title: Long-Read Sequencing and Proteomics Reveal Blood Transcriptome and Protein Expression Profiles in Multiple Primary Lung Cancers

city: Xining City

first: husai

last: Ma

address: No. 55, South Street, Chengzhong District, Xining City, Qinghai Province

institute: Qinghai Red Corss  Hospital

summary: Multiple primary cancers (MPC) refer to the occurrence of two or more independent primary malignant tumors in the same patient, either simultaneously or metachronously. Their clinical diagnosis and differential diagnosis are challenging, treatment strategies are complex, and the incidence has been increasing in recent years. However, there is still a lack of potential biomarkers that can be used for early identification and prognosis assessment, which has become a key bottleneck in current research. This study employed Oxford Nanopore Technologies (ONT) long-read transcriptome sequencing and data-independent acquisition (DIA) proteomics to perform integrated multi-omics analysis of Whole blood samples from patients with only primary lung cancer (OPLC), lung cancer with MPC, and healthy controls (HC), systematically characterizing the molecular features of MPC at both transcript and protein levels. The results showed that MPC patients exhibited significantly increased transcript complexity, with the numbers of differentially expressed genes (DEGs), differentially expressed transcripts (DETs), and differential transcript usage (DTU) events being substantially higher than those in OPLC and HC groups. Multiple genes displayed rich isoform diversity. Functional enrichment analysis indicated that MPC-specific molecules were significantly enriched in immune- and inflammation-related pathways such as NF-κB, NOD-like receptor, Toll-like receptor, and TNF signaling. By integrating gene, transcript, and protein expression profiles, several core molecules (e.g., ATP6AP2, APMAP, CIAO2A, and ITGB2) with consistent expression across multiple regulatory levels were identified, all showing significant and consistent alterations in MPC. This study reveals the molecular characteristics of transcript isoform complexity in the peripheral blood of MPC patients through long-read sequencing combined with proteomics, providing important theoretical insights for understanding the mechanisms of MPC and identifying potential targets.

overall_design: We used the Oxford nanopore technology (ONT) to sequence  from blood samples from four healthy individuals, four patients with a single primary lung cancer, and four patients with multiple primary cancers.

first: Yingkui

first: Guangzhi

last: Zhu

first: Huanjuan

first: Yanfei

last: Zhang

first: Hongwu

last: Tian

first: Jin

last: Cao

first: Shengwei

last: LI

first: Guowei

last: Liu

first: Husai


title: Macrophage repolarization by immune checkpoint blockade drives T-cell engagement in the tumor microenvironment

status: Public on Dec 02 2025

city: Phoenix

first: Jessica

last: Lancaster

state: Arizona

address: 5951 E Mayo Blvd

department: Immunology

country: USA

institute: Mayo Clinic

summary: Immunotherapy combinations can improve patient outcomes, yet the interactions within the tumor microenvironment (TME) that drive therapeutic synergy are poorly understood. Tumor establishment drives monocyte recruitment and differentiation into tumor-associated macrophages (TAMs), which have essential roles in coordinating immune responses and are thus attractive targets for therapeutic modulation. In a murine model of combination anti-PD-1 and anti-PD-L1 checkpoint blockade, tumor control was associated with increased infiltration of CD8+ T-cells and M1-like repolarization of TAMs.  Live-cell imaging of the tumor microenvironment revealed close contacts between tumor-infiltrating CD8+ T-cells and TAMs, in which the extent of the contact interfaces increased with combination immunotherapy. Treatment with anti-PD-L1 was able to increase macrophage expression of pro-inflammatory factors and phagocytic activity, suggesting a role for TAMs in reactivating CD8+ T-cells in the TME. However, co-treatment with anti-PD-1 was ultimately necessary for tumor control, indicating the need for combination targeting of the TME.

overall_design: Nanostring sequencing of B16-F10 tumors grown in C57BL6/J mice and treated with 200 ug IgG2a isotype control antibodies or a combination of 200ug anti-PD-1 and 200 ug anti-PD-L1 antibodies at day 16, using Nanostring's Tumor Signaling 360 panel.

first: Tina

last: Kwok

last: N Lancaster


title: Syndecan-1-targeted therapeutic antibody inhibits macropinocytosis and induces antitumor immunity in pancreatic cancer

city: Houston

first: Haoqiang

last: Ying

state: Texas

address: 1515 Holcombe Blvd

institute: M. D. Anderson Cancer Center

type: Expression profiling by high throughput sequencing

summary: Pancreatic ductal adenocarcinoma (PDAC) remains one of the deadliest malignancies, with a 5-year survival rate of just 13%. While the development and early clinical use of small molecules targeting oncogenic KRAS mutations, key drivers of PDAC, have shown promise, resistance to these targeted therapies remains a significant challenge. We recently identified Syndecan-1 (SDC1), a highly expressed heparan sulfate proteoglycan, as a critical KRAS effector protein that promotes nutrient salvage and tumor growth. Here, we report the development of a human-specific monoclonal antibody (anti-SDC1 mAb) that inhibits PDAC cell proliferation in vitro and suppresses PDAC tumor growth in vivo. Mechanistically, anti-SDC1 mAb blocks macropinocytosis and induces antibody-dependent cellular cytotoxicity (ADCC). In vivo, anti-SDC1 mAb synergizes with standard chemotherapy, KRAS* inhibitors, and immunotherapies, resulting in tumor regression and near-complete response. These findings highlight anti-SDC1 mAb as a promising therapeutic strategy for PDAC and potentially other KRAS* and SDC1-driven tumors.

overall_design: Human AsPC-1 PDAC cells were treated with isotype or 22B under 0.2mM Gln for 3 days. Total RNA was then extracted for mRNA-seq.

first: Zecheng

last: Yang

first: Jun

last: Yao

first: Wantong


title: Integrative RNA-seq and LASSO-COX analysis reveal Paeonol's key target gene in proliferation suppression and apoptosis-induced in Cervical Cancer

city: shanghai

first: yue

last: wu

address: zhanling RD 58-304

country: China

institute: shanghai biogenuinetech co., LTD.

summary:
Abstract
Background: The natural compound paeonol exhibits therapeutic promise against cervical carcinoma, though its precise molecular mechanisms remain undefined.
Methods: First, we treated human cervical cancer (HeLa) cells with different concentrations of paeonol. Cellular proliferation and apoptotic responses were evaluated via cell-counting kit 8 (CCK8) assays and flow cytometric analysis. Subsequent transcriptomic profiling employed RNA sequencing coupled with alternative splicing assessment to detect differentially expressed genes (DEGs). Protein interaction networks were established for pivotal DEGs, followed by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment investigations. Clinical data pertinent to cervical cancer were retrieved from The Cancer Genome Atlas (TCGA). Prognostic model development incorporated Kaplan–Meier survival estimation, Least Absolute Shrinkage and Selection Operator (LASSO) regression, alongside univariate and multivariate COX proportional hazards analyses, with model accuracy subsequently assessed. Finally, Quantitative reverse transcription polymerase chain reaction (qRT-PCR) validated DEG expression.
Results: Paeonol treatment suppressed proliferation while inducing apoptosis in HeLa cells. Transcriptomic and splicing analyses revealed 12 critical DEGs: NLRP1, FN1, NQO2, NREP, B4GALNT1, ANK3, FAM219A, ODF3B, MAPK15, EPGN, MUC1, and MEG3. Enrichment analyses indicated these DEGs principally associate with inflammatory processes and the biological regulation of cellular proliferation and apoptotic death. Analysis of clinical outcomes in 197 TCGA patients demonstrated significantly enhanced five-year survival probability within the low-risk cohort. FN1, NQO2, and ODF3B were incorporated into a prognostic signature following LASSO regression. Univariate and multivariate COX analyses identified T stage, tumor grade, and differential expression of these three genes as significant outcome predictors; the resultant prognostic model exhibited robust accuracy. qRT-PCR results corroborated the RNA sequencing data concerning DEG expression patterns.
Conclusion: Paeonol modulates HeLa cell proliferation and apoptosis through regulation of 12 key genes, including FN1. This activity involves governing inflammatory responses alongside cellular proliferation, migration, and differentiation processes. These findings offer a theoretical foundation supporting paeonol's potential clinical utility in cervical cancer management.

overall_design: Hela cells in the experimental groups were treated with 0.5 mg/mL paeonol, while the control group received vehicle control (paeonol-free medium). Following 24 hours of incubation, cell proliferation and RNA-seq analyses were conducted.

first: Yuhong

last: Shang

last: Jiang

first: Miao

last: Li

last: Gao

first: Lei

last: Xu

first: Peng

last: Ge


title: TP53-Agnostic Lethality through Combined Pan-HDAC and CDK Inhibition in Acute Myeloid Leukemia

city: Seoul

first: Marc

last: Diederich

address: Gwanak-ro, Gwanak-gu, 1

department: College of Pharmacy

country: South Korea

institute: Seoul National University

summary: Tumor protein 53 (TP53)-mutated acute myeloid leukemia (AML) is characterized by poor outcomes and the quick development of treatment resistance. Here, we report that simultaneous inhibition of cyclin-dependent kinases (CDKs) and histone deacetylases (HDACs) with dinaciclib and CAY10603, respectively, eliminates the therapeutic response gap between TP53-mutant and TP53 wild-type AML. Biochemical profiling showed that CAY10603 is not only HDAC6-selective but also exhibits pan-HDAC activity similar to suberoylanilide hydroxamic acid, enabling dual targeting of transcriptional and cell cycle pathways. Across parental wild-type lines and isogenic TP53 mutants, the combination consistently suppressed clonogenic growth, induced caspase-dependent apoptosis, and downregulated key regulators such as CDK2, CDK4/6, and their cyclins, while restoring the CDK inhibitor CDKN1A/p21. In an orthotopic NSG mouse model, dinaciclib + CAY10603 significantly reduced leukemia burden and extended survival without adverse toxicity. By “normalizing” TP53-mutant AML to respond like its wild-type counterpart, this pan-HDAC/multi-CDK blockade offers a TP53-agnostic therapeutic option and warrants clinical evaluation as a strategy that remains effective regardless of baseline allelic status.

overall_design: HL-60 and U937 cells were treated with HDAC inhibitors CAY10603 or SAHA for 24 hours at a concentration of 2 µM in three independent biological replicates.

first: Pottier

last: Aurelien

first: Li

last: Vladimir

first: Morceau

last: Franck

first: Diederich

last: Marc


title: Hypergravity Enhances Stretch Sensitivity in Rat Cardiomyocytes via Increased Expression and Activity of Stretch-Activated Channels

city: Moscow

first: Vadim

last: Mitrokhin

address: Ostrovityanova

country: Russia

institute: RNRMU

summary: This study investigated how 14-day 4g hypergravity exposure affects mechanosensitive ion channels (MSCs) in rat ventricular cardiomyocytes. RNA-seq revealed selective MSC remodeling: TRPM7 (+41%), TRPC1 (+68%), and TRAAK (+239%) were upregulated, while TRPV2 (-62%), Piezo2 (-58%), and Kir6.1 (-76%) were downregulated. Structural analysis showed 44% increased membrane capacitance and sarcomere elongation, consistent with hypertrophic remodeling. Patch-clamp studies demonstrated markedly hypersensitive stretch-activated currents (I_SAC), with activation at minimal displacements (1-2 μm) and enhanced responses under 4-10 μm stretch (107-222% of controls). These coordinated molecular, structural, and functional adaptations suggest targeted transcriptional reprogramming that balances enhanced Ca²⁺ signaling with stabilized K⁺ conductance, demonstrating cardiac mechanotransduction plasticity under gravitational stress with implications for cardiovascular risks in altered gravity environments.

overall_design:
Male Wistar rats (8 weeks old) were divided into two groups: hypergravity-exposed and control. The hypergravity group was subjected to 4g centrifugal force for 14 days (8 hours/day). Ventricular cardiomyocytes were isolated from both groups for RNA sequencing and electrophysiological analysis. RNA-Seq was performed to profile transcript levels of ion channels, while patch-clamp techniques assessed the sensitivity of stretch-activated currents (ISAC) to mechanical deformation. Data include raw RNA-Seq files and processed expression values for mechanically sensitive ion channels.

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


title: Age-dependent transcriptomic effects of morphine in the frontal cortex of female mice

city: Boston

first: Jonathan

middle: David

last: Lee

state: MA

address: 3 Blackfan Circle, Rm. 424

institute: BIDMC

summary: Increasing lifespans make health problems in the elderly such as opioid misuse a more prominent concern. Understanding the effects that opioids may have on the aged brain can help us address age-related concerns of opioid exposure. This study aimed to assess potential interactions between aging and opioid exposure. Three-month-old (young adult) and 19-month-old (aged) C57BL/6JN mice were assigned to either a morphine (3mg/kg, i.p.) or saline group. A conditioned placed preference (CPP) task was used to assess reward sensitivity, while rotarod and beam walk tests were used to assess sensorimotor coordination. To assess for potential age-dependent effects of morphine on gene expression, we performed RNA sequencing in the prefrontal cortex (PFC). We found that morphine induced CPP in both age groups. Our results indicate impaired motor coordination in aged mice; however, morphine did not significantly affect motor coordination in either age group, although a trend toward an increased number of slips was observed in morphine-treated aged mice. Transcriptomic analysis revealed more robust effects of morphine on gene expression in the aged brain compared to the young brain. Interestingly, we found limited overlap between morphine-regulated genes in young and old mice, suggesting that the molecular effects of morphine are age-dependent. Taken together, while we found no significant interactions between morphine (at the tested dose) and aging in the behavioral assays, morphine caused age-dependent gene expression changes. Our findings suggest that age should be considered when prescribing opioids and that age-specific therapeutics may help address opioid use disorder in the elderly.

overall_design: RNA-seq profiling of mouse prefrontal cortex; young adult or old female mice were treated with morphine (3mg/kg, i.p.) or saline

middle: D

first: Maria

last: Mavrikaki

first: Frank

middle: J

last: Slack


title: VILMIR is a trans-acting long noncoding RNA that enhances the host interferon response in human epithelial cells

city: Raleigh

first: Xinxia

last: Peng

address: 1060 William Moore Drive

institute: North Carolina State University

summary: Long noncoding RNAs (lncRNAs) have been found to play significant regulatory roles within antiviral and immune responses. We previously identified the novel lncRNA virus-inducible lncRNA modulator of interferon response (VILMIR), that was found to broadly regulate the host transcriptional response to interferon-beta (IFN-β) treatment in A549 human lung epithelial cells. Here, we investigated the mechanism by which VILMIR regulates the host interferon response in trans by identifying interacting proteins and gene regulatory networks of VILMIR. Through an RNA pull-down assay, we found that VILMIR interacted with both nuclear and cytoplasmic proteins in vitro, including the transcriptional regulators FUBP1 and PUF60 in the nucleus, as well as the antiviral proteins IFIT1 and IFIT3 and the aminoacyl-tRNA synthetases QARS1 and KARS1 in the cytoplasm. In addition, we found that overexpression of VILMIR in A549 cells resulted in an overall enhancement of host interferon response genes and identified a core set of interferon-stimulated genes that were consistently regulated by VILMIR knockdown and overexpression. Finally, we proposed several possible mechanisms by which VILMIR may interact with the identified proteins to regulate the interferon response, such as by interacting with FUBP1 and PUF60 in the nucleus to regulate host transcription in trans or by interacting with the IFIT proteins and aminoacyl-tRNA synthetases in the cytoplasm to regulate translation.

overall_design: To investigate the functional role of VILMIR during the human interferon response, we generated an A549 cell line overexpressing ectopic VILMIR as well as an empty vector control line. The control cell line as well as the VILMIR-overexpressing cell line were treated with or without human IFN-β at 1 ng/mL or 10 ng/mL for 6 hours in triplicate. RNA-sequencing and differential expression analysis was then performed to identify the impact of VILMIR overexpression on host transcription before and after IFN-β treatment.

first: Kristen

last: John

first: Ethan

last: Smith

first: Alexandra

last: Istishin

first: Nasif

last: Mahmood

first: Kayleigh

last: Diveley

first: Tammy

middle: S

last: Tollison

first: Susan

last: Carpenter


title: TET3 is a regulator and can be targeted for the intervention of myocardial fibrosis [ATAC-seq]

city: Nanjing

first: yujia

last: xue

address: Longmian Avenue 639

institute: China Pharmaceutical University

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Cardiac fibrosis as an integral part of the wound healing process on the one hand safeguards myocardial homeostasis but on the other hand contributes to adverse cardiac remodeling and loss of heart function eventually leading to heart failure (HF). Resident cardiac fibroblasts are the principal source of myofibroblasts that produce extracellular matrix proteins to mediate cardiac fibrosis. We investigated the involvement of ten-and-eleven translocator 3 (TET3) in this process focusing on the regulatory mechanism and translational potential. We report that TET3 depletion in cultured cardiac fibroblasts blocked transition to myofibroblasts in response to different pro-fibrogenic stimuli. Consistently, deletion of TET3 from quiescent or activated fibroblast (myofibroblast) attenuated cardiac fibrosis and rescued heart function in models of cardiac injury. Importantly, a small-molecule TET3-specific degrader Bobcat339 displayed therapeutic potential by mitigating cardiac fibrosis and normalizing heart function when administered post-surgery. Integrated transcriptomic analysis (RNA-seq and CUT&Tag-seq) identified the mechanosensor Piezo2 as a downstream target for TET3. Piezo2 deletion or inhibition dampened fibroblast activation in vitro and ameliorated cardiac fibrosis in vivo. Mechanistically, Piezo2 promoted fibroblast activation by modulating the activities of mechanosensitive transcription factors. Finally, relevance of the TET3-Piezo2 axis was verified in heart specimens collected from HF patients. Our data demonstrate that TET3 is a pivotal regulator of cardiac fibrosis and can be potentially targeted for the intervention of heart failure.

overall_design: Mouse primary cardiac fibroblasts were induced to undergo Tet3 knockout by transfection with Cre recombinase virus, followed by TGF-β treatment.

first: Chenghao

last: Zhu

first: Wenxuan

last: Hong

first: Yuwen

first: Yujia

last: Xue

first: Zemin

last: Fang

first: Dingsheng

last: Jinag

first: Yong

last: Xu

first: Ming

last: Kong


title: TET3 is a regulator and can be targeted for the intervention of myocardial fibrosis.


title: A chemical-genetic interaction between PAF1 and ENL/AF9 YEATS inhibition

city: La Jolla

first: Michael

last: Erb

state: CA

address: 10550 N. Torrey Pines

institute: The Scripps Research Institue

summary: Transcriptional regulatory proteins are frequent drivers of oncogenesis and common targets for drug discovery. The transcriptional co-activator, ENL, which binds to chromatin through an acetyllysine reader YEATS domain, is preferentially required for the survival and pathogenesis of acute leukemia. Small molecules that inhibit the ENL YEATS domain show anti-leukemia effects in preclinical models, which is thought to be caused by the downregulation of pro-leukemic ENL target genes. However, the transcriptional effects of ENL YEATS domain inhibitors have not been studied in models of intrinsic or acquired resistance and, therefore, the connection between proximal transcriptional effects and downstream anti-proliferative effects is poorly understood. To address this, we identified models of intrinsic and acquired resistance and used them to study the effects of ENL YEATS domain inhibitors. We first discovered that ENL YEATS domain inhibition produces similar transcriptional responses in naive models of sensitive and resistant leukemia. We then performed a CRISPR/Cas9-based genetic modifier screen and identified in-frame deletions of the essential transcriptional regulator, PAF1, that confer resistance to ENL YEATS domain inhibitors. Using isogenic models of PAF1-mediated resistance, we again found that the downregulation of ENL target genes is shared in both sensitive and resistant leukemia. Altogether, these data support the conclusion that the suppression of ENL target genes is not sufficient to explain the anti-leukemia effects of ENL antagonists.

overall_design: Samples Gene expression response to ENL antagonist, SR-0813. In this data submission there are the PAF1-1 and PAF1-4 mutant MOLM13 cells that have been selected in SR-0813 for an extended period of time and monitored by flow cytometry. These cells were removed from compound or DMSO for a passage and then re-treated with compound (10 µM SR-0813) or control (DMSO). Samples were compared to their respective DMSO control.

first: Paige

middle: A

last: Barta

first: Leopold

last: Garnar-Wortzel


title: 3D-ASC-Exos as a novel drug carrier for glioma treatment

city: Changchun

first: Boyin

last: Jia

state: Jilin

address: 2888 Xincheng Street

department: College of Animal Science and Technology

institute: Jilin Agricultural University

type: Non-coding RNA profiling by high throughput sequencing

summary: It is known that antler stem cell extract has an inhibitory effect on glioma growth. Therefore, it is speculated that antler stem cell-derived exosomes (ASC-Exos) can be used as a new drug carrier for the treatment of glioma. Due to the limitation of exosome sources, we constructed a 3D culture system to harvest more exosomes. In this experiment, compared with the 2D-ASC-Exos obtained from the traditional 2D culture, the 3D-ASC-Exos obtained from our constructed 3D culture system were concentrated nearly 30 times in the culture medium volume, which was more convenient for subsequent purification. The two forms of ASC-Exos had similar morphologies and surface markers, but 3D-ASC-Exos were enriched with more miRNAs related to tumor suppression. In vitro experiments demonstrated that 3D-ASC-exosomes loaded with temozolomide (TMZ) inhibited the proliferation, migration and invasion abilities of glioma cells and promoted the apoptosis of glioma cells. The vivo tumor-bearing mouse model demonstrated that 3D-ASC-Exos loaded with TMZ exerted tumor-suppressive effects by inhibiting tumor growth and promoting tumor apoptosis. Meanwhile, the treatment with 3D-ASC-Exos loaded with TMZ caused no damage to the various tissues and organs of mice compared with the TMZ group. In conclusion, 3D-ASC-Exos can be used as a novel drug carrier for the treatment of glioma.

overall_design: 3D-ASC-Exos  were collected from 3D culture system. Total RNA pooled was collected from 3D-ASC-Exos to construct a cDNA library. 3D-ASC-Exos miRNAs were analyzed by Illumina NovaSeq 6000 for high-throughput deep sequencing.

first: Xintong

last: Han

first: Xin

last: Li

first: Xinrui

last: Yan


title: Transcriptomic analysis of KMT5A-knockout healthy volunteer peripheral blood CD8+ T cells under activated conditions

city: Shanghai

first: Xiaoling

last: Tian

address: Dongchuan road,500

institute: East China Normal University

summary: This study aims to investigate the role of Lysine Methyltransferase 5A (KMT5A, also known as SETD8) in CD8+ T cell biology. CD8+ T cells were isolated from healthy volunteer PBMCs using magnetic bead-based negative selection. Isolated cells were subjected to CRISPR-Cas9 mediated gene editing targeting KMT5A (KMT5A-KO group) or a non-targeting control guide RNA (control group, CTRL) via ribonucleoprotein (RNP) complex electroporation. Cells were harvested at 72 hours post-electroporation for RNA extraction. RNA sequencing (RNA-seq) libraries were constructed using the Illumina TruSeq Stranded mRNA kit and sequenced on an Illumina platform to generate paired-end reads. Transcriptome profiles of KMT5A-KO CD8+ T cells were compared to those of CTRL cells to identify differentially expressed genes (DEGs) and elucidate potential pathways regulated by KMT5A in this cell type.

overall_design: This study employed a comparative transcriptomic approach to assess the effects of KMT5A gene knockout on CD8+ T cells. The primary comparison is between CD8+ T cells edited with KMT5A-targeting sgRNA RNP (KMT5A-KO group) and CD8+ T cells edited with non-targeting control sgRNA RNP (CTRL group). All samples were derived from CD8+ T cells isolated from healthy volunteer PBMC and processed in parallel.

first: Guolong

last: Liu


title: Construction of a single-cell transcriptome atlas for Pogostemon cablin embryoids reveals PcNAC048 as a dual regulator coordinating lateral root morphogenesis and patchouli alcohol biosynthesis

city: Guangzhou

first: xiaobing

last: wang

address: Higher Education Mega center, Panyu, Guangzhou

institute: Guangzhou University of Chinese Medicine

summary: We have employed a single cell sequencing approach using 10x Genomics scRNAseq to study the single-cell atlas of Pogostemon cablin globular somatic embryos (GSE) and cotyledonary somatic embryos (CSE). Pseudo-time analysis reconstructed the continuous differentiation trajectory of apical meristem cells and epidermal cells. Further research identified PcNAC048 as a potential transcription factor regulating embryonic cell differentiation, which promotes lateral root development and negatively regulates the biosynthesis of patchouli alcohol. Overall, this study provides theoretical support for the development of Pogostemon cablin germplasm resources and the regulation of medicinal compounds.

overall_design: Protoplasts of globular somatic embryos (GSE) and cotyledonic somatic embryos (CSE) in P. cablin were isolated by enzymatic hydrolysis and analyzed using ScRNA-seq

first: Xiaobing

last: Wang

first: Xiaojin

last: Ge

last: Zhong

first: Daidi

last: Wu

first: Zhiye

last: Yang

first: Ruoting

last: Zhan

first: Likai

last: Chen

first: Hua


title: The light-responsive chromatin accessibility landscape in rice [RNA-seq]

city: Chicago

first: CHUAN

last: HE

state: Illinois

address: 929 East 57th Street, GCIS E313

institute: University of Chicago

summary: Here, we show that light greatly altered chromatin accessibility during the switches from skotomorphogenesis to photomorphogenesis

overall_design: We first extracted the total RNA of samples. poly(A) RNA of each biological replicate was captured first for RNA-sequencing


title: RELB Reprograms Exhausted Tumor-Infiltrating Lymphocytes for Improved Adoptive Cell Therapy [proliferation]

city: Durham

first: Charles

last: Gersbach

state: NC

address: 101 Science Drive

institute: Duke University

summary: Tumor-infiltrating lymphocytes (TILs) are a promising autologous cell therapy to treat solid tumors. TILs are manufactured by expanding and reinfusing tumor-reactive T cells from tumor biopsies. Efficacy of TIL therapies has been limited by the heterogeneity of expanded TIL products and the high prevalence of dysfunctional exhausted CD8+ T cells (TEX). A subset of CD8+ TILs that are double-positive (DP) for CD103 and CD39 is enriched for tumor-reactive TILs across multiple cancer types, but also is susceptible to the TEX state.  We screened overexpression of all human transcription factors (TFs) to discover master regulators of expansion in DP TILs from non-small cell lung cancer patients.  RELB emerged as the dominant hit driving proliferation of DP TILs despite exhaustion induced by repeat stimulation, with a skew towards CD8+ cells. TCR-seq showed maintenance of TCR diversity from the initial TCR repertoire after multiple days of in vitro expansion driven by RELB. Transcriptome profiling of multiple RELB-expressing TIL subtypes revealed a shift towards a memory/costimulatory-like phenotype. Using a HER2-targeting CAR and tumor co-culture model, RELB conferred improved persistence after multiple tumor challenges in vitro and improved solid tumor control in mouse xenografts in vivo. Finally, co-culture of RELB-overexpressing TILs with patient-matched tumor organoids showed an increase in TIL product polyfunctionality, tumor reactivity, and tumor killing.

overall_design: CD8+ TILs were isolated using a TIL CD8+ magnetic isolation kit (REAlease® CD8 (TIL) MicroBead Kit, human, Miltenyi) and DP TILs were FACS sorted (n = 2 patient samples), activated with Transact (Miltenyi) according to manufacturer’s instructions, and transduced (n = 2 transduction replicates) with the MORF library at low MOI. Cells were expanded for 3 days, selected with puromycin at 1 µg/mL for 3 days, and then expanded for an additional 9 days. Cells were then re-activated at day 12 post-transduction with a 3:1 ratio of CD3/CD28 dynabeads (Invitrogen) to T cells, stained with CellTrace Violet (Invitrogen), and sorted for lower and upper 15% tails of CellTrace signal. All replicates were maintained and sorted at a minimum of 400x coverage. Cells were maintained at 1–2 × 106 cells ml−1 for all expansion steps unless otherwise indicated.

first: Christian

last: McRoberts Amador

first: Rachel

middle: E

last: Conover

first: Pamela

middle: K

last: Noldner

first: Ying

last: Zhou


title: RELB Reprograms Exhausted Tumor-Infiltrating Lymphocytes for Improved Adoptive Cell Therapy [TCR-Seq]

overall_design: RNA was isolated from distinct TIL patient samples (n=4) that were transduced with either Thy1.1, RELB, or nothing using Norgen’s Total RNA Purification Plus Kit and Takara’s SMARTer Human TCR a/b Profiling Kit v1 was used to generate TCR-seq libraries. Library generation and purification methods were carried out according to manufacturer protocol. Initial data processing was performed using MiXCR using the present for the SMARTer Human TCR a/b kit. The R package immunarch  was used to analyze effective number of types for TCR diversity across samples to account for both number of clones and relative abundance.


title: Triple checkpoint blockade of PD-1, Tim-1 3 and Lag-3, but not PD-1 alone, enhances adoptive T cell immunotherapy in a mouse model of ovarian cancer

city: Charlottesville

first: Gary

last: Owens

state: VA

address: 415 Lane Road MR5 Building Room 1322

institute: University of Virginia

summary: The five-year survival rate for ovarian cancer patients remains below 50%, underscoring the need for innovative therapies. One promising approach involves engineering T cells to specifically target proteins uniquely overexpressed in tumors, thereby controlling tumor growth without toxicity to healthy tissues. Mesothelin (Msln) contributes to the malignant and invasive phenotype in ovarian cancer and has limited expression in healthy cells, making it a candidate immunotherapy target. Our previous results in a mouse model of ovarian cancer demonstrated that T cells engineered to express a T cell receptor (TCR) targeting Msln (TCRMSLN) mediated therapeutic activity, delaying tumor growth and prolonging mouse survival. However, inhibitory ligands expressed in the tumor microenvironment (TME) interacted with inhibitory receptors on activated T cells, suppressing antitumor function. We hypothesized combining engineered T cells with checkpoint blockade would enhance T cell function in the TME and improve therapeutic efficacy, but administration of mono-specific antibodies targeting individual inhibitory pathways had no significant impact on T cell efficacy. By contrast, the combination of PD-1, Tim-3 and Lag-3 blockade with engineered T cells significantly improved T cell function and overall animal survival relative to treatment with antibody alone or TCRMSLN with singlet or doublet antibody combinations. Single-cell RNA sequencing revealed TCRMSLN T cells treated with the triplet antibody combination increased expression of genes involved in interferon responses and metabolic function, and reduced expression of genes associated with exhaustion. These results suggest that strategies to disrupt multiple inhibitory pathways simultaneously may be necessary for improved adoptive T cell therapy efficacy in patients.

overall_design: Mice were injected with an ovarian cancer cell line, and then we performer adoptive immunotherapy. We isolated engineered T cells that target Mesothelin (Msln) from these animals and performed scRNA-seq on them

first: Gabriel

middle: F

last: Alencar

first: Asmaa

middle: O

last: Mohamed

first: Madison

middle: G

last: Burnett

first: Samantha

last: Jean

first: Anders

last: Nelson

first: Yapeng

last: Su

first: Valentin

last: Voillet

first: Magdalia

middle: R

last: Suarez

first: Susan

last: Ruskin

first: Lam

last: Trieu

first: Jennifer

middle: L

last: Lam

first: Stefan

last: Bekiranov

first: Raphael

last: Gottardo

first: Philip

last: Greenberg

first: Kristin

last: Anderson


title: The light-responsive chromatin accessibility landscape in rice

first: CHUAN

last: HE

overall_design: Freshly collected seedlings were immediately chopped with a razor blade for 5 min in 100 μL lysis buffer (20 mM MES, pH 5.7, 0.6M Mannitol, 10 mM MgCl2, 10 mM KCl, 0.1% 2-mercaptoethanol and 0.2% Triton X-100, 1x PI (protease inhibitor)) on ice (two biological replicates were performed) according to our previous work(10) . Thereafter the chopped slurry was washed into 1 ml lysis buffer in a 1.5ml tube and incubated for 5 min on ice with gentle shaking. The mixture was filtered through a 40-μm filter into a new tube, followed by centrifugation at 500 g for 8 min (4 °C). Then, we resuspended the precipitation with 150 µl wash buffer (20 mM Tris-HCl pH 7.8, 10 mM MgCl2, 60 mM KAc pH 5.6). The above washed 50,000 nuclei were then tagged with TS-Tn5 at 37ºC for 30 min following the kit’s instructions (Vazyme, TD501). After tagging, the integration products were purified using DNA Purification Kit (ZYMO, D4004) and then amplified using Q5 high fidelity DNA polymerase (New England Biolabs, M0491S) for 10-15 cycles. The libraries were sequenced on Illumina HiSeq X Ten with paired-end reads of 150-bp


title: RELB Reprograms Exhausted Tumor-Infiltrating Lymphocytes for Improved Adoptive Cell Therapy [RNA-Seq]

first: Charles

last: Gersbach

overall_design: RNA was isolated from distinct TIL patient samples (n=4) that were transduced with either Thy1.1, RELB, or nothing using Norgen’s Total RNA Purification Plus Kit and submitted to Azenta for standard RNA-seq with polyA selection

first: Christian

last: McRoberts Amador

first: Pamela

last: Noldner

first: Ying

last: Zhou


title: ADAP-METTL3 modulates the inflammatory responses of macrophages via m6A modification of Spry1

city: Suzhou

first: Anqi

last: Dong

address: 199 Ren'ai Road

institute: Soochow university

summary: While N6-methyladenosine (m6A) RNA modification is implicated in macrophage inflammatory responses, its regulatory mechanisms remain elusive. Our prior studies demonstrated that a deficiency in immune adaptor protein ADAP promotes inflammation in TLR4-stimulated macrophages. Here, we show that ADAP binds to METTL3, and depletion of Mettl3 alleviates the hyperinflammation in Adap-/- macrophages, indicating METTL3 counteracts the anti-inflammatory function of ADAP in macrophages. Furthermore, LPS induces the METTL3-dependent m6A methylation of Spry1 mRNA in macrophages in the A6988 within the motif GGACU, which is further potentiated upon the depletion of ADAP. Moreover, this positive effect of ADAP deficiency on LPS induced m6A methylation of Spry1 mRNA is dependent on IGF2BP2, which specifically binds to and stabilize the m6A modified Spry1 mRNA that contributes to an exacerbation of the inflammation in Adap-/- macrophages via NF-κB activation in macrophages. Together, our findings unveil a reciprocal inhibition between ADAP and METTL3 fine-tunes the inflammatory responses of macrophages via modulation of the m6A methylation of Spry1 mRNA.

overall_design: Peritoneal macrophages from WT and Adap-/- mice were either unstimulated or stimulated with LPS (1 ug/ml, 6 h). Isolated RNA was subjected to MeRIP-seq.

first: Pengchao

last: Zhang

first: Fan

last: Yang

first: Yanqi

last: Wang

first: Hebin

last: Liu


title: HALO: Hierarchical Causal Modeling for Single Cell Multi-Omics Data

city: Pittsburgh

first: Eleanor

last: Valenzi

state: PA

address: 3459 Fifth Avenue

institute: University of Pittsburgh

summary: Though open chromatin may promote active transcription, gene expression responses may not be directly coordinated with changes in chromatin accessibility. Most existing methods for single-cell multi-omics data focus only on learning stationary and shared information among these modalities, overlooking modality-specific information delineating cellular states and dynamics resulting from causal relations among modalities. To account for this, the epigenome and transcriptome relationship can be characterized in relation to time as “coupled” (changing dependently) or “decoupled” (changing independently). We propose the framework HALO, which adopts a causal approach to model these temporal causal relations on two levels. On the representation level, HALO factorizes these two modalities into both coupled and decoupled latent representations, identifying the dynamic interplay between chromatin accessibility and transcription through temporal modulations in the latent space. On the individual gene level, HALO matches gene-peak pairs and characterizes changing dynamics between gene expression and local peaks with time. HALO reveals bipotency in a subset of AT2 cells that exhibit different decisions in lineage specification between systemic sclerosis (SSc) and normal conditions. We demonstrate that using coupled and decoupled information, HALO discovers analogous biological functions between modalities, distinguishes epigenetic factors for lineage specification, and identifies temporal cis-regulation interactions relevant to cellular differentiation and complex human diseases.

overall_design:
Explanted lung tissue from patients with systemic sclerosis-associated interstitial lung disease was collected at the time of lung transplant.  Donor control lung tissue was collected at the time of organ and tissue donation.  All samples were processed fresh for snATAC-seq or multiomic sequencing. Raw data are unavailable due to patient privacy concerns.

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

first: Mao

last: Haiyi

first: Jia

last: Minxue

first: Di

last: Marissa

first: Valenzi

last: Eleanor

first: Cai

last: Xiaoyu Tracy

first: Lafyatis

last: Robert

first: Zhang

last: Kun

first: Benos

last: Panagiotis


title: Effects of UBE2S on colorectal cancer cell line RKO

first: Jun

last: Li

state: None Selected

institute: Shanghai Jiao Tong University School of Medicine affiliated Ruijin Hospital

summary: To assess effects of UBE2S on colorectal cancer after UBE2S overexpression and knockdown in RKO cell line and to explore the potential signaling pathways, we analyzed the changes in mRNA expression levels in UBE2S-treated cells through RNA sequencing. This approach allowed us to determine the activation of relevant signaling pathways within the cells.


title: TET3 is a regulator and can be targeted for the intervention of myocardial fibrosis [RNA-seq]

first: yujia

last: xue

first: Chenghao

last: Zhu

first: Wenxuan

last: Hong

first: Yuwen

first: Yujia

last: Xue

first: Zemin

last: Fang

first: Dingsheng

last: Jinag

first: Yong

last: Xu

first: Ming

last: Kong


title: Comparison of gene expression between MACs and MIL4-MACs

city: Habikino

first: Akifumi

last: Matsuyama

state: Osaka

address: 3-7-1 Habikino

department: Center for reverse Translational Research

country: Japan

institute: Osaka Habikino Medical Center

summary: MIL4-MACs may release additional pro-angiogenic factors aside from IL-8 and could be classified as anti-inflammatory.

overall_design: To confirm the high levels of angiogenesis-related genes in MIL4-MACs, those of MIL4-MACs have been compared to those of MACs.

first: Teruhide

last: Yamaguchi


title: Comparative analysis of replicative and doxorubicin-induced senescence identifies conserved transcriptomic signatures in aging fibroblasts [RNA-Seq1]

city: Xi'an

first: Wei

last: Sun

state: Shaanxi

address: No. 229 Taibai North Road, Beilin District

department: College of Life Sciences and Medicine

institute: Northwest University

summary: We established parallel replicative and doxorubicin-induced senescence models in human BJ fibroblasts, demonstrating progressive morphological changes and SA-β-gal activity increases across passages (P3-P33). Molecular analyses confirmed expected senescence markers: p21/p53 upregulation and LMNB1 downregulation at RNA/protein levels. Transcriptomic profiling identified 734 upregulated (extracellular matrix organization) and 195 downregulated (cell cycle) genes. Comparative lncRNA analysis revealed 23 consistently upregulated candidates, with PURPL showing the most dramatic senescence-associated induction. These validated models provide a robust platform for mechanistic aging studies, with PURPL emerging as a key regulator of cellular senescence.

overall_design: RNA-seq of BJ (P3, P11, P24, P30, and P33) and DOX-induced cells.

first: Jie

last: Wang

first: Yongqiang

last: Ning

last: Wei


title: PURPL bidirectionally controls senescence: depletion rejuvenates fibroblasts while overexpression accelerates aging [RNA-Seq2]

summary: ellular senescence is a fundamental driver of aging and age-related diseases, with long non-coding RNAs (lncRNAs) emerging as critical regulators of this process. Here, we identify PURPL as a bidirectional modulator of senescence in human fibroblasts. Through comparative transcriptomic analysis of replicative and doxorubicin-induced senescence models, we discovered PURPL as the most significantly upregulated lncRNA common to both systems. CRISPR interference (CRISPRi)-mediated PURPL depletion in senescent cells reversed hallmark aging phenotypes, including restoration of LMNB1 expression, suppression of p21, and reduced SA-β-gal activity. Conversely, PURPL overexpression in young fibroblasts recapitulated senescence signatures, inducing extracellular matrix reorganization and cell cycle arrest pathways. RNA-seq revealed PURPL overexpression dysregulated 1,633 genes, mirroring natural aging transcriptomes. These opposing effects establish PURPL as a central regulator of cellular senescence, suggesting its therapeutic potential for age-related conditions. Our findings provide mechanistic insights into lncRNA-mediated senescence control and highlight PURPL as a novel target for aging intervention strategies.

overall_design: RNA-seq of CRISPRi-mediated PURPL knockdown and control cell lines, alongside PURPL-overexpressing and corresponding control cell lines

first: Xiao

last: Yang


title: Osteocytic Wnt generates RANKL for osteoclastogenesis via TGFβ signaling

city: Chongqing

first: Yujiao

last: Liu

state: Chongqing

address: No.1 Xueyuan Road, Yuzhong District

department: College of Laboratory Medicine,

institute: Chongqing Medical University

summary: Wnt signaling is generally believed to inhibit osteoclast differentiation (OcD) by upregulating its target Opg expression; however, we unexpectedly found that Wnt mice with activation of Wnt/β-catenin signaling in osteocytes have higher OcD and bone resorption while generating more bone (PNAS 2015,112: E478). Here, we show cellular and molecular mechanisms for this unpredictable function of osteocytic Wnt on OcD. Isolated osteocytes (daCOt) of the Wnt mice display higher expression of RANKL and Opg with increased RANKL/Opg ratio, promoting OcD of bone marrow monocytes/macrophages to enhance bone resorption. RANKL-siRNA knocks down RANKL to reduce this OcD. In daCOt transcriptome analysis, the enrichment of KEGG pathways highlights the largest correlation of OcD with TGFβ signaling, which is activated by highly expressed Tgfb1 and Tgfb2, identified by heat-map analysis, and verified in a dose-dependent manner in Wnt agonist C91-treated wild-type osteocytes. Moreover, TGFβ signaling increases RANKL expression in dose and time-dependent manners, respectively with higher RANKL/Opg ratio and OcD. Conversely, TGFβ signaling inhibition reverses these effects. Importantly, mice with osteocytic deletion of the only TGFβRII decreases osteoclast number by 48.2% to 3.0/mm in Oc.N/B.Pm with markedly reduced expression of RANKL both at protein and mRNA levels in the osteocytes of cancellous bone in distal femurs. Furthermore, luciferase assay with 3kb-RANKL promoter region reveals that TGFβ signaling activates RANKL transcription. In summary, osteocytic Wnt promotes OcD mainly by activating TGFβ signaling, which activates RANKL gene transcription, opening up a novel avenue for the treatment or application of osteoclast or bone resorption-related diseases.

overall_design: The RNA from primary osteocytes of three wild-type (wt) littermate mice and primary  osteocytes of three transgenic mice with activated Wnt signaling was extracted for high-throughput sequencing analysis.

first: Lizhou

last: Zhao

first: Molin


title: A Stem and Progenitor Cell-Derived Gene Expression Signature is Prognostic for Survival in Myelofibrosis

city: Toronto

first: Alexander

middle: James

last: Murison

state: Ontario

address: 101 College Street, TMDT room 11-706

country: Canada

institute: University Health Network

summary: We hypothesized that transcriptomic features among disease-driving hematopoietic stem and progenitor cells (HSPC) could improve the current paradigm for risk stratification in myelofibrosis (MF). Therefore, we performed bulk RNA-seq on blood from 358 MF patients split into training and test cohorts (ClinicalTrials.gov Identifier: NCT02760238). Single-cell (sc) RNA-seq data from Lin-CD34+ MF HSPCs were used to guide the development of a prognostic model trained on the bulk RNA-seq data. A NanoString assay was used to validate our final model in two independent cohorts of MF patients. We identified a 24-gene weighted-sum expression score (termed MPN24) that was prognostic of overall survival (OS). In two independent cohorts of 170 and 99 MF patients, MPN24-High scoring patients had worse 5-year OS compared to MPN24-Low scoring patients (25.5% vs 86.9%, HR = 9.81, P = 4.0e-11 and 23.5% vs 75.9%, HR = 6.40, P = 1.8e-7, respectively). MPN24-High was also associated with clinical, mutation and karyotypic features known to confer adverse risk in MF. The MPN24 score retained independent prognostic significance in multivariable analysis incorporating these covariates as well as existing risk stratification models including DIPSS, DIPSS Plus, MIPSS70, and MIPSS70 Plus v2.0, adding significant prognostic value to these baseline risk stratification models (P = 1.6e-5, 4.0e-6, 1.5e-6, 5.4e-4, respectively). The MPN24 score was particularly useful in improving risk-stratification of DIPSS-Intermediate-1 and Intermediate-2 and MIPSS70 Intermediate and High-risk patients. An HSPC-derived gene expression score is associated with overall survival independent of existing clinical and genomic prognostic variables and improves risk stratification in MF.

overall_design: We profiled the peripheral blood mononuclear cells (PB-MNCs) from 358 myelofibrosis patients (PMc1 cohort) and correlated transcriptomic profiles to overall survival to derive a 24-gene prognostic score (termed, MPN24). We benchmarked the ability of a custom NanoString-based assay to measure the 24-gene score against RNA-seq in 59 PMc1 patient samples. Additionally, we validated MPN24 using the NanoString assay in PB-MNCs from two independent cohorts consisting of 170 (PMc2 cohort) and 99 (MSK cohort) myelofibrosis patients.

first: Jessie

last: Medeiros

first: Andy

last: Zeng

first: Michelle

last: Chan-Seng-Yue

first: Hyerin

last: Kim

first: Tristan

last: Woo

first: Suraj

last: Bansal

first: Jessica

last: McLeod

first: Sagar

last: Kothari

first: Andrea

last: Arruda

first: Hubert

last: Tsui

first: Jaime

last: Claudio

first: Dawn

last: Maze

first: Hassan

last: Sibai

first: Mark

last: Minden

first: James

last: Kennedy

first: Christopher

last: Famulare

first: Raajit

last: Rampal

first: Jean

middle: C

first: John

last: Dick

first: Vikas

last: Gupta


title: Transcriptomic architecture of weak versus strong fear extinction learning uncovers extinction-suppressive role of Tia1

city: New Haven

first: In-Jung

state: CT

address: 300 George Street

department: Ophthalmology

institute: Yale University

summary:
BACKGROUND: Extinction, the capacity for an individual to inhibit or diminish maladaptive fear memories, is a critical aspect of fear processing. In humans, weak extinction learning is often observed in anxiety and fear-related disorders, such as PTSD. However, the mechanisms behind regulating extinction and determining individual variability in extinction learning remain poorly understood.
METHODS: To investigate the molecular basis of inter-individual and sex-related differences in the ability to extinguish fear, extinction learning was analyzed in inbred wild-type male and female mice. Extinction tests were combined with in vivo profiling of the differential hippocampal transcriptomes associated with weak and strong extinction learning and genetic manipulations to extend our transcriptomic findings.
RESULTS: We identified significant sex-dependent and -independent differences in hippocampal gene expression between weak and strong extinction learner animals. Especially surprising was a very high transcriptomic overlap between weak learner males and females, showing upregulation of multiple genes associated with neurotoxic insult and cellular stress, including a major stress regulator, a prion-like Tia1. Viral overexpression of Tia1 in the dorsal hippocampus caused sex-independent dysregulation of microglia and diminished fear extinction learning in animals of both sexes.
CONCLUSIONS: Altogether, we demonstrated the brain-based transcriptomic architecture associated with weak versus strong fear extinction learning in male and female mammalian subjects and identified the sex-independent extinction-suppressive role of hippocampal Tia1 upregulation. Our results should help better understand the mechanisms underlying individual, and sex-dependent differences in extinction and inform novel therapeutic targets for pharmacological extinction augmentation strategies in fear-related disorders.

overall_design: Transcriptom analysis of the hippocampi from the wild-type inbread male female mice with differing an amenability to fear extinction

first: Maria

middle: I

last: Bonilla

first: Hae-Lim

last: Lee

first: Stephen

last: Foster

first: Andrii

last: Rudenko


title: Inducible CD147 up-regulation boosts persistent SARS-CoV-2 infection triggering severe COVID-19 independent of ACE2 [scRNA-seq]

city: Xi’an

first: Ping

last: Zhu

address: Changle Xi Road

institute: Fourth Military Medical University

summary: The high mortality caused by severe COVID-19 poses challenges to public health. However, the pathogenesis of severe cases remains a puzzle. Here, we find that SARS-CoV-2 infection boosts CD147 inducible up-regulation, causing persistent virus infection and severe pathological lesions. Specifically, inducible expression of CD147 is transcriptionally regulated by the aryl hydrocarbon receptor (AHR) upon virus infection, while membrane-bound ACE2 decreases, thus indicating that CD147 is a predominant receptor responsible for persistent virus infection. Meanwhile, SARS-CoV-2 infection triggers immune imbalance by promoting cell death of CD4+ T and B cells and abnormal cell communications in rhesus macaque model. Meplazumab, a humanized CD147 antibody, effectively inhibits virus entry and cytokine level, and restores immune balance. We further resolve the cryo-EM structure of CD147-spike complex, and validate five pairs of residues at the interaction interface, which is blocked by Meplazumab via steric hindrance effect. Our findings uncover the pathogenesis of severe COVID-19.

sample_organism:
Macaca mulatta
Mus musculus

platform_organism:
Macaca mulatta
Mus musculus

overall_design: The rhesus macaques (n=16) were challenged with 106 TCID50 SARS-CoV-2 (Beta strain) by intratracheal routes at 0 dpi and divided into four groups. At 1 dpi, the rhesus macaques were injected intravenously with MPZ or 3E8 antibody. The groups were listed as follows: Ctrl group (PBS), MPZ group (MPZ, 10 mg/kg), 3E8 group (3E8, 10 mg/kg), and MPZ+3E8 group (MPZ, 5 mg/kg; 3E8, 5 mg/kg). At 7 dpi, the rhesus macaques were euthanized to collect the lung tissues for scRNA-seq and spatial transcriptomics analysis. Meanwhile, the lung tissues from four healthy rhesus macaques were used as the control. The hCD147 mice (n=4) were also challenged with 105 TCID50 SARS-CoV-2 (Beta strain) intranasally at 0 dpi. At 3 dpi, the mice were euthanized to collect the lung tissues for scRNA-seq and spatial transcriptomics analysis.

first: Zhi-Nan

last: Chen

first: Huijie

last: Bian

first: Liang

first: Ke

last: Wang

first: Qiang

last: Huang


title: Inducible CD147 up-regulation boosts persistent SARS-CoV-2 infection triggering severe COVID-19 independent of ACE2 [Spatial Transcriptomics]


title: Long non-coding RNA RAB30-DT promotes the progression of hepatocellular carcinoma by enhancing cancer cell stemness

city: Shenzhen City

first: lantian

last: zhang

state: Guangdong Province

address: Haiyuan Road, Futian District

institute: The University of Hong Kong-Shenzhen Hospital

summary: Hepatocellular carcinoma (HCC) is a common malignant tumor with a high mortality rate, and improved treatment strategies and better prognostic markers are conducive to the treatment of HCC. Long non-coding Rnas (lncRNAs) have emerged as key regulators of cancer development. However, the exact impact of lncRNAs on the development and progression of HCC remains unclear. Our previous bioinformatics analysis found that RAB30-DT was closely related to the occurrence and development of HCC. In order to study the molecular mechanism of lncRNA RAB30-DT's regulation of HCC occurrence and development, SK-Hep1 HCC cell line with stable lncRNA RAB30-DT knockdown was constructed and bulk RNA-seq sequencing was performed. Analysis results: Knocking down lncRNA RAB30-DT can affect pathways related to tumor stemness and DNA damage, revealing the molecular mechanism of lncRNA RAB30-DT's regulation of HCC occurrence and development, and providing a new target for HCC treatment.

overall_design: Knockdown of lncRNA RAB30-DT was performed on the SK-Hep1 cell line, and RNA was extracted from both the control and knockdown groups of SK-Hep1 cells for RNA-seq analysis.


title: Transcriptome Profiling of Fast-Glycolytic and Slow-Oxidative Muscle Fibers in Chickens under Short- and Long-Term High-Fat Diets

city: chengdu

first: donghao

address: No. 211, Huimin Road, Wenjiang District, Chengdu City

institute: sichuan agricultural university

summary: Skeletal muscle fiber composition influences meat quality and metabolic regulation in poultry, yet fiber-type–specific responses to high-fat diet (HFD) remain poorly understood. This study examined the effects of short-term (SHFD) and long-term (LHFD) HFD feeding on fast/glycolytic (pectoralis major, PEM) and slow/oxidative (soleus, SOL) muscles in Guangyuan grey chickens. Histological analysis showed SOL fibers underwent oxidative-to-glycolytic transition under LHFD, accompanied by mitochondrial disruption. PEM accumulated more lipids and showed early metabolic stress. Transcriptome profiling revealed 3840 differentially expressed genes between PEM and SOL, with 1761 constitutively different. Under SHFD, SOL activated protective pathways including PPAR and autophagy, while PEM showed limited adaptation. LHFD induced further downregulation of fatty acid metabolism and structural genes (e.g., ALPK1, CA12, PM20D2, SLC27A1, and GADD45G) in PEM, but SOL maintained or enhanced expression of genes (e.g., NR4A3, PRKAG2, NOCT, PPARA, and SLC6A6) involved in muscle organization and lipid processing. Temporal clustering highlighted progressive divergence in transcriptional responses. These results suggest SOL fibers exhibit greater resilience to lipid overload than PEM fibers. Our findings provide insight into the molecular basis of muscle-type–specific adaptations to dietary fat and offer targets for improving metabolic health and meat quality in poultry.

overall_design: This study aimed to investigate the muscle-type–specific transcriptional responses to different durations of high-fat diet (HFD) feeding in Guangyuan Grey chickens. A total of 270 150-day-old hens with similar body weights were randomly assigned to three dietary groups: long-term high-fat diet (LHFD, 60 days), short-term high-fat diet (SHFD, 15 days), and a normal control diet (CONT). Each group included 9 replicates with 10 birds per replicate. At 210 days of age, nine birds per group were randomly selected and humanely euthanized after 12 hours of fasting. Immediately postmortem, the pectoralis major muscle (PEM) and soleus muscle (SOL) were dissected from the same birds. Tissue blocks (~0.5 × 0.5 × 1.0 cm) were excised from the ventral (skin-facing) side of PEM and from the widest region of SOL, excluding the superficial layer. Each sample was divided into two portions: one was fixed for histological analysis, and the other was rapidly snap-frozen in liquid nitrogen and stored at −80 °C for RNA extraction and transcriptomic analysis. PEM is composed primarily of fast-twitch glycolytic fibers, whereas SOL contains a mixture of oxidative and glycolytic fibers, making them ideal models for studying muscle fiber type–specific metabolic adaptations. RNA sequencing was conducted to compare gene expression profiles across the six sample groups (CONT-PEM, SHFD-PEM, LHFD-PEM, CONT-SOL, SHFD-SOL, LHFD-SOL). This design allowed for the identification of core transcriptional differences between muscle types and diet-induced transcriptional remodeling, particularly focusing on lipid metabolism, mitochondrial function, and fiber-type transformation.


title: C13 bacterial consortium is sensitive to C. jejuni infection which impacts host-derived miRNAs targeting bacterial and host genes

city: Gainesville

first: Christian

last: Jobin

state: Florida

department: Medicine

institute: University of Florida

summary: C. jejuni infection altered C13 bacterial composition and modified host extracellular vesicle (EV)-derived miRNA expression profile, associated with changes in microbial virulence gene expression. Host-derived miRNAs may be an important component of host-microbiome communication.

overall_design: Three groups of Germ Free (GF) Il10-/- mice were used in this study: 1) mice colonized with a defined consortium of 13 bacterial isolates (C13) representing the four most prominent phyla in the mouse gut.  2) C13 plus Campylobacter jejuni (C. jejuni) 81-176 and 3) GF alone.

first: Siddhi

last: Chitre

first: Raad

last: Gharaibeh


title: ILC1 quiescence confers protection against MCMV infections during undernutrition

city: Akita

first: Megumi

last: Tatematsu

department: Department of Medical Biology

institute: Akita University Graduate School of Medicine

summary: In the liver, group 1 innate lymphoid cells (ILCs) comprise 10–20% tissue-resident ILC1 cells and 80–90% conventional NK cells. Both cells contribute to early defense against virus infection by secreting IFN-γ. However, the distinct role of ILC1 cells in viral infections remains incompletely understood. Here, we identified that ILC1 cells outnumbered NK cells, constituting 80–90% of group 1 ILCs in the liver during undernutrition. Mechanistically, undernutrition significantly reduced NK cell numbers, while hepatic ILC1 cells entered a quiescent state that allowed them to survive in an mTORC1-dependent manner. Metabolically, quiescent ILC1 cells exhibited greater glucose uptake than NK cells and efficiently utilized it via oxidative phosphorylation, which was induced by mTORC1. Finally, ILC1-deficient mice succumbed to MCMV infection under undernourished conditions, but not when fed ad libitum. These findings suggest a non-redundant function of hepatic ILC1 cells in protecting the host against MCMV infection during undernutrition.

overall_design: RNA-seq profiling of NK cells and ILC1 cells from wild type mice and Rptor flox Ncr1-Cre mice fed ad libitum or calorie-restricted.


title: TET3 is a regulator and can be targeted for the intervention of myocardial fibrosis [ChIP-seq]

city: Nanjing

first: yujia

last: xue

address: Longmian Avenue 639

institute: China Pharmaceutical University

summary: Cardiac fibrosis as an integral part of the wound healing process on the one hand safeguards myocardial homeostasis but on the other hand contributes to adverse cardiac remodeling and loss of heart function eventually leading to heart failure (HF). Resident cardiac fibroblasts are the principal source of myofibroblasts that produce extracellular matrix proteins to mediate cardiac fibrosis. We investigated the involvement of ten-and-eleven translocator 3 (TET3) in this process focusing on the regulatory mechanism and translational potential. We report that TET3 depletion in cultured cardiac fibroblasts blocked transition to myofibroblasts in response to different pro-fibrogenic stimuli. Consistently, deletion of TET3 from quiescent or activated fibroblast (myofibroblast) attenuated cardiac fibrosis and rescued heart function in models of cardiac injury. Importantly, a small-molecule TET3-specific degrader Bobcat339 displayed therapeutic potential by mitigating cardiac fibrosis and normalizing heart function when administered post-surgery. Integrated transcriptomic analysis (RNA-seq and CUT&Tag-seq) identified the mechanosensor Piezo2 as a downstream target for TET3. Piezo2 deletion or inhibition dampened fibroblast activation in vitro and ameliorated cardiac fibrosis in vivo. Mechanistically, Piezo2 promoted fibroblast activation by modulating the activities of mechanosensitive transcription factors. Finally, relevance of the TET3-Piezo2 axis was verified in heart specimens collected from HF patients. Our data demonstrate that TET3 is a pivotal regulator of cardiac fibrosis and can be potentially targeted for the intervention of heart failure.

overall_design: chromatin immunoprecipitaion sequencing(chip-seq) with CST JunB(C37F9) in mouse primary cardiac fibroblast.

first: Chenghao

first: Wenxuan

last: Hong

first: Yuwen

first: Yujia

last: Xue

first: Zemin

last: Fang

first: Dingsheng

last: Jinag

first: Yong

last: Xu

first: Ming

last: Kong


title: Targeting Pre-existing Club-Like Cells in Prostate Cancer Potentiates Androgen Deprivation Therapy

first: Gilles

last: Laverny

address: 1 rue Laurent fries

country: France

institute: IGBMC

summary: A critical knowledge gap in prostate cancer research is understanding whether castration-tolerant progenitor-like cells that reside in treatment-naïve tumors play a direct role in therapy resistance and tumor progression. Herein, we reveal that the castration tolerance of LSCmed (Lin-, Sca-1+, CD49fmed) progenitor cells, the mouse equivalent of human prostatic Club cells, arises not from intrinsic properties but from significant transcriptional reprogramming. Utilizing single-cell RNA sequencing of LSCmed cells isolated from prostate-specific Pten-deficient (Ptenpc-/-) mice, we identify the emergence of castration-resistant LSCmed cells enriched in stem-like features, driven by the transcription factor Fosl1/AP-1. We demonstrate that cells exhibiting Ptenpc-/- LSCmed characteristics are prevalent in the aggressive mesenchymal stem-like prostate cancer (MSPC) subtype recently identified in human castration-resistant prostate cancer (CRPC). Furthermore, our findings show that the dual-targeting agents JQ-1 and CX-6258—focused on Fosl1/AP-1 and PIM kinases, respectively—effectively suppress both the progenitor properties and the growth of mouse and human MSPC surrogates in vitro and in vivo. Thus, early eradication of castration-tolerant Club-like cells presents a promising therapeutic strategy to mitigate prostate cancer progression toward CRPC.

overall_design: Two groups of experimental mice (n=8 each) were castrated and after 4-day recovery, we started treatment with JQ-1 (daily) and CX-6258 (bi-weekly) versus vehicle (Fig. 6C). After 28 days, mice were euthanized, then prostates were harvested, weighed, and processed for histopathological analyses or functional assays. Two mice per condition were used to perform digital spatial profiling using the GeoMx.

first: Vanessa

last: Friedrich

first: Jacques-Emmanuel

last: Guidotti

first: Vincent

last: Goffin

first: Tao

last: Ye


title: Expanding the clinical and genetic spectrum of GLUL-related developmental and epileptic encephalopathy

city: Seoul

first: Seungbok

last: Lee

address: 101, Daehak-ro, Jongno-gu

department: Department of Genomic Medicine

institute: Seoul National University Hospital

summary: We presented analyses of the GLUL-related developmental and epileptic encephalopathy with detailed clinical descriptions and identified novel pathogenic variants. Comparative analysis of genotypes and phenotypes revealed the diverse nature of the disease, expanding our knowledge about the genetic and clinical spectrum of GLUL-related disorder.

overall_design: RNASeq for a muscle specimen of a patient with a novel GLUL variant


title: Identification of a small-conductance mechanosensitive channel that plays a role in L-glutamate export in Bacillus methanolicus

city: Trondheim

first: Luciana

last: Fernandes de Brito

address: Kjemi 3, 3.237, Gløshaugen, Sem Sælandsvei 6/8

department: Department of Biotechnology and food science

country: Norway

institute: Norwegian University of Science and Technology

summary: Several small-conductance mechanosensitive channels (MscS) are well-researched l-glutamate exporters in bacterial workhorses employed in industrial biotechnology. Bacillus methanolicus, a methylotrophic bacterium, holds promise as a sustainable platform for l-glutamate bioprocesses utilizing methanol as a feedstock. Our research on B. methanolicus MGA3 identifies its MscS as the sole mechanosensitive channel in this organism and a key exporter of the amino acid l-glutamate, providing valuable insights into its potential for industrial applications. Transcriptomic analysis of B. methanolicus wild type cultured on an l-glutamate production medium revealed downregulation of fatty acid biosynthesis genes, suggesting that fatty acid metabolism is influenced by l-glutamate overproduction, with consequent changes in membrane fluidity likely driving mechanosensitive channel-mediated l-glutamate efflux. The MscS-like channel in B. methanolicus shares structural and functional similarities with MscS in Escherichia coli and Corynebacterium glutamicum. In silico structural predictions show that MGA3 MscS forms a homoheptameric structure with a transmembrane TM-barrel, resembling that of E. coli MscS. The opening mechanism of the channel, driven by membrane dynamics, involves coordinated rotation and flipping of its transmembrane helices, with variations in lipid composition potentially influencing the channel’s activity. Additionally, excess supplementation of biotin, a crucial coenzyme for fatty acid biosynthesis’ carboxylases, led to reduced l-glutamate biosynthesis in MGA3. Finally, metabolic engineering experiments inducing MscS gain- and loss-of-function further confirmed the channel’s critical role in B. methanolicus amino acid production, proportionally enhancing and reducing l-glutamate efflux, respectively. These findings open doors to novel strategies for engineering B. methanolicus and related methylotrophic organisms for efficient production and export of amino acids and other value-added products.

overall_design: B. methanolicus MGA3 cultivated in MVcM vs. MVcMii media

first: Davide

last: Luciano

first: Marta

last: Irla

first: David

last: Virant

first: Gaston

last: Courtade

first: Trygve

last: Brautaset


title: Bulk RNA sequencing of prostate tissues from BPH model mice and treatment groups

city: Beijing

first: Zheng

last: Li

state: Beijing

address: No. 8 Xishiku Street, Xicheng District

institute: Peking University First hospital

summary: We performed bulk RNA sequencing on fresh prostate tissue serial sections from five randomly selected mice per group.

overall_design:
Differential expression analysis for BPH vs. Control (Figure 6i), BPH + rmCXCL13 vs. BPH (Figure 6k), and BPH + rmCXCL13 + anti-CD4 vs. BPH + rmCXCL13 (Figure 6m) highlighted significant changes in senescence-related genes.
Prostate tissues from BALB/c mice were divided into Control, BPH, BPH + rmCXCL13, and BPH + rmCXCL13 + anti-CD4 groups. Each group contained five mice. RNA was extracted and subjected to bulk RNA-seq. Due to technical issues, raw sequencing files were lost. Processed expression matrices are available and included in this submission.

***************************************************************
Submitter note: Raw sequencing data are not available due to storage limitations
***************************************************************

first: XiaofeI

last: Wang

first: Zhifu

last: Liu

first: Senmao

first: Shuai

last: Hu

first: Wei

last: Yu


title: Gene expression profiling of senescent and non-senescent BPH epithelial cells

summary: RNA-seq analysis of human BPH prostate epithelial cells classified by SA-β-gal staining status.

overall_design: Epithelial cells were isolated from human BPH tissue and divided into senescent (SA-β-gal+) and non-senescent (SA-β-gal−) groups. RNA-seq was performed and gene expression profiles were compared.

first: Frozen tissue sections from human BPH prostate samples were serially sectioned. The first section was subjected to SA-β-gal staining to identify senescent (positive) and non-senescent (negative) regions. The adjacent second section was then used for microdissection to isolate epithelial cells from the corresponding regions based on their senescence status. Total RNA was extracted from the microdissected senescent and non-senescent epithelial cells

last: followed by RNA-seq to profile transcriptomic differences.

first: Xingxing

last: Tang

first: Jie

last: Jin


title: PURPL plays a role in epigenetic regulation of gene expression [ChIP-Seq]

last: Sun

summary: Cellular senescence involves heterochromatin reorganization and aberrant gene expression. We demonstrate that the nuclear-enriched lncRNA PURPL regulates senescence-associated chromatin dynamics by modulating H3K9me3 deposition. PURPL knockdown increased global H3K9me3 levels while its overexpression reduced this repressive mark. Genome-wide analysis revealed 411 loci with PURPL-dependent H3K9me3 changes, including the aging-related gene SERPINE1, where H3K9me3 accumulation correlated with transcriptional repression. These findings establish PURPL as an epigenetic regulator of senescence through targeted control of heterochromatin formation.

overall_design: ChIP-seq of CRISPRi-mediated PURPL knockdown and control cell lines.

first: Xiao

last: Yang


title: STING-induced programmed necrosis modulates the collagen-rich immunosuppressive microenvironment in meningiomas [RNA-seq]

city: Chicago

first: Mark

last: Youngblood

state: IL

address: 676 N. St. Clair Street; Suite 2210

department: Department of Neurological Surgery

institute: Northwestern University

summary: Up to one-quarter of meningiomas recur within five years of resection; however, there are currently no established adjuvant therapies for tumors that are intractable to surgery and radiation. Using integrated epigenetic and transcriptional profiling, we identify widespread expression of STING across multiple cell populations within the meningioma microenvironment, associated with increased chromatin accessibility in neoplastic cells and promoter hypomethylation. Ex vivo single-cell suspensions of resected human meningiomas treated with the STING agonist 8803 induced direct cytotoxicity within 72 hours, including even in the absence of CD45+ immune cells. Pharmacologic inhibitors and RNA-sequencing revealed the association of programmed necrotic cell death pathways that require reactive oxygen species production with STING agonist8803 efficacy, which also required reactive oxygen species production. Proteolytic maturation of the pore-forming protein gasdermin D (GSDMD) on meningioma cancer cells contributeds to 8803-induced cytotoxicity, and  the suggested release of pro-inflammatory damage-associated molecular pattern molecules that further upon 8803-induced cytotoxicity, which further potentiatedpotentiates STING activation of innate immune cell activation and responses through toll-like receptors activation.  Across four different preclinical murine meningioma models, 8803 reduced tumormeningioma volume, and survival significantly increased survival was significantly improved after treatment of across three orthotopic tumorsmodels. These therapeutic effects arewere Treatment with 8803 across multiple pre-clinical orthotopic meningioma models exhibited improved survival associated with marked immunological tumor infiltration and cytotoxicity, tumor stromal remodeling through macrophage elaborated metalloproteinases, and downregulation of the TIM3 and LAIR1 widespread cytotoxicity and immunologic infiltration during the therapeutic window, including down-regulation of severalthe dominant mechanisms of meningioma-mediated immune suppression. , such as TIM3 and LAIR1, alongside the LAIR ligand collagen through macrophage elaborated MMP production. Our results reveal widespread synergistic expression of STING in neoplastic and myeloid populations of meningiomas, identify novel mechanisms of STING signalingcytotoxicity inthrough tumor cells, andsuggest a role for STING activation in degradation of collagen-rich tumor microenvironments the impact on the extracellular matrix,  and clarify a new treatment indication for the STING agonist 8803.

overall_design: Ex-vivo, patient derived meningiomas were processed as single cell suspensions post resection. RNA-seq profiling was applied to each patient sample with both a control treatment, and an 8803 STING-agoinst treatment

middle: W

first: Shashwat

last: Tripathi

first: Hinda

last: Najem

first: Harrshavasan

last: Congivaram

first: Mateo

last: Gomez

first: Amy

middle: B

last: Heimberger


title: Transcriptome profiling of senescent and non-senescent BPH epithelial cells after CD4⁺ T cell co-culture

summary: Human primary CD4⁺ T cells were co-cultured with either senescent or non-senescent primary BPH epithelial cells for 6 hours. BPH epithelial cells were then isolated and subjected to RNA sequencing. The experiment was repeated three times independently.

overall_design: Primary human CD4⁺ T cells were co-cultured for 6 hours with senescent or non-senescent BPH epithelial cells, which were pre-identified by SA-β-gal staining. Following co-culture, epithelial cells were isolated and subjected to transcriptome profiling (bulk RNA-seq). The study included three biological replicates per group.

first: Xiaofei


title: STING-induced programmed necrosis modulates the collagen-rich immunosuppressive microenvironment in meningiomas [scMulti-omics]

overall_design: Single nucleus multiome (RNA-seq + ATAC-seq) profiling of frozen meningioma specimens post resection.


title: Expression data from rat skeletal muscle at 0 and 48 hours of postmortem interval

city: Ciudad de México

first: Mariano

last: Guardado Estrada

state: Ciudad de México

address: Circuito de la Investigación SN

country: Mexico

institute: UNAM

summary:
Accurate estimation of the postmortem interval is critical in forensic investigations to determine the time since death. While traditional methods offer limited precision, molecular approaches based on gene expression have shown promise. mRNA retains detectable and quantifiable expression changes that reflect biological processes occurring after death. In this study, we analyzed the transcriptome of rat skeletal muscle at 0 hours (control) and 48 hours of post mortem interval to identify differentially expressed mRNAs that may serve as potential biomarkers, aiming to enhance the accuracy and reliability of PMI estimation in forensic science.
Microarray analysis using the Affymetrix Clariom S Rat array (Affymetrix) facilitated a comprehensive overview of the skeletal muscle transcriptome at 48 hours of postmortem interval. This platform allowed for the simultaneous quantification of thousands of transcripts, revealing widespread gene expression alterations associated with the postmortem interval. The resulting transcriptomic profile not only provided insight into molecular changes occurring after death but also highlighted specific mRNAs as potential biomarker candidates for PMI estimation in forensic contexts.

overall_design: This study aimed to characterize global transcriptomic changes in rat skeletal muscle at 0 hours (control) and 48 hours of postmortem interval. A total of 10 adult male Wistar rats were divided into the two experimental groups. Rats in the 48 h-PMI group were maintained at 25 °C in a climatic chamber until tissue collection, whereas control rats were sampled immediately after euthanasia. External and internal postmortem signs were documented for all animals. 200 mg of femoral muscle was collected from each rat and total RNA was extracted using TRIzol™ reagent and evaluated for concentration and integrity. Transcriptomic profiling was performed using the Affymetrix Clariom S Rat Gene Expression Array. For each sample, 10 ng of total RNA was amplified and biotin-labeled using the GeneChip™ 3´ IVT Pico Kit. Arrays were hybridized, washed, stained, and scanned using standard Affymetrix workflows.


title: Cancer therapy induced effects on lung organoids as suitable in vitro model II

city: Essen

first: Bettina

last: Budeus

address: Virchowstrasse 173

department: Institute for Cell Biology

country: Germany

institute: University Hospital Essen

summary: Generation of lung organoids as model for genotoxic stress

overall_design: iPSC-derived lung organoids were either treated with radiation or not prior RNAseq analysis

first: Chiara

last: Kroepel

first: Lisa-Marie

last: Stasch

first: Diana

last: Klein


title: Protamine expression in somatic cells condenses chromatin and disrupts transcription without altering DNA methylation

city: Aachen

first: Deepika

last: Puri

address: Pauwelsstraße 20

institute: Helmholtz Institute For Biomedical Engineering, Uniklinik Aachen

type: Methylation profiling by genome tiling array

summary: Protamines play a crucial role in nuclear condensation during spermiogenesis, a process associated with significant chromatin remodeling and replacement of histones by protamines. While much research has focused on the function of protamines in sperm development and fertility, their effects in non-sperm cells remain largely unexplored. In this study, we investigated the impact of overexpressing murine and human protamine 1 and 2 (PRM1 and PRM2) on nuclear architecture, histone eviction, DNA methylation, and transcription in HEK293T cells and mesenchymal stromal cells (MSCs). Overexpression of protamines resulted in nuclear condensation and particularly PRM1 showed notable enrichment in nucleoli and cycle abnormalities. Immunofluorescence staining indicated a significant reduction in specific histone modifications (H3K9me3, H3K4me1, and H3K27Ac) in response to protamine expression, especially in MSCs. Interestingly, despite these changes in nuclear organization, the methylome remained largely stable upon protamine expression. However, particularly expression of protamines significantly diminished transcription, especially of the ribosomal genes upon PRM1 expression. Our studies indicate that PRM1 and PRM2 condense distinct genomic regions in somatic cells, resulting in widespread silencing of gene expression.

overall_design: HEK293T cells were transfected with Empty vector, human PRM1-EFGP and  human PRM2-EGFP. 3 days after transfection cells were sorted and high EGFP cells were used to isolate DNA and Illumina beadchip Epic Array Analysis was performed

first: Alexandra

last: Bott

first: Monica

last: Varona Baranda

first: Esra

last: Dursun Torlak

first: Gina Esther

last: Merges

first: Hubert

last: Schorle

first: Wolfgang

last: Wagner


title: A Stem and Progenitor Cell-Derived Gene Expression Signature is Prognostic for Survival in Myelofibrosis [RNA-Seq]

first: Alexander

last: Murison

summary: We hypothesized that transcriptomic features among disease-driving hematopoietic stem and progenitor cells (HSPC) could improve the current paradigm for risk stratification in myelofibrosis (MF). Therefore, we performed bulk RNA-seq on blood from 358 MF patients split into training and test cohorts (ClinicalTrials.gov Identifier: NCT02760238). Single-cell (sc) RNA-seq data from Lin-CD34+ MF HSPCs were used to guide the development of a prognostic model trained on the bulk RNA-seq data. A NanoString assay was used to validate our final model in two independent cohorts of MF patients. We identified a 24-gene weighted-sum expression score (termed MPN24) that was prognostic of overall survival (OS). In two independent cohorts of 170 and 99 MF patients, MPN24-High scoring patients had worse 5-year OS compared to MPN24-Low scoring patients (25.5% vs 86.9%, HR = 9.81, P = 4.0e-11 and 23.5% vs 75.9%, HR = 6.40, P = 1.8e-7, respectively). MPN24-High was also associated with clinical, mutation and karyotypic features known to confer adverse risk in MF. The MPN24 score retained independent prognostic significance in multivariable analysis incorporating these covariates as well as existing risk stratification models including DIPSS, DIPSS Plus, MIPSS70, and MIPSS70 Plus v2.0, adding significant prognostic value to these baseline risk stratification models (P = 1.6e-5, 4.0e-6, 1.5e-6, 5.4e-4, respectively). The MPN24 score was particularly useful in improving risk-stratification of DIPSS-Intermediate-1 and Intermediate-2 and MIPSS70 Intermediate and High-risk patients. An HSPC-derived gene expression score is associated with overall survival independent of existing clinical and genomic prognostic variables and improves risk stratification in MF.

overall_design:
We profiled the peripheral blood mononuclear cells (PB-MNCs) from 358 myelofibrosis patients (PMc1 cohort) and correlated transcriptomic profiles to overall survival to derive a 24-gene prognostic score (termed, MPN24). We benchmarked the ability of a custom NanoString-based assay to measure the 24-gene score against RNA-seq in 59 PMc1 patient samples. Additionally, we validated MPN24 using the NanoString assay in PB-MNCs from two independent cohorts consisting of 170 (PMc2 cohort) and 99 (MSK cohort) myelofibrosis patients.

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

first: Jessie

last: Medeiros

first: Andy

last: Zeng

first: Michelle

last: Chan-Seng-Yue

first: Hyerin

last: Kim

first: Tristan

last: Woo

first: Suraj

last: Bansal

first: Jessica

last: McLeod

first: Sagar

last: Kothari

first: Andrea

last: Arruda

last: Tsui

first: Jaime

last: Claudio

first: Dawn

last: Maze

first: Hassan

last: Sibai

first: Mark

last: Minden

first: James

last: Kennedy

first: Christopher

last: Famulare

first: Raajit

last: Rampal

first: Jean

last: Wang

first: John

last: Dick

first: Vikas

last: Gupta


title: Asthma and Allergy Array DNA methylation in Child Buccal Swabs at 5 Years of Age

city: Beaverton

first: Lyndsey

middle: Elizabeth

last: Shorey-Kendrick

state: Oregon

address: 505 NW 185th Ave

institute: Oregon Health and Science University

summary: We previously reported improved respiratory outcomes in babies born to pregnant smokers supplemented with vitamin C (500 mg/day) versus placebo in a randomized clinical trial.  Improved respiratory outcomes persisted to 5 years of age and were associated with buccal DNA methylation (DNAm) measured using the InfiniumMethylationEPIC array. The objective of this study was to examine associations of vitamin C treatment and lung function with buccal DNAm at asthma and allergy loci not covered by the EPIC platform.

overall_design:
We profiled DNAm at 36,999 CpGs in loci previously associated with asthma or allergic diseases using custom content Asthma&Allergy arrays in 137 subjects (65 placebo; 72 vitamin C) with pulmonary function testing (PFT) at the 5-year visit. We examined the association of buccal DNAm with 1) vitamin C treatment vs placebo, 2) forced expiratory flow between 25-75% of expired volume (FEF25-75) and 3)  wheeze at 4-6 years of age.
The data submitted is part of a double-blind randomized clinical trial and so the patients and investigators are still blind to treatment assignment.

first: L

first: C

middle: T

last: McEvoy

first: R

last: Tepper

first: E

last: Spindel


title: Isolation and immortalization of human portal vein endothelial cells: a novel research tool for investigating splanchnic vasculature

city: Barcelona

first: Genís

last: Camprecios

state: Catalonia

address: Rosselló 153

department: Hepatic Hemodynamics

country: Spain

institute: IDIBAPS

summary: Cirrhosis profoundly impacts extrahepatic vasculature, particularly altering the portal venous system (PVS), leading to increased portal pressure, portosystemic collaterals, and portal vein thrombosis, which heightens morbidity and reduced survival in patients with liver disease. Despite extensive research on intrahepatic vasculature in cirrhosis, understanding of extrahepatic changes in the splanchnic territory at the molecular level remains limited. The distinct characteristics of the PVS suggest a unique endothelial profile crucial for elucidating vascular dysfunction in liver diseases. However, current research is hampered by the inaccessibility of the portal vein and imperfect preclinical models. Here, we aim to isolate and immortalize primary human portal vein endothelial cells (hPVECs) to enhance understanding of pathophysiological changes during liver disease and establish a platform for future drug testing.

overall_design: hPVECs and inferior cava vein (ICV) endothelial cells (ECs) were isolated from human portal vein or ICV, obtained during hepatic transplantation, using trypsinization, mechanical scratching, and FACS. Subsequently, hPVECs were immortalized (iPVECs) with lentiviral particles expressing the SV40 large T-antigen. iPVECs maintained endothelial phenotype and behavior along with specific endothelial gene expression profiles distinct from systemic ECs. We then expanded the obtained cells and performed bulk RNAseq with hPVEC and hICVEC passage 1 cells iPVEC passage 20 cells. Comparative gene expression analysis was performed between hICVEC vs hPVECs and also compared to iPVEC.

first: Virginia

last: Hernández-Gea

first: Aina

last: Anton

last: Campreciós


title: Activating PIK3CA mutation promotes overgrowth of adipose tissue via inhibiting lipophagy in macrodactyly

city: Shanghai

first: Yating

last: Yin

address: Zhizaoju Road#693

institute: Shanghai ninth people’s hospital

summary: Excessive proliferation and lipid accumulation of adipose tissue are the main pathological alterations in macrodactyly. Our previous studies found that macrodactyly exhibits abnormal lipid metabolism and inhibited autophagy, but the underlying mechanisms remain unclear. This study aims to investigate the regulatory mechanisms of autophagy in macrodactyly. The therapeutic impact and underlying mechanisms of autophagy on lipid accumulation, induced by a gain-of-function mutation of PIK3CA in macrodactyly, were assessed with respect to autophagy, lipid metabolism, oxidative stress, and deubiquitination. Autophagy deficiency resulting from PIK3CA mutation in macrodactyly led to excessive accumulation of adipose tissue. Lipid accumulation can be mitigated by inducing lipophagy of lipid droplets (LDs) in adipose derived stem cells of macrodactyly (Mac-ADSCs). The subsequent increase in free fatty acids (FFA) led to mitochondrial oxidative stress in Mac-ADSCs. Inducing autophagy exacerbated mitochondrial oxidative stress in Mac-ADSCs, thereby contributing to apoptosis. Additionally, the ablation of the deubiquitinase USP15 facilitated the degradation of LDs in Mac-ADSCs, through ubiquitin-dependent macrolipophagy. USP15 inhibitor reduced lipid accumulation in macrodactyly adipose tissue xenografts. In conclusion, activating PIK3CA mutation promotes excessive proliferation and lipid accumulation of Mac-ADSCs by inhibiting lipophagy. Targeted inhibition of USP15 may serve as a promising therapeutic approach for treating macrodactyly.

overall_design: To investigate the pathological alterations associated with macrodactyly, we conducted RNA sequencing (RNA-Seq) analysis on the adipose tissues of polydactyly (Pol-AT) and macrodactyly (Mac-AT).

first: Xiao

last: Zhang

first: Hailei

last: Mao

first: Bin


title: Precise Excision of Expanded GGC Repeats in NOTCH2NLC via CRISPR/Cas9 for Treating Neuronal Intranuclear Inclusion Disease

city: Changsha

first: Yongcheng

last: Pan

state: Hunan

address: 87 Xiangya Rd

institute: Key Laboratory of Hunan Province in Neurodegenerative Disorders, Xiangya Hospital, Central South University

summary: Neuronal intranuclear inclusion disease (NIID) is an adult-onset neurodegenerative disease caused by expanded GGC repeats in the 5’ untranslated region of the human-specific NOTCH2NLC gene. The high sequence similarity between NOTCH2NLC and its paralogs poses a significant challenge for precise gene editing. Here, we develop a CRISPR/spCas9-based gene-editing strategy that precisely excises the expanded GGC repeats in NOTCH2NLC without detectable off-target effects on the highly homologous NOTCH2/NOTCH2NL family genes (<2% sequence divergence at this locus). The efficacy, specificity and safety of this approach are rigorously validated across multiple experimental models, including human cell lines, NIID iPSCs, and our previously established transgenic NIID mouse model. Our results demonstrate that precise excision of the expanded GGC repeats effectively alleviates NIID-related neuropathological, molecular and behavioral abnormalities. This study establishes the proof of concept for genome editing as a therapeutic strategy for NIID and other related repeat expansion disorders.

overall_design: We analyzed transcriptomic changes in mouse tissues and human NPCs pre- and post-gene therapy using RNA-seq.

first: Qiong

last: Liu

first: Nina

last: Xie

first: Beisha

last: Tang


title: Bone marrow and blood demonstrate distinct immune reconstitution patterns and correlations with relapse post-transplant

city: New York

first: Alan

last: Hanash

state: NY

address: 1275 York Ave

institute: Memorial Sloan Kettering Cancer Center

summary: The bone marrow represents the tumor microenvironment for many hematologic malignancies and a potentially critical site for alloimmunity following hematopoietic transplantation. Despite the importance of immune reconstitution (IR) post-transplant, marrow IR data are limited, and insights are largely derived from studies of peripheral blood (PB). We investigated lymphocyte IR longitudinally in marrow (n=110) and PB (n=115) samples from adults undergoing allogeneic transplantation for hematologic malignancies (n=33). This transplant cohort included a diverse representation of graft sources (mobilized peripheral blood, CD34-selected grafts, and umbilical cord blood) and degrees of HLA mismatch. Natural killer (NK) cells quickly expanded within the first 30 days post-transplant in both marrow and PB, but were then outnumbered by T cells in PB after day 100. In contrast, NK cells remained dominant in the marrow at day 100 (p<0.01, paired Wilcoxon signed-rank test), and thereafter marrow T and NK cell frequencies were similar throughout year-one. Tissue-specific features post-transplant included fewer regulatory T cells, more innate lymphoid cells, and increased CD69 expression on lymphocytes in marrow compared to PB. Furthermore, day 100 PD1 expression on marrow T cells was greater in non-relapsing patients than those who subsequently relapsed. These findings reveal persistent NK dominance of the marrow early post- transplant and suggest correlations between marrow immunity and clinical transplant outcomes.

overall_design:
Bone marrow was obtained 100 days after allogeneic stem cell transplantation. We sorted live, CD45+ cells and performed single-cell RNA sequencing. Using these data we profiled lymphocyte populations present in the bone marrow in the two patients, including  host/donor chimerism analysis at the single-cell level.
*******************************************************
Raw data not provided due to patient privacy concerns
*******************************************************

first: Susan

last: DeWolf

first: Jason

last: Kuttiyara

first: Paola

last: Vinci

first: Teng

last: Fei

first: John

last: Slingerland

first: Zoe

last: Katsamakis

first: Joshua

last: Fein

first: Brianna

last: Gipson

first: Rachel

last: Lorenc

first: Victoria

last: Zinsmeyer

first: LeeAnna

last: Marcello

first: Paul

last: Giardina

first: William

last: Donohoe

first: Gunjan

last: Shah

first: Richard

last: Lin

first: Esperanza

last: Papadopoulos

first: Boglarka

last: Gyurkocza

first: Brian

last: Shaffer

first: Martin

last: Tallman

first: Ioannis

last: Politikos

first: Sergio

last: Giralt

first: Juliet

last: Barker

first: Miguel-Angel

last: Perales

first: Roni

last: Tamari

first: Omar

last: Abdel-Wahab

first: Christina

last: Cho

first: Katharine

last: Hsu

first: Jonathan

middle: U

last: Peled

first: Marcel

last: van den Brink

middle: M


title: Ischemic stroke of different etiologies undergoing mechanical thrombectomy

city: Pamplona

first: Josune

last: Orbe

address: Avenida Pio XII 55

institute: CIMA

summary: Accurate ischemic stroke (IS) etiologic diagnosis provides an appropriate secondary prevention strategy and better outcome. Currently, 45% of IS are cryptogenic urging to find novel approaches to enhance diagnostic precision and secondary stroke prevention. OBJETIVE: To study the transcriptomic content of thrombus extracellular vesicles (EVs) to delineate the molecular profile and potential biomarkers of IS etiologies. RESULTS:30 consecutive patients with moderate or severe ischemic stroke (IS) and different IS etiologies were included to perform the transcriptomic analysis of thrombus EVs comprising: atherothrombotic (AT, n=10), Cardioembolic (CE, n=10), and embolic stroke of undetermined source (ESUS, n=10). The RNASeq analysis of EVs detected clusters of differentially expressed genes (DEGs) related to IS etiologies.

overall_design: Transcriptome of patients with ischemic stroke of atherothrombotic (AT, n=10), cardioembolic (CE, n=10) or undetermined etiology (ESUS, n=10) were compared in thrombus extracellular vesicles .

first: J

first: FJD

last: Machado

first: C

last: Roncal

first: M

last: Herrera


title: A spatiotemporal atlas of orchiectomy-induced modulation of cellular composition and gene expression in the mouse prostate

city: Ann Arbor

first: Evan

last: Keller

state: Michigan

address: 1109 Geddes Avenue

department: Urology

institute: University of Michigan

summary: Mice models are commonly used to further our understanding of the function of the prostate and response to androgen depletion. We used both spatial transcriptomics and single cell transcriptomics to determine how the mouse prostate degrades after castration. We identified changes in each of the prostate lobes that couuld be tracked over time as well as changes in the immune profile and of the composition of fibroblasts within the prostate. In addition to several previously identified fibroblast subtypes that we were able to show were altered in response to androgen depletion, we identified a novel fibroblast subtype present only in the castrated prostate.

overall_design: Four serial sections of prostates from castrated mice 10 and 20 days after castration were sequenced by 10x Visium to test for differential spatial gene expression after castration.

first: Greg

last: Shelley

first: May

last: Alison

first: Allison

last: May

first: Jinlu

last: Dai

first: Sethu

last: Pitchiaya

first: Arvind

last: Rao