskell

first: Lisa

last: Gloss

first: Michael

last: Konkel


title: Met32 governs transcriptional control of sulfur metabolic flexibility and resistance to reactive sulfur species in the human fungal pathogen Candida albicans

city: Montreal

first: Adnane

last: Sellam

address: 5000, rue Bélanger

institute: Universite de Montreal

summary: Although considerable advances were made in understanding the metabolic machinery that enable bacteria to utilize alternative sulfur sources, many aspects of this process remain understudied in fungi. To explore the genetic circuit by which the highly prevalent opportunistic yeast Candida albicans control sulfur utilization, we characterized the transcriptional atlas associated with sulfur starvation in this yeast. We identified many putative desulfonation enzymes that were differentially transcribed and showed that Jlp12, an α-ketoglutarate dioxygenase, was critical for the utilization of different sulfur sources found in many niches of the human host. We also uncovered that the Zinc-finger transcription factor Met32 acts as a master transcriptional regulator, that modulates genes of sulfur utilization including Jpl12. Importantly, we found that the C. albicans Met32 functions exclusively in the control of sulfur utilization genes, while it governs methionine biosynthesis in the S. cerevisiae lineage. This work also identified Seo13 as the first major facilitator superfamily transporter in fungi that transports the alternative sulfur source glutathione, under the direct control of Met32. Furthermore, we showed that Met32 modulates C. albicans tolerance sulfite excess by tuning the basal transcriptional level of the superoxide dismutase Sod1. This underscores the dual role of Met32 in the breakdown of sulfur-containing metabolites and the neutralization of the resulting reactive sulfur species (RSS). Our study delineates a new mechanism by which fungal pathogens utilize sulfur sources and neutralize RSS, and underscores its importance for fungal fitness in vivo.

overall_design: In this study, we identified transcripts modulated under sulfur starvation by comparing the transcriptome of cells growing in sulfur-lacking medium (SLD) to those growing in sulfur-replete medium (SLD supplemented with either taurine, mixture of methionine/cysteine, ammonium sulfate or glutathione) in the pathogenic yeast Candida albicans. We also identified transcripts that were modulated in the mutant of the transcription factor Met32 (met32). The transcriptional profile of met32 grown in sulfur-free (SLD) or sulfur-supplemented medium (SLD + taurine or SLD + glutathione) was compared to that of the wild-type (WT) strain grown under similar conditions.

first: Anagha C

last: Menon

first: Faiza

last: Tebbji

first: Antony T

last: Vincent


title: CAF drives spheroid formation and pleural dissemination in NSCLC

city: Okayama

first: Shuta

last: Tomida

state: Okayama

address: 2-5-1 Shikata-cho, Kita-ku

department: Center for Comprehensive Genomic Medicine

country: Japan

institute: Okayama University Hospital

summary: Pleural dissemination in non-small cell lung cancer (NSCLC) refers to the direct spread of cancer cells from the primary tumor site to the extrapulmonary thoracic cavity, where they colonize the pleura. This study aimed to investigate the impact of cancer-associated fibroblasts (CAFs) on the development of pleural dissemination in NSCLC. Through in vitro co-culture and in vivo pleural dissemination models, we identified that CAFs promote cancer cell migration, anchorage-independent growth, adhesion, invasion, and spheroid formation of cancer cells, leading to the development of pleural dissemination. RNA sequencing analysis revealed that CAFs upregulated the expression of Cellular Communication Network Factor 1 (CCN1), a matricellular protein, in cancer cells.

overall_design: RNA-seq profiling of wildtype PC9, H1299, H1975, HCC827 NSCLC cell lines and their co-culture cell lines with CAFs

first: Naoki

last: Matsuda

first: Ken

last: Suzawa

first: Yin

last: Min Thu