title: Graded constitutive NF-κB signaling in B cells drives SLL/CLL-like lymphomagenesis and overcomes microenvironment dependence [M002]

city: Munich

first: Marc

last: Schmidt-Supprian

state: Bavaria

address: Einsteinstr. 25

department: Tranlational Cancer Research

institute: TUM

summary: Aberrant activation of NF-κB transcription factors is a hallmark of human lymphoma. Many lymphoma- as well as microenvironment-associated alterations mediating enhanced NF-κB signaling occur upstream of the IκB Kinase complex and its key kinase IKK2, therefore affecting additional pathways. Here, we specifically investigated the effects of graded canonical NF-κB activation in mouse B cells, induced through the expression of one or two copies of a constitutively active IKK2 variant (IKK2ca). Strong canonical NF-κB signaling drives an early expansion of B1a cells, culminating in lethal lymphomagenesis with complete penetrance. These B cell malignancies resemble human small lymphocytic lymphoma (SLL) and chronic lymphocytic leukemia (CLL) with respect to disease course, gene expression and stereotypic B cell receptor clonality. Mice with less pronounced canonical NF-κB activation presented delayed, more heterogeneous lymphomagenesis with lower penetrance, highlighting NF-κB dose-dependent effects. Mechanistically, we show that constitutive IKK2 signals provide a profound cell-intrinsic competitive advantage to B1a cells and strongly synergize with TCL1 overexpression, resulting in a severely accelerated and aggravated CLL-like disease. In addition, strong constitutive NF-κB activation overcomes the critical dependency of TC1tg lymphoma cells on obligate environmental maintenance signals. In conclusion, we provide direct in vivo proof for canonical NF-κB signals as an oncogenic driver in an animal model, and demonstrate reduced tumor microenvironment dependency as a key NF-κB-mediated mechanism in lymphomagenesis. Our findings underscore the pivotal role of this pathway in human SLL/CLL and its potential as a therapeutic target, particularly for aggressive/refractory disease.

overall_design: RNA-seq profiling of B1a cells across four genotypes (Ikk2het, Ikk2hom, TCL1tg, TCL1tgIkk2het) at three time points (3 months, 9 months, and old age) to study tumor development (CLL/SLL) and progression.

first: Valeria

last: Soberón

first: Lena

last: Osswald

first: Andrew

last: Moore

first: Gene

last: Swinerd

first: Jingyu

last: Chen

first: Seren

last: Baygün

first: Carina

last: Diehl

first: Laura

last: Kraus

first: Vanessa

last: Gölling

first: Ricarda

last: Trapp

first: Pankaj

last: Singroul

first: Thomas

last: O'Neill

first: Sabrina

last: Bortoluzzi

first: Daniel

last: Kovacs

first: Tim

last: Ammon

first: Yuliia

last: Hubarzhevska

first: Rupert

last: Öllinger

first: Sebastian

last: Müller

first: Olga

last: Baranov

last: Engleitner

first: Yoshiteru

last: Sasaki

first: Felix

last: Gillhuber

first: Sonja

last: Grath

first: Oliver

last: Weigert

first: Andreas

last: Rosenwald

first: Klaus

last: Rajewsky

first: Katja

last: Steiger

first: Florian

last: Bassermann

first: Roland

last: Rad

last: Krappmann

first: Ingo

last: Ringshausen


title: Transcriptomic profiling of mouse brain in PGRN KO, NesGrnOE-KOBG, EGFL7-PGRNdko and floxed control mice (Grnflfl)

status: Public on Mar 11 2026

city: Frankfurt

first: Irmgard

last: Tegeder

state: Hessen

address: Theodor Stern Kai 7

department: Clinical Pharmacology

country: Germany

institute: Goethe University Frankfurt

type: Expression profiling by high throughput sequencing

summary:
Transcriptomic profiling of mouse brain in old progranulin deficient mice with a complete loss of progranulin (PGRN KO), with restoration of neuronal progranulin (NesGrnOE-KOBG), in progranulin-EGFL7 double deficient mice (EGFL7-PGRNdko) and floxed control mice (Grnflfl)
Genetic progranulin deficiency in humans causes Frontotemporal Dementia (FTD). Progranulin knockout mice (PGRNko) are a model for the disease albeit cognitive impairment in mice is subtle. The predominant FTD-phenotype in mice are hyperactivity, sugar craving, compulsiveness, skin lesions owing to exessive grooming and general health issues in old mice such as anal prolaps. Progranulin in primarily expressed in neurons and in myeloid derived immune cells including microglia. Progranulin deficient microglia are neuron-aggressive and are believed to contribute to excessive synaptic pruning. It is not known if progranulin that is normally neurotrophic and keeps microglia in a neuron-supportive phenotype has to come from within microglia or is a progranulin-signal from neurons to microglia. To dissect out the differential contribution we generated mice which express progranulin in neurons via Nestin-driven restoration of progranulin expression in a progranulin knockout background. In addition we assessed additional deletion of EGFL7, which behaves as progranulin competitor for binding/activation of NOTCH receptors. Hence, this study describes and compared the transcriptome of old mice in four mouse lines: full progranulin knockout (PGRNko), neuronal resoration of PGRN on a knockout background (NesGrnOE-koBG), progranulin & EGFL7 double deletion (EGFL7-PGRNdko) and floxed progranulin control mice (Grn flfl). Mice were 60-70 weeks old at the the time of tissue collection. Total RNA was extracted with Qiagen RNAeasy micro-kits and sequencing librariers were prepared by Novogene according to standard protocols for mRNA sequencing. RNA seq was done on Illumina NovaSeq 6000. The sequence alignment was done with standard proceduers including quality filtering and adapter trimming with Qiagen's CLC Genomic Workbench. The TMM (trimmed mean od M-values) algorithm  was used for read normalization. Total reads, RPKM and TPM are also provided in "Processed data" files.

overall_design: PGRNko, NesGrnOE-knBG, EGFL7-PGRNdko and Grn-flfl mice were observed throughout life in multiple behavioral tests including at the end Intellicage analysis of cognition, social behavior and compulsiveness, excessive grooming. At the end mice were euthanized, the brain was rapidly removed, cerebellum, olfactory buld and brainstem were cut off, the brain was cut sagitally and the two halves were separately snap frozen in liquid nitrogen and then stored at -80°C until RNA extraction (one half).

first: Matthias

last: Klein


title: A regulatory circuit operated by KDELR1 and KDELR3 fine-tunes the composition of the early secretory pathway

city: Milan

first: Jose

middle: Manuel

last: Garcia-Manteiga

address: Via Olgettina 58

department: Center for Translational Genomics and Bioinformatics

country: Italy

institute: OSR - San Raffaele Scientific Institute

summary: KDEL receptors (KDELRs) prevent the secretion of soluble chaperones and enzymes meant to reside in the endoplasmic reticulum. While a single KDELR exists in yeast (ERD2), three isoforms are present in mammals, displaying high sequence similarity (73-83%). However, the phylogenetic conservation of the differences and the diverse tissue distribution of the three KDELRs suggest functional specialization. We show here that, while all three receptors can prevent the secretion of KDEL-bearing clients, KDELR1 and KDELR3 regulate in opposite ways the production of AGR2, a key assistant of mucin folding. AGR2 transcripts increase dramatically upon silencing KDELR3 but decrease when KDELR1 is downregulated. Silencing ERp44, but not other ER residents, phenocopies KDELR3 knockdown, suggesting that AGR2 regulation depends on ERp44-KDELR3 interactions. Our findings identify a novel regulatory circuit that controls the molecular composition of the early secretory pathway based on specific interactions between KDELRs and ER residents.

overall_design: HeLa cells were subjected to silencing of the three human KDELRs or of ERp44 using lipofectamine RNAi max. Five replicates for each condition were used. 72 hours after silencing cells were collected and RNA extracted. Libraries of cDNA were prepared with TruSeq mRNA Library Prep Kit (Illumina) and sequenced using the Novaseq instrument (Illumina) 100 bp single-end up to 30 million reads. After trimming the adapter sequences from reads (Trimmomatic), sequences were aligned to the human hg38 genome using the STAR (v2.5.3a) (https://github.com/alexdobin/STAR/) and counted using feature Counts (http://bioconductor.org/packages/release/bioc/html/Rsubread.html) with the gene annotation from Gencode (version31). The R/Bioconduction package ‘limma’ was used to conduct differential expression analysis using standard procedures (Ritchie et al., 2015).

first: Federica C

last: Palazzo

first: Jose M.

first: Roberto

last: Sitia

first: Tiziana

last: Anelli


title: Over-expression of human mutated APP transgene in the mouse brain induces differential gene expression profiles depending on the age of exposure

city: Bonn

first: Kan

last: Xie

address: Venusberg-Campus 1/99

department: Translational Gerontology

institute: German Center for Neurodegenerative Diseases

summary: Alzheimer´s disease (AD) is the most prevalent neurodegenerative disorder causing about two third of all dementia cases in the elderly. Although AD pathological features have been intensively examined in numerous scientific studies employing invertebrate and vertebrate model organisms, the exact underlying mechanisms remain poorly understood. Extracellular accumulation of amyloid beta (Aβ) peptides represents a pathological hallmark of AD and it has been considered as the initiation point of AD by the popular amyloid hypothesis. In the current study, we aim to address whether expression of a mutated APP transgene (APPSweInd) would provoke differential neurological outcomes depending on the age of exposure in mice. Towards this end, we employed a tetracycline-based inducible genetic system to target APPSweInd transgene expression to defined age intervals initiated at adulthood, i.e., between 6-18 months (= mid-aged group) and 12-24 months (= aged group). Surprisingly, over-expression of mutant APP reduced the transcriptional levels of genes embedded in the cholinergic network exclusively in the aged mouse brain, while gene expression profiles related to inflammatory activation and exacerbated gliosis were shared between age groups. Hence, our data support the notion that AD pathology may rely on a disruption of the cholinergic system that is triggered through an altered response of the aged brain towards toxic Aβ aggregates.

overall_design: Using an inducible tet-off system over-expressing mutant human APP under the control of doxycycline, the brain transcriptomic profiles between mice expressing human APP between 12-24 months of age vs. 6-18 months of age were compared.

first: Devon

last: Ryan

first: Stefan

last: Bonn

first: Dan

last: Ehninger


title: Single-cell RNA sequencing reveals that the immunosuppression in landscape induced by chronic stress in colorectal cancer

city: Shanghai

first: Yingru

last: Zhang

state: Shanghai

address: No. 528, Zhangheng Road, Pudong New Area

country: China

institute: Shanghai University of Traditional Chinese Medicine Shuguang Hospital

summary: The high prevalence of depressive disorders in individuals with cancer and their contribution to tumour progression is a topic that is gradually gaining attention. Recent evidence has shown that there are prominent connections between immune gene variants and mood disorders. The ho_x0002_meostasis of the tumour immune microenvironment (TIME) and the infiltration and activation of immune cells play a very important role in the antitumour effect. In this study, we established a compound mouse model with chronic unpredictable mild stress (CUMS) and orthotopic colorectal cancer to simulate colorectal cancer (CRC) patients with depression. Using 10✕Genomics single_x0002_cell transcriptome sequencing technology, we profiled nearly 30,000 cells from tumour samples of 8 mice from the control and CUMS groups, revealed that immune cells in tumours under a chronic stress state trend toward a more immunosuppressive and exhaustive status, and described the crosstalk between the overall inflammatory environment and immunosuppressive landscape to provide mechanistic information or efficacious strategies for immune-oncology treatments in CRC with depressive disorders.

overall_design: Single-cell RNA-seq data of four BALB/c mice randomly divided into control and CUMS groups

first: Yan

last: Wang


title: Graded constitutive NF-κB signaling in B cells drives SLL/CLL-like lymphomagenesis and overcomes microenvironment dependence

first: Marc

last: Schmidt-Supprian

overall_design: RNA-seq profiling of B1a cells across four genotypes (Ikk2het, Ikk2hom, TCL1tg, TCL1tgIkk2het) at three time points (3 months, 6 months, 9 months, and old age) to study tumor development (CLL/SLL) and progression.

first: Valeria

last: Soberón

first: Lena

last: Osswald

first: Andrew

last: Moore

first: Gene

last: Swinerd

first: Jingyu

last: Chen

first: Seren

last: Baygün

first: Carina

last: Diehl

first: Laura

last: Kraus

first: Vanessa

last: Gölling

first: Ricarda

last: Trapp

first: Pankaj

last: Singroul

first: Thomas

last: O'Neill

first: Sabrina

last: Bortoluzzi

first: Daniel

last: Kovacs

first: Tim

last: Ammon

first: Yuliia

last: Hubarzhevska

first: Rupert

last: Öllinger

first: Sebastian

last: Müller

first: Olga

last: Baranov

last: Engleitner

first: Yoshiteru

last: Sasaki

first: Felix

last: Gillhuber

first: Sonja

last: Grath

first: Oliver

last: Weigert

first: Andreas

last: Rosenwald

first: Klaus

last: Rajewsky

first: Katja

last: Steiger


title: Herpes simplex virus 1 (HSV-1) R-loops are targets for APOBEC-mediated innate immunity [DRIP-seq]

city: Debrecen

first: Márton

last: Miskei

address: Egyetem tér1.

country: Hungary

institute: University of Debrecen

summary: APOBEC cytidine deaminase enzymes play a crucial role in antiviral innate immunity as they introduce mutations in viral genomes, restrict viral replication, and promote host defense against infections. Special three-stranded nucleic acid structures, called R-loops, have been considered as efficient targets for APOBEC-mediated mutagenesis because they present an ideal substrate, a persistent single-stranded DNA, for the enzyme. However, the relationship between R-loops and APOBEC enzyme activity has not been verified and remains to be elucidated. Here, we reveal a mechanistic link between R-loop formation, APOBEC enzyme binding, and APOBEC mutagenesis at viral R-loop targets during virus infection. In a cellular model involving human T lymphoblastoid leukemia cells infected with Herpes simplex virus 1 (HSV-1), we show that APOBEC3A and APOBEC3G binding sites and mutagenesis signatures occur mainly in viral R-loop structures. C-to-T mutagenesis primarily targets R-loops, which are specifically concentrated in HSV-1 genes that are essential for virus assembly and function. Among these genes, we identified mutation hotspots resulting in amino acid changes, likely leading to inactivation of HSV-1 protein functions. Collectively, our findings show that viral R-loops serve as potent targets for APOBEC-induced mutagenesis, representing an effective strategy to inactivate essential viral proteins and compromise viral activity.

overall_design: DRIP-seq experiments to determine the R-loops on the HSV-1 genome.

first: Marton

first: Lorant

last: Szekvolgyi


title: Characterization of the TREM-1 Signaling Landscape in Human Neutrophils

city: Mainz

first: Matthias

last: Klein

address: Langenbeckstrasse 1

department: Inst. for Immunology

institute: University Medical Center Mainz

summary: The Triggering Receptor Expressed on Myeloid Cells (TREM)-1 is a member of the Immunoglobulin superfamily. Beyond its role in acute and chronic inflammatory processes, TREM-1 is also involved in cancer emergence and progression. Advanced information about the TREM-1 signaling cascade may reveal novel targets for treating inflammatory and cancer diseases. We investigated the transcriptomic profile of PMN after TREM-1 ligation that made us available to link the previously analyzed kinome activity to the resulting transcriptomic profile. In sum, we revealed several signaling pathways being active after TREM-1 ligation that are associated with various biological processed and diseases.

overall_design: RNA-seq profiling of purified human PMN after TREM-1 ligation compared to an isotype matched control as well as after TLR-4 ligation compared to a medium control.

first: Frederic

last: Ries

first: Nora

last: Rogmann

first: Sophie

last: Toebben

first: Federico

last: Marini

first: Florian

last: Heidel

first: Markus

last: Radsak


title: Herpes simplex virus 1 (HSV-1) R-loops are targets for APOBEC-mediated innate immunity [ChIP-Seq]

type: Genome binding/occupancy profiling by high throughput sequencing

overall_design: ChIP-seq experiments to find APOBEC3A and APOBEC3G binding sites on the HSV-1 genome.


title: Unveiling Alternate Pathways for SARS-CoV-2 Infection via Extracellular Vesicle-Mediated Transfer of ACE2 and TMPRSS

city: NEW YORK

first: Ya-Wen

last: Chen

state: NY

address: 1425 MADISON AVE, Icahn 13-20A, Box 1189

department: Otolaryngology

country: USA

institute: Icahn School of Medicine at Mount Sinai

summary: he COVID-19 pandemic, caused by SARS-CoV-2, has underscored the urgency of understanding viral entry mechanisms to develop effective therapeutic strategies. SARS-CoV-2 primarily exploits angiotensin-converting enzyme 2 (ACE2) as its entry receptor and relies on the serine protease TMPRSS2 to prime its spike protein, enabling membrane fusion and infection. Traditionally, TMPRSS2 has been described as a cell surface protein, but our study reveals that in human lung epithelial cells, TMPRSS2 is largely absent from the plasma membrane and instead resides intracellularly. We show that TMPRSS2 is secreted together with ACE2 in extracellular vesicles (EVs) from lung epithelial cells, which are subsequently taken up by non-epithelial cells, specifically alveolar macrophages, endothelial cells, and pericytes, that do not express TMPRSS2 or ACE2 mRNA under homeostatic conditions. This EV uptake deposits ACE2 and TMPRSS2 protein onto recipient cells, equipping them for SARS-CoV-2 entry. By transferring these viral entry proteins, EVs expand the spectrum of susceptible cell types in the lung, offering a new explanation for how the virus can infect diverse cell populations and cause widespread tissue damage. Identifying EVs as vehicles for delivering functional ACE2 and TMPRSS2 across cell types reveals previously unrecognized pathway of viral entry with important implications for not only COVID-19 pathogenesis but also for other viral infections that exploit similar entry mechanisms. These findings open new avenues for therapeutic intervention aimed at disrupting EV-mediated protein transfer, potentially limiting viral dissemination and severity, and may also represent a generalizable mechanism exploited by other viral pathogens, highlighting the potential relevance of EV-mediated protein transfer beyond SARS-CoV-2.

overall_design: An IPF lung was dissociated and sorted into 4 populations based on the cell surface protein expressions of TMPRSS2 and EPCAM.

first: Lu

last: Tian

first: Remi

last: Klotz

first: Min

last: Yu


title: Multi-Omics Atlas-Assisted Discovery of Transcription Factors for Selective T Cell State Programming

city: San Diego

first: April

middle: Elizabeth

last: Williams

state: California

address: 10010 N Torrey Pines Rd

department: IGC

institute: Salk Institute for Biological Studies

summary: Transcription factors (TFs) regulate the differentiation of T cells into diverse states with distinct functionalities. To precisely program desired T cell states in viral infections and cancers, we generated a comprehensive transcriptional and epigenetic atlas of nine CD8+ T cell differentiation states for TF activity prediction. Our analysis catalogued TF activity fingerprints of each state, uncovering new regulatory mechanisms that govern selective cell state differentiation. Leveraging this platform, we focused on two critical T cell states in tumor and virus control: terminally exhausted T cells (TEXterm), which are dysfunctional, and tissue-resident memory T cells (TRM), which are protective. Despite their functional differences, these states share significant transcriptional and anatomical similarities, making it both challenging and essential to engineer T cells that avoid TEXterm differentiation while preserving beneficial TRM characteristics. Through CRISPR screening combined with single-cell RNA sequencing (Perturb-seq), we validated the specific TFs driving the TEXterm state and confirmed the accuracy of TF specificity predictions. Importantly, we discovered novel TEXterm-specific TFs such as ZSCAN20, JDP2, and ZFP324. The deletion of these TEXterm states-specific TFs in T cells enhanced tumor control and synergized with immune checkpoint blockade. Additionally, this study identified multi-state TFs like HIC1 and GFI1, which are vital for both TEXterm and TRM states. Furthermore, our global TF community analysis and Perturb-seq experiments revealed how TFs differentially regulate key processes in TRM and TEXterm cells, uncovering new biological pathways like protein catabolism that are specifically linked to TEXterm differentiation. In summary, our platform systematically identifies TF programs across diverse T cell states, facilitating the engineering of specific T cell states to improve tumor control and providing insights into the cellular mechanisms underlying their functional disparities.

overall_design: P14 (gp33 specific CD8+T cells) are transduced with dual gRNA retrovirus (vector1 or vector2) encoding EGFP to introduce CRISPR knock out. The cells were stained with CITE-seq antibodies (HTOs), and EGFP pos P14 T cells were isolated by Fluorescence-activated cell sorting (FACS).

first: H

middle: K

last: Chung

first: Ming

last: Sun

middle: E

first: Susan

last: Keach


title: LIVER EXERCISE FACTOR REVERSES AGING AND ALZHEIMER’S-RELATED MEMORY LOSS BY TARGETING BRAIN VASCULATURE

city: San Francisco

first: Karishma

middle: Joanne Bhatia

last: Pratt

state: CA

address: 513 Parnassus Ave, Med. Sci. Bldg, Room S1364

institute: UCSF

summary: Blood factors transfer the benefits of exercise to the aged brain. However, mechanisms to restore cognition and the translational application to neurodegenerative disorders are unknown. Here we show that liver-derived exercise factor glycosylphosphatidylinositol (GPI)-specific phospholipase D1(Gpld1)–a GPI-degrading enzyme–reverses aging and Alzheimer’s-related memory loss by targeting brain vasculature. Increasing Gpld1 rejuvenated transcriptional hippocampal aging signatures at a single cell level and rescued memory in aged mice. Mechanistically, we identified GPI-anchored tissue-nonspecific alkaline phosphatase (TNAP) on brain vasculature as a GPLD1 substrate. Mimicking an age-related increase in cerebrovascular TNAP impaired memory in young mice and mitigated Gpld1-mediated cognitive benefits in aged mice. Inhibiting TNAP activity recapitulated the benefits of Gpld1 in old age. In an Alzheimer’s disease model, increasing Gpld1, or inhibiting TNAP, ameliorated A pathology and improved cognitive deficits. Collectively, our data identify cerebrovasculature as a gatekeeper of the restorative cognitive benefits of a liver-to-brain exercise axis.

overall_design: scRNA sequencing experiment from hippocampi of aged mice treated with GPLD1 mutant, GPLD1, and TNAP inhibitor. scRNA sequencing experiment from hippocampi of WT mice, 5xFAD mice treated with GFP control, and 5xFAD mice treated with GPLD1

first: Gregor

last: Bieri


title: The purinergic receptor P2rx7 mediated ATP sensing is required to prevent bone aging by directing mitochondrial fitness of MSCs

city: Beijing

first: Zimeng

last: Zhuang

address: Haidian District

department: Department of Orthodontics

institute: Peking University School and Hospital of Stomatology

summary: Bone aging contributes to osteoporosis and impaired bone formation, mechanisms intricately linked to alterations in mesenchymal stem cells (MSCs) that include a metabolic decline whose precise mechanisms remain elusive. In this study, P2rx7 was identified as a key regulator of mitochondrial fitness in bone marrow MSCs during the bone aging process. To validate the role of P2rx7 in bone metabolism and aging, we deleted P2rx7 by using P2rx7 knockout mice and the mice demonstrated premature and exacerbated bone aging. What we want to explore is how P2rx7 knockout affects the genome changes of bone cells.

overall_design: To investigate how P2rx7 knockout influenced the bone, we constructed P2rx7 knockout mice. Femurs of three different Widetype (WT) mice or P2rx7 knockout mice aged 6-8 weeks were collected the bone marrow were flushed out and used for RNA-seq.


title: Transient Elevation in Cellular Glucose Uptake Exacerbates Pressure-Overload Induced Cardiac Hypertrophy and Dysfunction

city: Birmingham

first: Sayan

last: Bakshi

state: Alabama

department: Department of Pathology

institute: University of Alabama at Birmingham

type: Methylation profiling by high throughput sequencing

summary: BACKGROUND: Clinical studies support that prior hyperglycemia can lead to long-lasting adverse cardiovascular effects, referred to as “glycemic memory”. Several studies support that epigenetic modifications, specifically DNA methylation changes, influenced by altered metabolic pathways, may play a key role in this phenomenon. The objective of this study was to ascertain the gene expression and corresponding DNA methylation signatures linked to glycemic memory, and to investigate if these changes, in long-term, lead to worsened cardiovascular effects upon pressure overload.  METHODS AND RESULTS: Using inducible and cardiomyocyte specific GLUT4 overexpression transgenic mice, we induced glucose delivery for 2 weeks, followed by another 2 weeks of basal uptake. Next, we employed RNA-sequencing and reduced representation bisulfite sequencing and discovered differential gene expression and DNA methylation signatures that persisted even after cellular glucose levels reverted to normal. Significant changes across both expression and methylation enriched pathways associated with adverse cardiac events, indicating a potential glycemic memory response. To understand the long-term effects of these changes, we also performed sham or transverse aortic constriction (TAC) surgery at this four-week timepoint to induce pressure overload-mediated secondary stress. Mice were then followed for an additional 8 weeks and assessed for contractile function by echocardiography, cellular remodeling, and molecular analysis. Pressure overload led to an exacerbated hypertrophic response and cardiac dysfunction. Subsequent qPCR-mediated expression analysis identified molecular changes akin to heart failure, worsened cardiac fibrosis, and impaired oxidative stress.  CONCLUSIONS: These results indicate an altered transcriptome and DNA methylome upon prior transient enhanced glucose delivery, which evokes a glycemic memory response, responsible for long-term heart failure predisposition. We also identified molecular signatures of differential expression and methylation, as well as potential hubs that may provide novel targets for mechanistic understanding and therapeutic interventions.

overall_design: To identify the gene expression signatures linked to glycemic memory, we used doxycycline (DOX) inducible and cardiomyocyte specific GLUT4 overexpression (mG4H) transgenic mice. We induced glucose delivery for 2 weeks in mG4H mice (ON), followed by another 2 weeks of basal uptake (OFF). Next, we employed reduced representation bisulfite sequencing to discover differential DNA methylation changes that persisted even after cellular glucose levels reverted to normal (mG4H-OFF compared to mG4H-ON; adjusted by respective controls). For each group, DNA methylation sequencing was performed on one sample that contained 10 biological replicates.

first: Luke

middle: A

last: Potter

first: Adam

middle: R

last: Wende


title: Gene expression profile at single cell level of lymphocytes extracted from the mesenteric lymph nodes of TCRMOG animals colonized with either Salmonella expressing MOG or OVA

city: Basel

first: DBM Bioinformatics

last: Core Facility

state: BS

address: Hebelstrasse 20

department: Departement of Biomedicine

country: Switzerland

institute: University of Basel

summary: To analyze if expresion of a self-epitope (in this case an epitope of MOG (myleing oligodendrocyte glycoprotein)) in the gut can trigger an autoimmune reaction, TCRMOG (harbor MOG-reactive T cells) were colonized with a gut restricted strain of Salmonella expressing either an epitope of MOG or ovalbumin (OVA). We then compared the gene expression profile from immune cells within the mesenteric lymph node to assess (autoreactive) immune cell activation.

overall_design: Germ-free TCRMOG (or 2D2) mice were mono-colonized with either Salmonella expressing an epitope of MOG (MOG35-55) or OVA (323-339). Mesenteric lymph nodes were extracted 3 weeks post colonization, put through a cell strainer to receive a single cell suspension and then used for gene expression analysis. 3 Mice were used per experimental group.

first: Anne-Katrin

last: Pröbstel

first: Lena

last: Siewert


title: Stable Bioreactor Control Reveals Acidic pH–Driven Metabolic Reprogramming and Mitochondrial Dysfunction in Human Lymphoblastoid Cells

city: THUWAL

first: Samhan

middle: M.

last: Alsolami

address: Thuwal 23955

department: BESE

country: Saudi Arabia

institute: KAUST

summary: Human cells require pH regulation to maintain physiological function, yet the molecular consequences of acidic environments remain incompletely understood. Here, we employ a gas-only bioreactor to control pH, oxygen, and temperature. Integrated Omics analyses reveal that acidic pH induces a glycolytic metabolic shift, suppresses proliferation, and promotes accumulation of lactate and oncometabolites alongside mitochondrial dysfunction. Acidic conditions increase reactive oxygen species (ROS) and activate inflammatory and immune pathways, leading to heteroplasmic enrichment of a pathogenic mitochondrial mutation. Acidic pH depletes intracellular NAD⁺, partly driven by PARP1 activation. Restoring NAD⁺ through nicotinamide mononucleotide (NMN) supplementation partially rescues proliferation and stress-associated transcription, while elevating NAD+ levels by NMN or PARP1 inhibition reverses heteroplasmic enrichment of mutant mitochondrial DNA. These findings underscore the role of pH homeostasis in coordinating metabolism, redox balance, and immune signaling, and identify NAD⁺ metabolism as a mechanistic link between acidic microenvironments, mitochondrial genome instability, and immune–metabolic remodeling.

overall_design: B-LCL treated with pH 6.8 and pH 7.4 in controlled environment

first: Mo

last: Li


title: Effect of ITPRIPL1 treatment on gene expression of human peripheral blood mononuclear cells

first: Yibo

last: Zhang

address: 131 Dongan Road, Xuhui District, Shanghai

department: Institutes of Biomedical Sciences

institute: Fudan University

summary: ITPRIPL1, an inhibitory ligand of CD3ε, is expressed in immune privileged testis and multiple tumors.Human PBMCs were treated with or without ITPRIPL1 to test transcriptome alteration at the single-cell level. We performed the scRNA-seq analysis of the ITPRIPL1-treated and ITPRIPL1-untreated groups based on the 10X Genomics platform.

overall_design: sc-RNA-seq of the single cells from ITPRIPL1-treated and ITPRIPL1-untreated Human PBMCs

first: Shouyan

last: Deng

first: Jie

last: Xu


title: A single small molecule-based human embryo model reveals a requirement for V-ATPase in mammalian blastocyst cavitation

summary:
Human naïve pluripotent stem cells (nPSCs) can be induced by various combinations of
signaling factors to generate blastocyst-like structures, termed blastoids. Despite rapid
progress in human blastoid models, their potential to uncover fundamental mechanisms of
early human development remains limited, leaving key morphogenetic processes poorly
understood. Here, we describe a simple and robust system in which dimethyl sulfoxide
(DMSO) alone induces blastoid formation from human nPSCs. This model recapitulates
key pre- and post-implantation features and exhibits enhanced polar trophectoderm (TE)
organization, more efficient attachment within an implantation-relevant window, improved
epiblast lumenogenesis associated with amniotic cavity formation, and more robust,
sustained expansion of embryonic lineages following attachment. Using this system, we
reveal a previously unrecognized mechanism underlying TE cavitation and identify
lysosome-associated genes — particularly subunits of the proton pump V-ATPase — as
essential regulators of blastoid cavitation. DMSO treatment upregulates key V-ATPase
2
subunits (ATP6V0A4 and ATP6V1B1), which are also enriched in the TE of human
embryos. Genetic or pharmacological inhibition of V-ATPase activity disrupts lysosomal
acidification, blocks intracellular vacuole formation, and impairs blastoid cavitation,
whereas overexpression of V-ATPase subunits rescues this phenotype. Furthermore,
genetic and pharmacological perturbations of V-ATPase function significantly
compromise cavitation in both mouse and human blastocysts. Finally, DMSO treatment
induces membrane biomechanical changes characteristic of early embryonic development,
suggesting a mode of action distinct from conventional small-molecule, signaling pathway-
based induction strategies. This simple DMSO-based blastoid model recapitulates key
aspects of human blastocyst development and reveals a conserved requirement for V-
ATPase-mediated lysosomal acidification during early mammalian embryogenesis.

overall_design: nPSCs were treated with and without DMSO along with A83, PD03, and both PDA83.


title: A renal function for 2,3-bisphosphoglycerate mutase (BPGM)

city: Berlin

first: Robert

last: Labes

address: Charitéplatz 1

department: Institute of Translational Physiology

institute: Charité - Universitätsmedizin Berlin

summary: The enzyme 2,3 bisphosphoglycerate mutase (BPGM) catalyzes the production of 2,3-bisphosphoglycerate (2,3-BPG). The main role of 2,3-BPG is to shift the equilibrium of hemoglobin toward the deoxy-state, accounting for the notion that 2,3-BPG production would appear to be solely useful in cells which contain hemoglobin. 2,3-BPG level determines the baseline O2-Hb affinity at rest, which shows a broad diversity among human population. The kidney is the site of oxygen sensing and erythropoietin synthesis through the action of the hypoxia-inducible factor (HIF) transcription factors, especially HIF-2. We observed a marked BPGM expression in distal parts of renal tubules that is elevated during kidney injury, thus, indicating cellular functions of BPGM not related to oxygen binding to hemoglobin. We, thus, explored the function of renal BPGM, particularly as related to down-stream metabolic effects.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for MEF cells under control or hyperosmolar conditions with and without BPGM knockdown.

first: Michael

last: Faehling

first: Christian

last: Rosenberger


title: Whole transcriptome gene expression in Alzheimer's disease (AD) patients [lncRNA]

city: Boca Raton

first: Massimo

last: Caputi

address: 777 Glades Road

institute: Florida Atlantic University

type: Non-coding RNA profiling by high throughput sequencing

summary: Alzheimer disease (AD) is the most common type of dementia. It is a progressive disease initially characterized by memory loss and leading to loss of the ability to respond to the environment. Currently, there is no single molecular or imaging test that can establish an AD diagnosis. A clinical diagnosis is often the results of multiple cognitive tests, spinal fluid or other bodily fluid tests for specific biomarkers and brain imaging, and it is often a diagnosis of exclusion for other diseases. We utilized a whole transcriptome sequencing approach to analyze gene expression in AD patients and control subjects blood samples with the goal of developing a novel panel of biomarkers for the disease. Whole blood sample were collected from 50 AD patients and 50 matched controls. AD patients were defined by a clinical diagnosis supported by 3 cognitive tests (MMSE, ADAS, CDR). Healthy donors have an unremarkable clinical history and their scores in the 3 cognitive tests are within normal ranges.

overall_design: Whole transcriptome sequencing of peripheral blood from 49 Alzheimer's disease patients and 49 matched controls.

first: Sean

last: Paz

first: Janet

last: Robishaw


title: Dissecting the cellular architecture of breast cancer brain metastases reveals prognostically distinct immune landscapes [snRNA-Seq]

status: Public on Mar 10 2026

city: Heidelberg

first: Christel

last: Herold-Mende

department: Department of Neurosurgery

institute: University Hospital Heidelberg

summary: Breast cancer brain metastases (BCBM) are a severe condition with high demand for improved personalized treatment, but a comprehensive understanding of BCBM immune-microenvironment heterogeneity and susceptibility to immunotherapy is lacking. Here, we multimodally profile the immune niche in a vast, clinically well-annotated cohort of 156 BCBM applying tissue cytometry, bulk and single nuclei RNA-sequencing, flow cytometry, and spatial transcriptomics, complemented by functional studies in patient-derived models. Integrative analyses reveal two immune landscapes predicting prolonged patient survival and not deducible from paired primary tumors: 1) BCBM with a high proportion of CD8 tissue-resident-like memory T cells as major players of tumor immune control. 2) BCBM containing tertiary lymphoid structures. Surrogate signatures of these landscapes are prognostic in independent BCBM and primary breast cancer cohorts, are associated with fewer metastases, and predict immunotherapy response. Our work provides critical insights into anti-tumor immunity in BCBM and identifies novel biomarkers with translational relevance.

overall_design:
We performed single nuclei RNA-sequencing (snRNA-seq) of 28 BCBM. We focused on the TNBC subtype, since TNBC is associated with inferior prognosis of BCBM patients and at the same time is considered immunogenic with ICB use in the clinic. Samples were selected representing the full cohort in terms of CD8 T cell infiltration rate and ecotype variety. We sequenced single nuclei rather than cells for a comprehensive view, preventing potential cell loss by harsh tissue dissociation, and after optimizing the nuclei isolation protocol. This involved controlled tissue preparation and improved in silico doublet detection applying our previously developed demultiplexing pipeline using patient-specific single nucleotide variants (SNP).

***************************************************************
Due to data protection regulations, the raw sequencing data have been deposited at GHGA: https://data.ghga.de/dataset/GHGAD48917471237543
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last: Jassowicz

first: Fangyoumin

last: Feng

first: Rolf

last: Warta

first: Marc

last: Zapatka

first: Martina

last: Seiffert


title: DMSO Drives Trophectoderm and Blastoid Formation from Human Naïve Pluripotent Stem Cells without Lineage-Specifying Signals

city: barcelona

first: aintzane

last: rueda

state: Choose State

address: C/ del Dr. Trueta, 179, 08005 Barcelona

country: Spain

institute: Sequentia Biotech SL.

summary: This study investigates the role of dimethyl sulfoxide (DMSO) in facilitating the formation of human trophectoderm cells and, consequently, blastoids. Current in vitro derivation methods for trophectoderm cells, integral to the formation of blastocysts, implantation, and placental development, are restricted to early pluripotent stem cells such as naïve pluripotent stem cells (nPSCs). The use of PD03 and A83-01 has proven effective for creating human trophectoderm-like cells and subsequent blastocyst-like structures, or blastoids. However, our research introduces DMSO as a significant factor in the differentiation of nPSCs into trophectoderm cells. DMSO is known for its effect on reducing pluripotency and promoting differentiation in human primed embryonic stem cells, and thus we hypothesized its potential in inducing the trophectoderm program in nPSCs. The results of our experiments show that DMSO treatment not only induces morphological changes similar to trophectoderm fate but also results in the downregulation of pluripotency genes and upregulation of trophectoderm markers. The trophectoderm cells resulting from DMSO treatment are able to further differentiate into trophectoderm subtypes. Thus, DMSO could potentially serve as an efficient tool for promoting the formation of human blastoids.

overall_design:
These blastoids were produced using PALLY protocol with and without DMSO. Also, these blastoids were allowed to go on to model peri-implantation in vitro under the same treatment.
D1-D4 samples represent technical replicates.

last: Alsolami

first: Yiqing

last: Jin

first: Yingzi

first: Ismail

last: Shakir

first: Alfonso

last: Saera-Vila

first: Arun

last: Chandrasekaran

first: Gerardo

last: Ramos Mandujano


title: KLF2 overrides the resident memory CD8 T cell differentiation program, in opposition to KLF3

city: Minneapolis

first: Christine

middle: H

last: O'Connor

address: 117 Pleasant St. SE

department: Minnesota Supercomputer Institute

institute: University of Minnesota

summary: Numerous transcriptional regulators have been associated with the differentiation pathways that lead to recirculating versus tissue-resident memory T cells. However, it is unclear whether independent, coordinated expression of these regulators is required to determine residency versus recirculation, or whether there is a hierarchy, with some factors playing a dominant role in controlling T cell trafficking. We report that ablation of the gene encoding Kruppel-like factor 2 (KLF2) during CD8+ T cell activation leads to rapid transcriptional reprograming, such that effector T cells fail to recirculate and prematurely acquire canonical phenotypic and transcriptional characteristics of resident memory cells (TRM). Klf2-deficient memory CD8+ T cells retained the capacity to undergo recall responses, including in vivo pathogen control. These data suggest KLF2 diverts CD8+ T cells from the TRM differentiation program. In contrast, ablation of another member of the KLF family, KLF3, enhanced differentiation of some recirculating T cell subsets and limited production of TRM in lymphoid tissues. However, both KLF2 and KLF3 were required for differentiation of long-lived effector cells, suggesting cooperation between these factors in some situations. These findings indicate KLFs occupy a central nexus in coordinating activated CD8+ T cell differentiation and trafficking.

overall_design: Congenically distinct WT or KLF2-GFP P14 CD8+ T cells were negatively enriched from spleen and activated with plate bound α-CD3 and α-CD28 with rhIL-2 for 48 hours, and then electroporated with sgRNAs/Cas9 complexes for target (Klf2 or Klf3) or control (Cd19 or Thy1) genes through a Lonza 4DNucleofector X Unit, using published conditions (Seki et al. 2018). Following culture in rhIL-2 for 24-48h, 5x10e4 CRISPR-modified cells were co-adoptively transferred into congenically distinct host mice, which received LCMV-Arm infection (2x10e5 PFU) within 2h of transfer. CRISPR modified donor P14 CD8+ T cells were FACS sorted from the spleen, 8- or 25-days post LCMV infection, RNA isolated and submitted for sequencing through the University of Minnesota Genomics Center.

first: Taylor

last: DePauw

first: Kexin

last: Gai

first: Jian

last: Shen

first: Nicholas

middle: J

last: Maurics

first: Ka Hyun

last: Rhee

first: Weiguo

last: Cui

first: Changwei

last: Peng

first: Stephen

middle: C

last: Jameson


title: Distribution of big tau isoforms in the human central and peripheral nervous system

city: New York

first: Joshua

last: Orrick

address: 194 E 2nd St

department: Genetics & Genomics

institute: Mount Sinai School of Medicine

summary: This project aims to characterize the distribution of “big tau,” a longer tau isoform expressed in the peripheral nervous system (PNS) and select central nervous system (CNS) regions, and to examine its relationship with aging and neurodegeneration. We performed mass spectrometric sequencing of big tau sequence and mapped its distribution across the human nervous system. Postmortem samples included brains from Alzheimer’s disease (AD), disease controls, and amyotrophic lateral sclerosis (ALS); spinal cord from young controls, disease controls and ALS; and peripheral nerves. Big and small tau levels were also quantified in the cerebrospinal fluid (CSF) from young normal controls, amyloid positive and amyloid negative participants. To reinforce the presence of big tau in the brain, iso-seq was performed on post-mortem brain tissue samples.

overall_design: MAPT-focused iso-seq was performed on five caudate and five middle frontal gyrus post-mortem frozen brain tissue samples from individual control donors with European ancestry.

first: Chiara

last: Pedicone

first: Alison

last: Goate


title: Evolutionary Analysis of Transcriptional Regulation Mediated by Cdx2 in Rodents

city: Shenzhen

first: Guipeng

last: Li

address: 1088 Xueyuan Avenue

institute: Southern University of Science and Technology

sample_organism:
Mus musculus
Rattus norvegicus

platform_organism:
Mus musculus
Rattus norvegicus

overall_design: RNA-seq, ATAC-seq and CDX2 ChIP-seq of mES cells and RMES cells

first: Wei

last: Chen


title: Whole transcriptome gene expression in Alzheimer's disease (AD) patients [sRNA]

first: Massimo

last: Caputi

first: Sean

last: Paz

first: Janet

last: Robishaw


title: Transcriptomics and experiments verified that astaxanthin reduces the apoptosis of nerve cells in ischemic stroke by inhibiting the toll-like receptor signaling pathway

city: 吉林省长春市

first: Wang

last: Xu

address: 吉林省长春市净月开发区博硕路

department: college of traditional chinese medicine

institute: Changchun University of Chinese Medicine

summary:
Background and Objective. Inflammation is an important pathological process in ischemic stroke (IS). Astaxanthin (ATX) is a natural product with neuroprotection effects. However, the mechanism of ATX on anti-inflammatory after IS is not clear. The aim of this study was to investigate the mechanism of ATX on anti-inflammatory after IS.
Methods. Male Sprague-Dawley rats were used to establish a model of middle cerebral artery occlusion (MCAO) on one side, and were pre-treated with gavage of ATX for 7 days. One day after MCAO, the brain tissues were collected. Transcriptomic sequencing, flow cytometry, brain water content, western blot, HE staining and ELISA were analyzed to evaluated the brain damage.
Results. ATX treatment has improved the neurological deficits, reduced brain edema, and apoptosis. Also, ATX has reduced inflammation and apoptosis related proteins such as TLR4, MyD88, NFκB, IL1β, IL6, Cyt C and Caspsae3.
Conclusions. ATX can improve nerve damage after IS, and these protective effects were realized by anti-inflammatory and anti-apoptosis. This protective mechanism involves the TLR4/MyD88/NF-κB signaling pathway


title: Whole transcriptome gene expression in Alzheimer's disease (AD) patients

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

overall_design: Refer to individual Series


title: Contribution of cytotoxic CD8 T cells, neutrophils and type 1 interferon signaling to hyperinflammatory pathology in HIV associated TB meningitis

city: London

first: James

middle: Royce

last: Barnacle

state: London

address: Francis Crick Institute, 1 Midland Road

department: Infectious Disease

country: United Kingdom

institute: Imperial College London

summary: Immune dysregulation contributes to death and disability in tuberculous meningitis. People living with HIV have the highest risk of TBM, greatest death and disability, and the least evidence that anti-inflammatory therapy improves the poor outcome. Improving therapy relies on a more refined understanding of the host immune response. Using single-cell RNA sequencing, we characterised 188,983 lumbar CSF cells from 25 adults with HIV-associated TBM and identified correlates of bacterial load. There was heterogeneity in cell composition between patients, but cytotoxic CD8 T cells with low cytokine expression were consistently predominant with a large number expressing GZMK, known to activate complement. In microbiologically-confirmed TBM, there was greater cytotoxicity in T, NK and γδ cells, and higher type 1 interferon stimulation in T and B lymphocytes. Neutrophils expressed markers suggesting heightened cytokine stimulation, enhanced effector function, and IL-8-mediated peripheral neutrophil recruitment. In a separate longitudinal cohort, type 1 interferon signalling increased in the blood and CSF following treatment initiation. Overall, findings indicate a hyper-inflammatory immune response in the CSF of HIV-associated TBM patients characterised by an accumulation of granzyme-rich cytotoxic CD8 T cells, highly activated neutrophils and host-detrimental type 1 IFN signalling.

overall_design: Bulk RNA sequencing was performed on cerebrospinal fluid taken three, seven and/or 28 days post-enrolment from patients with HIV-associated TB meningitis.

first: Barnacle

last: James

first: Bangani

last: Nonzakazi

first: Slawinski

last: Hubert

first: Barrington

last: Christopher

first: Wilkinson

last: Katalin

first: Stek

last: Cari

first: Lai

last: Rachel

first: Meintjes

last: Graeme

first: Robertson

last: Brian

first: Gengenbacher

last: Martin

first: Davis

last: Angharad

first: Barber

last: Daniel

first: O'Garra

last: Anne

last: Robert J


first: Samhan

last: Alsolami

first: Mo

last: Li


title: Promoter mutagenesis and a massively parallel reporter screen of the MAPT locus identifies cis-regulatory elements and genetic variation effects [RNA-seq]

city: Huntsville

first: Jesse

middle: Nicholas

last: Cochran

state: AL

address: 601 Genome Way

institute: HudsonAlpha Institute for Biotechnology

summary: Tau neurofibrillary tangles are a hallmark of several neurodegenerative diseases called tauopathies, including frontotemporal dementia and Alzheimer's Disease. Ongoing clinical trials for tauopathies seek to reduce Tau in the brain through immunotherapy, antisense oligonucleotides, and siRNA. MAPT codes for Tau, therefore understanding how the MAPT gene is regulated and the effect of genetic variation at its regulatory elements is likely to have high relevance for tauopathies. We screened a ~3 Mb region including the MAPT locus using 2 different massively parallel reporter assay (MPRA) strategies in KOLF2.1J h-NGN2 neurons and HEK293FT cells, identifying previously unannotated cis-regulatory elements (CREs). Using CRISPR interference (CRISPRi) in mixed neuron cultures, we identified a new CRE for MAPT, as well as 2 CREs for another nearby gene of interest, KANSL1. Known genetic variation from the Alzheimer's Disease sequencing project was tested in a separate MPRA at the top CREs near the MAPT gene, identifying variants with altered regulatory effects including those at previously identified CREs for MAPT. Using a saturation mutagenesis screen of a 2,000 bp region encompassing the MAPT promoter, we assessed regulatory effects of each possible single nucleotide variant in this region. We identified several neuron-specific regulatory variant effects at this region, including a high confidence binding site for the transcription factors EGR2, ZBTB14 and TCLF5 at a region of high MPRA activity and genetic conservation.

overall_design: RNAseq of  CRISPRi  performed in KOLF2.1J mixed neuron cultures. NPCs were transduced with CRISPRi contstructs targeting the AAV safe harbor locus (negative control), MAPT promoter (positive control), as well as CREs of interest (r1-r14). The cells were  differentiated into mixed neuron cultures, and RNAseq was performed to determine effect on MAPT expression.

first: Rebecca

middle: M

last: Hauser

first: Nicholas


title: Genetic deletion of cytoglobin exacerbates cardiac hypertrophy and inhibits cardiac fibroblast activation independent of changes in blood pressure

city: Albany

first: David

last: Jourd'heuil

address: 43 New Scotland Ave

department: Molecular and Cellular Physiology

institute: Albany Medical College

summary: Hypertension-mediated left ventricular hypertrophy and cardiac fibrosis often precede heart failure. Recent studies indicate that cytoglobin (Cygb), a globin expressed in the vasculature, increases systemic blood pressure. The present work aims to determine the role of Cygb in angiotensin II (Ang II)-induced cardiac hypertrophy and fibrosis in the mouse. Methods: Males and females global Cygb knockout (Cygb−/−), and wildtype (Cygb+/+) mice were treated with Ang II (1.5 µg/kg/day) for two weeks via subcutaneous osmotic minipumps. Cardiac function was assessed through echocardiography, and hearts were analyzed for changes in hypertrophy, fibrosis, and gene expression. Functional studies were also performed in isolated cardiac fibroblasts. Results: Cygb−/− mice from both sexes showed an increase in cardiac hypertrophy over Cygb+/+ mice. Cardiac functions were also depressed in Cygb−/− males with no changes in females. Importantly, genetic deletion of Cygb did not affect systemic blood pressure in mice, at baseline or after Ang II treatment. We established that Cygb was expressed in fibroblasts and pericytes in humans and mice hearts. Finally, we found that Cygb−/− cardiac fibroblast did not upregulate the expression of genes associated with myofibroblasts following treatment with Ang II. This was reversed following expression of human cytoglobin. Conclusions: Our findings indicate that Cygb plays a protective role in the mouse heart during Ang II-induced cardiac stress. This is the first study detailing the function of Cygb in the heart as a regulator of cardiac hypertrophy. This study also reveals a role for Cygb in regulating cardiac fibroblast activation by Ang II. NEW & NOTEWORTHY: We identified cytoglobin as an important globin in cardiac pathophysiology. Genetic deletion of cytoglobin led to exacerbation of angiotensin II-mediated cardiac hypertrophy in the absence of any effect on systemic blood pressure. Cytoglobin is expressed in cardiac fibroblasts and pericytes and is required for cardiac fibroblast activation to myofibroblast. The present study reveals for the first time a role for cytoglobin in regulating angiotensin II signaling.

overall_design: Bulk RNA-seq was performed on left ventricular tissue from global cytoglobin knockout (Cygb−/−) mice and wild-type (WT) littermate controls after 2 weeks of continuous infusion with angiotensin II (Ang II) via subcutaneous osmotic pumps.Left ventricles were harvested at the end of the 2-week infusion period, RNA was isolated from each individual mouse, and sequencing libraries were prepared for gene expression profiling by bulk RNA sequencing. This design enables comparison of the transcriptional response of the left ventricle to Ang II between WT and Cygb knockout mice in order to identify gene expression programs and pathways that are altered by the loss of cytoglobin during Ang II–induced cardiac stress.

first: Le Gia Cat

last: Pham

first: Kurrim

last: Gilliard

first: Frances

first: Sarahann

last: Mistretta

first: John

last: Schwarz

first: Harold

last: Singer


title: Effects of Rg3@PACVs with NIR in a Rat Model of Pulmonary Ischemia–Reperfusion Injury

city: guangzhou

first: Haoxiang

last: Yuan

address: 28 Qiaozhong Middle Road, Liwan District

department: Department of Thoracic Surgery and Oncology

institute: The First Affiliated Hospital of Guangzhou Medical University

summary: Pulmonary ischemia–reperfusion injury (IRI) is characterized by excessive inflammation, oxidative stress, and mitochondrial dysfunction, leading to severe lung damage. To elucidate the molecular mechanisms underlying the protective effects of a macrophage membrane-coated mesoporous polydopamine nanoparticle system loaded with Ginsenoside Rg3 (Rg3@PACVs) combined with near-infrared (NIR) irradiation, we performed transcriptome sequencing of rat lung tissues. Differential gene expression profiles were compared between IRI and IRI + Rg3@PACVs + NIR groups. A total of 209 upregulated and 332 downregulated genes were identified in the treatment group relative to IRI controls. Functional enrichment analyses revealed that Rg3@PACVs + NIR pretreatment enhanced pathways related to endoplasmic reticulum stress response, PI3K-AKT signaling, oxygen transport, protein folding, hypoxia response, TCA cycle activity, ATP-dependent processes, and cell cycle progression. In contrast, inflammation-associated pathways including TLR/NF-κB signaling, cytokine binding, cell adhesion molecules, neutrophil extracellular trap formation, IL-1β production, and leukocyte trans-endothelial migration were significantly suppressed. GSEA further demonstrated reduced oxidative stress (cytochrome P450 activity) and enhanced FOXO signaling. These findings indicate that Rg3@PACVs + NIR mitigates pulmonary IRI by coordinately suppressing inflammatory responses, alleviating oxidative stress, and restoring mitochondrial homeostasis, thereby promoting lung tissue repair.

overall_design: The male SD rats were subjected to pulmonary ischemia–reperfusion injury and randomly assigned to two groups: IRI (n = 3) and IRI + Rg3@PACVs + NIR (n = 3). Total RNA was extracted from the left lung tissues for high-throughput sequencing.

first: Bo

last: Zeng

first: Pu

last: Shen

first: Jiancheng

last: Deng

first: Ying

last: Chen

first: Meiyu

last: Huang

first: Wentao

last: Wu

first: Xin

last: Xu

first: Xuanlin

last: Zhang

first: Xue

last: Liu

first: Xudong

first: Zhijin

last: Fan

first: Jianxing

last: He


title: Contribution of cytotoxic CD8 T cells, neutrophils and type 1 interferon signaling to hyperinflammatory pathology in HIV associated TB meningitis [RNA-Seq]

first: James

last: Barnacle

overall_design: Bulk RNA sequencing was performed on blood samples for adults enrolled in the LASER-TBM study with HIV-associated TBM at baseline and at five further timepoints during treatment.

first: Nonzakazi

last: Bangani

first: Hubert

last: Slawinski

first: Christopher

last: Barrington

first: Katalin

last: Wilkinson

first: Cari

last: Stek

first: Rachel

last: Lai

first: Graeme

last: Meintjes

first: Brian

last: Robertson

first: Martin

last: Gengenbacher

first: Angharad

last: Davis

first: Daniel

last: Barber

first: Anne

last: O'Garra

first: Robert


title: Functional analysis of NtWRKY70 regulating tobacco defense against Rhizoctonia solani AG3-TB

city: Shenyang

first: Yan

last: Li

address: No. 120 Dongling Road, Shenhe District

department: Plant Protection College

institute: Shenyang Agricultural University

summary: Rhizoctonia solani AG3-TB is a destructive soil-borne necrotrophic fungus that causes tobacco target spot disease. WRKY70 serves as a critical regulatory hub integrating salicylic acid (SA) and jasmonic acid (JA) signaling pathways to modulate defense responses. Previous studies have revealed that NtWRKY70 is significantly upregulated following R. solani AG3-TB infection, but its critical role in the resistance mechanism remains unclear.

overall_design: In this study, bioinformatics and experimental analyses demonstrated that NtWRKY70 encodes a WRKY family protein with a conserved WRKYGQK motif, lacking signal peptides and transmembrane domains, possessing nuclear localization and autoactivation activity. We constructed tobacco genetic transformation materials via virus-induced gene silencing (VIGS), CRISPR/Cas9-mediated knockout, and overexpression technologies demonstrated that NtWRKY70 was an important gene against R. solani AG3-TB. Transcriptomic analysis and reverse transcription quantitative PCR (RT-qPCR)  validation identified five key differentially expressed genes (DEGs) upregulated in OE-NtWRKY70 plants, including SARD1, Cytochrome P450, AOS1, acidic endochitinase P and CH3L2. In addition, exogenous application of SA and methyl jasmonate (MeJA) induced expression of NtWRKY70 and enhanced host resistance. Collectively, this study confirms that NtWRKY70 confers tobacco resistance to R. solani AG3-TB by coordinating SA and JA signaling pathways and activating downstream defense-related genes.


title: Phage-encoded CasPRs transcriptionally silence diverse CRISPR-Cas systems [RNA-Seq]

city: Seattle

first: Alexander

last: Meeske

state: WA

address: 750 Republican St

institute: University of Washington

summary: Anti-CRISPRs (Acrs) are diverse proteins or RNAs that protect invading phages and plasmids from host CRISPR-Cas immunity. Most Acrs neutralize their cognate Cas proteins via direct physical interaction. Here we describe CasPRs, a particularly widespread family of DNA-binding Acrs that recognize specific sequence motifs within cas gene coding regions, thereby blocking RNA polymerase and silencing transcription. We demonstrate that eight diverse CasPRs bind to the cas8b gene to repress the type I-B CRISPR-Cas system in its native host, Listeria seeligeri. Meanwhile, a CasPR from Streptococcus dysgalactiae silences type II-A CRISPR-Cas immunity by binding to the cas9 coding sequence. We found that one CasPR is required to inhibit CRISPR immunity during lysogeny by its host prophage. Taken together, our results indicate that members of the CasPR family have diverged to silence completely unrelated CRISPR types, and suggest transcriptional repression is a common mode of phage-mediated immune antagonism.

overall_design: RNA-seq in the presence and absence of CasPR expression in Listeria seeligeri.

first: Edith

last: Sanderson

first: Julieta

last: Peralta

first: Sam

last: Nouwens

first: Luke

last: Oriolt

first: Victoria

last: Hayes

first: Brett

last: Kaiser


title: Transcriptomic Analysis of Argon Effects on LPS-stimulated Primary Microglia

city: Nantong

first: Xia

state: Jiangsu

address: No. 9 Seyuan Road

institute: Nantong University

summary: The role of inflammation in psychiatric disorders, such as anxiety, has gained increasing attention, with the hippocampus being identified as a key region involved in emotional regulation. Argon has been reported to alleviate the symptoms of psychiatric disorders; however, its underlying mechanism remains unclear. In this study, we found that argon significantly suppressed lipopolysaccharide-induced anxiety-like behaviors and attenuated hippocampal neuronal hyperexcitability in mice. By analyzing neuronal excitability following microglial depletion and subsequent repopulation, as well as assessing microglial morphology, we confirmed that microglia are key targets of inflammation and validated the inhibitory effects of argon. Electrophysiological studies and transcriptome sequencing revealed that argon inhibited the microglia-released inflammatory cytokines via the Toll-like receptor 4/nuclear factor-κB signaling pathway, thereby improving the excitability of hippocampal neurons. Collectively, this study provides evidence that the regulation of microglia may be the underlying mechanism by which argon ameliorates neuroinflammation-induced anxiety-like behaviors.

overall_design: Primary microglia were isolated from brain of fetal Sprague–Dawley (SD) rats and cultured under standard conditions. Cells were assigned to three groups: (1) control, treated with vehicle solution; (2) LPS treatment, treated with 100 ng/mL lipopolysaccharide (LPS) for 4 hours; and (3) LPS + Argon treatment, treated with LPS (100 ng/mL) followed immediately by Argon exposure for 3 hours.Each treatment group included three independent biological replicates. RNA was extracted from each replicate and subjected to RNA sequencing (RNA-seq) to profile gene expression changes in response to LPS and Argon treatment.

first: Jie

last: Cheng

first: Baiyang

last: Zheng

first: Shusu

last: Luo

first: Yuan

last: Yuan

first: Xiaobo

last: Wu

first: Zhenglin

last: Jiang


title: Surveillance of aneuploidy by oxidative stress sensing through ATPAF1

city: Shanghai

first: Xiaoai

last: Lyu

address: 567 West Yangsi Road

institute: New York University Shanghai

summary: Aneuploidy, the aberrant number of chromosomes, is a major consequence of chromosomal instability (CIN) and a hallmark of cancer. Aneuploidy imposes significant fitness costs related to proteotoxic, replicative, mitotic, and metabolic stress. However, the molecular basis by which cellular surveillance systems sense these aneuploidy-associated stresses remains poorly understood. Using a genome-wide CRISPR-Cas9 screen, we identified ATPAF1 as a novel aneuploidy sensing gene. Loss of ATPAF1 confers proliferation advantage in CIN conditions and enriched aneuploid population by reducing apoptosis. Mechanistically, we establish a bidirectional relationship between aneuploidy onset and oxidative stress, revealing that ATPAF1 functions as a specific sensor for aneuploidy-associated reactive oxygen species. Our pan-cancer analysis demonstrates a positive correlation between the dysregulations of oxidative stress response pathways and increased aneuploidy severity, suggesting how cancer cells can actively reprogram oxidative stress responses to strive for survival despite CIN. Taken together, our work elucidates a novel mechanism of aneuploidy surveillance mediated by ATPAF1-dependent oxidative stress sensing, offering potential therapeutic targets or biomarkers for overcoming cancer aneuploidy.

overall_design: CRISPR-seq of HCT116 cells containing a GeCKO-v2 KO library treated with 0.7 μM AZ3146 for 14-days or without AZ3146

first: Jungseog

last: Kang

first: Xinhe

last: Xue


title: Mitochondrial-Localized Keratin 17 Promotes Chemoresistance in Basal-Like Pancreatic Cancer

city: New Haven

first: Luisa

last: Escobar-Hoyos

state: CT

address: 15 York Street PO box 208040

department: Therapeutic Radiology

institute: Yale University

summary: The basal-like molecular subtype of pancreatic ductal adenocarcinoma (PDAC) is highly lethal and therapy resistant. A better understanding of the underlying molecular mechanisms driving this aggressive tumor subtype is necessary for the development of effective therapies. Notably, upregulation of keratin 17 (K17) in cancer is associated with poor patient outcome and the basal-like PDAC subtype. Here, we identified a critical dependency of basal-like PDACs on de novo pyrimidine biosynthesis, driven by intra-mitochondrial K17. Mechanistically, K17 translocated into the mitochondrial intermembrane space via a mitochondrial localization sequence (MLS) recognized by the translocase of the outer mitochondrial membrane 20 (TOM20). In the mitochondria, K17 bound to and stabilized dihydroorotate dehydrogenase (DHODH), the rate-limiting enzyme of de novo pyrimidine biosynthesis, by preventing its ubiquitination-mediated degradation. Blocking the entry of K17 into the mitochondria sensitized cancer cells to gemcitabine, a pyrimidine analog and standard chemotherapeutic agent. In animal studies, pharmacologic inhibition of DHODH combined with gemcitabine treatment decreased tumor growth and doubled survival in mice bearing K17⁺ but not K17⁻ PDAC. These findings define a mitochondrial role for K17 in driving pyrimidine biosynthesis and uncover a metabolic vulnerability in K17⁺ basal-like PDACs that can be therapeutically targeted.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: RNA-seq profiling of KPC empty vector/K17, L3.6 wild type/K17-knockout, and PANC-1 empty vector/K17 cells.

first: Chun-Hao

last: Pan

first: Yinghuan

first: Monisankar

last: Ghosh

first: Md Afjalus

last: Siraj

first: Robert

last: Tseng

first: Nina

middle: V

last: Chaika

first: John

middle: D

last: Haley

first: Bahman

last: Khalvatifahlylani

first: David

last: Tuveson

first: Hardik

last: Patel

first: Muaz

last: Faruque

first: Girish

last: Rajacharya

first: Katie

middle: L

last: Donnelly

first: Cindy

last: Leiton

first: Carlos Mejia

last: Arbelaez

first: Haoting

last: Chen

first: Sumedha

last: Chowdhury

first: Shayan

last: Sarkar

first: Lyanne Delgado

last: Coka

first: Lucia

last: Roa-Peña

first: Michael

last: Horowitz

first: Natalia

last: Marchenko

first: Pankaj

last: Singh

first: Kenneth

last: Shroyer


title: Genome-wide profiling identifies the genetic dependencies of cell death following EGFR inhibition

city: Worcester

last: Lee

state: MA

address: 55 N Lake Ave

institute: Umass Chan Medical School

summary: EGFR is a proto-oncogene that is mutationally activated in a variety of cancers. Small molecule inhibitors targeting EGFR can effectively slow the progression of disease, and in some settings, these drugs even cause dramatic tumor regression. However, responses to EGFR inhibitors are rarely durable, and the mechanisms contributing to response variation remain unclear. In particular, several distinct mechanisms have been proposed to explain how EGFR inhibition activates cell death, and a consensus has yet to emerge. In this study, we use functional genomics with specialized analyses to infer how genetic perturbations affect the drug-induced death rate. Our data clarify that inhibition of PI3K signaling drives the lethality of EGFR inhibition. Inhibition of other pathways downstream of EGFR, including the RAS-MAPK pathway, promotes growth suppression but not the lethal effects of EGFR inhibitors. Taken together, our study provides a “reference map” for the genome-wide genetic dependencies of lethality in response to EGFR inhibitors.

overall_design: Genome-wide CRISPR KO screen of PC9-Cas9 cells treated with 10 µM erlotinib, 0.1 µM osimertinib, 5 µM osimertinib, or DMSO for 72 hours. T0 samples were taken immediately prior to drugging.

first: Sydney

last: Porto

first: Gavin

last: Birdsall

first: Nicholas

middle: W

last: Harper

first: Megan

last: Honeywell

first: Scott

last: Leighow

first: Tiana E

last: Naylor

first: Kelly M

last: Ward

first: Mika K

last: Wesley

first: Justin

last: Pritchard


title: Dissecting the cellular architecture of breast cancer brain metastases reveals prognostically distinct immune landscapes [RNA-Seq]

first: Christel

last: Herold-Mende

overall_design:
This dataset comprises bulk RNA sequencing data generated from cryopreserved breast cancer brain metastasis tissues obtained from patients treated at the Department of Neurosurgery, University Hospital Heidelberg. The samples were processed to extract total RNA and subsequently subjected to high-throughput sequencing to capture transcriptomic profiles. These data served as the discovery cohort for the development of the tissue-resident memory T cell (TRM) gene signature, which was derived through a supervised machine-learning approach. The TRM signature was subsequently evaluated and validated in independent external cohorts to assess its robustness and clinical relevance.
All raw sequencing data have been deposited at https://data.ghga.de/dataset/GHGAD28055171876192

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

last: Jassowicz

first: Rolf

last: Warta


title: RNA-seq data for P. putida KT2440 under diverse conditions for regulatory network analysis

city: La Jolla

first: Heera

last: Bajpe

address: 9500 Gilman Dr.

institute: UC San Diego

summary: This dataset consists of RNA-seq data for P. putida KT2440 to enable comparison of transcriptional regulatory networks across Pseudomonas strains.

overall_design: Expression profiling data for P. putida KT2440 under a diverse set of conditions (different media types, osmotic stress, oxidative stress, nitrogen starvation, iron starvation, zinc ion stress, copper ion stress, antibiotic treatment, mixed carbon source treatment) were generated through RNA sequencing.

first: Ying

last: Hefner

first: Richard

last: Szubin

first: Jaemin

last: Sung

first: Bernhard

middle: O

last: Palsson


title: Genome-wide profiling identifies the genetic dependencies of cell death following EGFR inhibition [RNA-Seq]

first: Michael

last: Lee

overall_design: RNA-sequencing of PC9 and H1650 non-small cell lung cancer cell lines with and without EGFR inhibitors 10 uM erlotnib and .1 uM osimertinib at 0 and 36 hours.

first: Sydney

last: Porto

first: Tiana

first: Kelly

first: Mika

middle: K


title: Diet Composition Shapes Sleep, Quality of Life, and Gene Expression in Obesity: A Randomized Trial.

city: Oxford

first: Matthew

last: Baxter

address: John Radcliffe Hospital

department: RDM

institute: University of Oxford

summary: Obesity affects quality of life and sleep, making effective weight loss an attractive intervention.  The role of diet composition on outcomes has not been determined. Our aim is to compare a ketogenic diet against an isocaloric balanced diet for effects on quality of life, sleep, circadian rhythm and gene expression in obesity. Twenty-four people affected by obesity completed the study. Volunteers were randomized to a 1200 Kcal ketogenic diet or balanced diet for 30 days, and all were encouraged to walk 30 minutes every day. Questionnaires for assessing quality of life and sleep were administered at baseline and at the end of the study. In a subgroup of 20 people actigraphy was performed to objectively capture sleep parameters, and circulating whole blood gene expression analysis was undertaken. Quality of life assessed with the IWQOL-Lite significantly improved after both diets (p<0.001), with the ketogenic diet showing a greater effect (p=0.006). The ketogenic diet also resulted in greater improvement in subjective sleep quality (p=0.023).  Both diets improved daytime sleepiness (balanced diet p=0.012; ketogenic diet p=0.021). The ketogenic diet induced a greater weight loss (p=0.026). Gene expression analysis revealed greater changes in lipid metabolism with ketogenic diet. No significant changes in circadian genes were seen.  Weight loss with diet intervention significantly improved quality of life of people with obesity, already after 30 days.  The ketogenic diet resulted in greater improvements, in parallel with greater weight loss. Both diets influenced the expression of many metabolic genes in whole blood, with greater changes in lipid metabolism seen with ketogenic diet. Weight loss with diet intervention is effective to improve quality of life, sleep, and daytime sleepiness symptoms.

overall_design: RNA-seq profiling of whole blood gene expression analysis in 20 individuals, stratified by actigraphy derived sleep parameters.

first: Elena

last: Gangitano

first: Giorgio

last: Caratti

first: David

last: Ray


title: Adenosine-specific transcriptional programs in murine connective tissue type mast cells

city: Heidelberg

first: Qihua

last: Liang

state: Baden-Württemberg

address: Im Neuenheimer Feld 366

department: Institute of Pharmacology

institute: Universitat Heidelberg

summary: Mast cells are tissue-resident immune cells that are critical for the pathogenesis of allergic and inflammatory disorders. Their physiological functions include host defense against parasites and, more recently, food quality control through antigen avoidance. The purine nucleoside adenosine (ADO), like other mast cell activators, such as antigens or Mrgprb2 agonists, increases intracellular Ca2+ concentration; however, it fails to induce degranulation of preformed mediators when applied to mast cells alone, and there is limited knowledge of whether ADO evokes the de novo synthesis and release of inflammatory mediators in tissue mast cells.  An unbiased genome-wide analysis of gene expression triggered by various mast cell activators should enable identification of the gene program specifically activated by ADO in mast cells and thereby reveal new components of the associated inflammatory responses. Here, we performed bulk RNA sequencing in primary murine peritoneal mast cells (PMCs) as connective tissue mast cells. By comparing responses evoked by ADO stimulation with those of the Mrgprb2 agonist compound 48/80 and antigens activating FcεRI receptors, we identified 393 genes uniquely regulated by ADO, including genes encoding the de novo synthesized mediators transforming growth factor α and interleukin 7. Transcription factor activity inference, protein classification, functional enrichment analysis, protein interaction network, and topology analysis revealed a distinct ADO-specific transcriptional gene program involved in phosphoinositide signaling, vesicle trafficking, glycolysis, mitochondrial activity, and cell cycle arrest. The functional relevance of the identified de novo synthesized mediators for ADO-evoked inflammatory reactions can be evaluated in future studies.

overall_design: RNA-Seq profiling of  murine connective tissue type mast cells with five experimental conditions: CONTROL, adenosine (ADO, 10 µM) for 2 hours, compound 48/80 (C48/80, 50µg/mL) for 6 hours, antigen (dinitrophenyl (DNP)-human serum albumin 100 ng/mL) for 2 hours, and antigen (dinitrophenyl (DNP)-human serum albumin 100 ng/mL) for 6 hours.

first: Volodymyr

last: Tsvilovskyy

first: Anouar

last: Belkacemi

first: Merima

last: Bukva

first: Christin

last: Richter

first: Nicole

last: Ludwig

first: Andreas

last: Keller

first: Marc

last: Freichel


title: RNA-seq data for P. syringae pv. tomato DC3000 under diverse conditions for regulatory network analysis

first: Heera

last: Bajpe

summary: This dataset consists of RNA-seq data for P. syringae pv. tomato DC3000 to enable comparison of transcriptional regulatory networks across Pseudomonas strains.

sample_organism: Pseudomonas syringae pv. tomato str. DC3000

overall_design: Expression profiling data for P. syringae pv. tomato DC3000 under a diverse set of conditions (different media types, osmotic stress, oxidative stress, nitrogen starvation, zinc ion stress, copper ion stress, antibiotic treatment) were generated through RNA sequencing.

first: Ying

last: Hefner

first: Richard

last: Szubin

first: Jaemin

first: Bernhard


title: Contribution of cytotoxic CD8 T cells, neutrophils and type 1 interferon signaling to hyperinflammatory pathology in HIV associated TB meningitis [TBM_scRNAseq]

first: James

last: Barnacle

overall_design: Single cell RNA sequencing was performed using 10x Fixed RNA Profiling on cerebrospinal fluid taken seven days post-enrolment from patients with HIV-associated TB meningitis.

first: Barnacle

last: James

first: Bangani

last: Nonzakazi

first: Slawinski

last: Hubert

first: Barrington

last: Christopher

first: Wilkinson

last: Katalin

first: Stek

last: Cari

first: Lai

last: Rachel

first: Meintjes

last: Graeme

first: Robertson

last: Brian

first: Gengenbacher

last: Martin

first: Davis

last: Angharad

first: Barber

last: Daniel

first: O'Garra

last: Anne

last: Robert J


title: Single-cell RNAseq identifies persistent epithelial and immune dysfunction in PwCF by mitigating inter-individual sampling heterogeneity

city: Chicago

first: Manu

last: Jain

state: IL

address: 303 E Superio

institute: Northwestern University

summary: Nasal epithelium (NE) is used as a surrogate for lower airway epithelium in cystic fibrosis (CF). However, NE is composed of diverse cell types with unique functions; changes to this heterogeneous tissue in disease can be cell-type specific. To explore the impact of tissue heterogeneity, we examine NE gene expression and cell-type heterogeneity using single-cell transcriptomics of people with CF (PwCF) and healthy volunteers (HV). We also compare cell-type abundance in paired single-cell and bulk transcriptomic samples.

overall_design:
Single-cell RNAseq (scRNAseq) was performed on nasal curettage samples from 9 PwCF and 7 HVs. Paired bulk RNAseq samples were obtained from a subeset of 5 PwCF and 5 HVs.

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

first: Thaddeus

last: Cybulski

first: Marc

last: Sala

first: Trevor

last: Nicholson

first: Zasu

last: Klug

first: Rachel

last: Nelson

first: Zhan

last: Yu

first: Yuliana

last: Sokolenko

first: Estefani

last: Diaz

first: Suchitra

last: Swaminathan

first: Ziyan

last: Lu

first: Nikolay

middle: S

last: Markov

first: Edward

middle: F

last: McKone

first: Alexander

last: Misharin


title: Sorting nexin 5 is necessary for MHC Class II antigen presentation and host defense against Mycobacterium tuberculosis infection.

city: Dallas

first: Michael

last: Shiloh

state: TX

address: 5323 Harry Hines Blvd. Y9.308

department: Internal Medicine

institute: University of Texas Southwestern Medical Center - UTSW

summary: Tuberculosis (TB) remains one of the leading causes of death from a single infectious agent worldwide, yet the host pathways that regulate antigen presentation and lung inflammation during Mycobacterium tuberculosis (Mtb) infection are incompletely defined. Sorting nexin 5 (SNX5) is a phox homology (PX) and bin, amphiphysin, rvs (BAR) domain protein best known for roles in endosomal trafficking, antigen processing, and antiviral host defense, but its contribution to immunity during Mtb infection is unknown. Here, we show that SNX5-deficient mice exhibit markedly increased mortality following low-dose aerosol infection despite unchanged pulmonary bacterial burden compared to wild-type mice. Snx5-/- mice developed exacerbated lung inflammation without major alterations in immune cell recruitment. In macrophages, Snx5-/- did not affect phagocytosis, vacuolar maturation, intracellular bacterial control, or global transcriptional responses to Mtb, but was required for efficient MHC class II antigen presentation. Snx5 deficiency reduced antigen degradation, limited peptide loading onto MHC-II and impaired activation of antigen-specific CD4+ T cells without altering surface MHC-II abundance or expression of costimulatory molecules. Together, these findings identify SNX5 as a previously unrecognized regulator of MHC-II peptide loading that shapes inflammatory outcomes during pulmonary Mtb infection, highlighting a role for endosomal sorting machinery in immunity to intracellular pathogens.

overall_design: Bone marrow cells from Snx5 Knockout or wild-type  C57BL/6J from 3 different animals were plated in 6-well plates (10ˆ6) and infected or not with Mtb (MOI of 5). After 4 or 24 hrs of infection, samples were lysed using TRIzol and RNA was isolated using phenol & the Qiagen RNeasy Mini Kit.  Then, we performed bulk RNA-sequencing.

first: Kubra

last: Naqvi

first: Victoria

last: Ektnitphong

first: Samuel

last: Alvares-Arguedas

first: Kathryn

last: Rahlwes

first: Priscila

last: Campos


title: Single-Cell Analyses Identify Independent Aging Processes that Compete to Determine Cellular Fate in Budding Yeast -- bulk RNA-Seq

city: South San Francisco

first: Nathaniel

last: Thayer

address: 1130 Veteran's Blvd

institute: Calico Life Sciences LLC

summary: Phenotypic heterogeneity is prevalent during aging, yet its underlying molecular drivers remain poorly understood. In budding yeast, two distinct aging trajectories, characterized by either ribosomal DNA (rDNA) instability or mitochondrial decline, have been proposed to be mutually exclusive. Here, we systematically dissect the heterogeneity among aging yeast cells by combining single-cell transcriptomics with longitudinal fluorescence microscopy. Our data reveals distinct transcriptional responses that emerge in aging cells, highlighted by loss of rDNA silencing, a hypoxia response, and the Environmental Stress Response (ESR). Contrary to expectation, we establish that ESR induction is not caused by rDNA instability but is instead a consequence of an early decline in mitochondrial membrane potential (MMP). However, the ESR is merely a biomarker of this decline and not itself a determinant of lifespan. While rDNA instability and mitochondrial dysfunction are anti-correlated as terminal phenotypes, we find that they are not necessarily mutually exclusive and can instead proceed concurrently within individual cells. Targeted genetic perturbations that are specific for one pathway do not impinge on the other, which is in contradiction to the idea of mutual inhibition between the two. We therefore propose a "competing hazards model", where independent aging processes progress in parallel, and the observed mode of death is determined by which process first reaches a catastrophic failure point. Our work untangles the causal links between several aging pathways and provides a new framework for understanding how distinct aging trajectories emerge from independent molecular events.

overall_design: Budding yeast cells (WT, msn2∆ and msn2∆msn4∆) were harvested as young and aged to perform bulk RNS-Seq.

first: Manuel

last: Hotz

middle: G

last: Kroll-Ling

first: Daniel

middle: E

last: Gottschling


title: Single-Cell Analyses Identify Independent Aging Processes that Compete to Determine Cellular Fate in Budding Yeast -- scRNA-seq

overall_design: Budding yeast cells (young or aged) were either directly used for single-cell RNA-Seq (timecourse samples) or sorted into subpopulations before scRNA-Seq. SRP40 samples were sorted into high and low fluorescent subpopulations, MMP samples were sorted into high and low MMP ratio subpopulations (as decribed in 10.1083/jcb.202505110).

first: Rachel


title: Axonal dying back of upper motor neurons in human ALS

first: Fabien

last: Dachet

address: 1200 W Harrison St

institute: University of Illinois Chicago

summary: Patients with amyotrophic lateral sclerosis (ALS) present with arm, leg, or bulbar weakness with or without spasticity. While genetics plays an important role, it cannot explain why symptoms start focally or how upper (UMN) and lower motor neuron (LMN) systems are linked. Here, we examined the clinicopathological relationships between UMN and LMN disease in ten ALS patients. Detailed clinical assessments were collected with tissues from the motor cortex, brainstem, and spinal cord via a rapid autopsy protocol. Tissues were stained for UMN/LMN, myelin, axons, microglia, and pTDP43. Total RNA-sequencing was performed in the medulla, cervical, and lumbar spinal cords from each patient to identify pathways enriched at sites of disease onset. None of the patients had symptoms of frontotemporal dementia (FTD), but all had focal sites of clinical onset and spasticity, indicating both UMN and LMN involvement. Postmortem examination showed LMN degeneration and microglial activation were highest at sites of disease onset. In contrast, UMN degeneration of the corticospinal tract (CST) was present equally at all levels of the spinal cord up through the medulla, regardless of the site of disease onset. Surprisingly, there was no UMN degeneration of cortical motor neurons and their proximal axons down to the medulla. Similarly, while extensive pTDP43 aggregates were seen in degenerating LMNs, no pTDP43 aggregates were seen in UMN cell bodies or their axons. Mechanistically, RNA-sequencing implicated inflammatory pathways, especially at sites of disease onset. Our findings suggest that many ALS patients without FTD have a dying back of UMN axons, independent of the site of disease onset, which stops in the brainstem with preservation of cortical motor neurons and their proximal axons. UMN axonal degeneration could be directly triggered by LMN degeneration and inflammation.

overall_design: This project examines how upper and lower motor neuron pathology are linked in amyotrophic lateral sclerosis (ALS). Using detailed clinical data, postmortem tissue analysis, and RNA-sequencing of various CNS regions, we relate sites of symptom onset to underlying molecular and pathological changes.

first: Haley

last: Cropper

first: Fozia

last: Mir

first: Jianguo

last: Liu

first: Vidushi

middle: R

last: Srivastava

first: Mohammed

last: Ramizuddin

first: Kylie

last: Kopecky

first: Ebony

last: Mocanu

first: Qin Li

last: Jiang

first: Madhu

last: Soni

first: Tibor

last: Valyi-Nagy

first: Diana

last: Mnatsakanova

first: Charles

last: Abrams

first: Fei

last: Song

first: Jeffrey

middle: A

last: Loeb


title: Comprehensive single-nucleus molecular and cell profiling features for cardiac tissues across platforms [10X_snRNAseq]

city: Saint Louis

first: Haewon

last: Shin

state: MO

address: 660 S Euclid

institute: Washington University School of Medicine in St. Louis

summary: Single-nucleus RNA sequencing (snRNA-seq) enables transcriptomic profiling of complex tissues such as the heart, where dissociation into intact single cells is challenging. Among commercially available snRNA-seq platforms, 10x Genomics Chromium and Takara Bio’s ICELL8 cx represent technically distinct approaches, yet systematic comparisons in cardiac tissue are limited. Here, we benchmarked the 3′ Chromium, 5′ Chromium, and ICELL8 cx platforms using nuclei isolated from mouse and human heart tissues, including healthy donors and patients with non-ischemic cardiomyopathy. We assessed nuclei capture efficiency, library complexity, transcript detection sensitivity, RNA biotype coverage, ambient RNA contamination, and cell-type resolution. The Chromium platforms achieved higher nuclear throughput with robust capture efficiency and low ambient RNA contamination, whereas ICELL8 cx provided deeper transcriptomic coverage per nucleus, detecting a broader range of RNA biotypes, including low-abundance non-coding RNAs, albeit with lower nuclei yield. All platforms consistently resolved major cardiac cell populations. Together, these results inform selection of snRNA-seq workflows for cardiac tissue studies, highlighting trade-offs between cellular throughput and transcriptomic depth.

overall_design: Ethical Approval for Human Specimens The study is compliant with all relevant ethical regulations and has been approved by the Washington University School of Medicine Institutional Review Board (IRB #201104172). Informed consent was obtained from each patient prior to tissue collection by Washington University School of Medicine, and no compensation was provided in exchange for subject participation in the study. All demographic and clinical data have been de-identified.  Inclusion Criteria Strict inclusion criteria were employed before tissue collection and any patients with human immunodeficiency virus or hepatitis were excluded. Healthy donor heart tissue samples were collected from patients with stable ejection fraction with no known history of cardiovascular disease and with a non-cardiac cause of death/transplant. NICM heart tissue samples were collected from patients with HF independent of ischemia and no known history of myocardial infarction.  Human single cell isolation for snRNA-seq Frozen myocardial tissue from left ventricular assist device (LVAD) cores or the corresponding region from the apex of explanted donors were minced with a razor blade and dissociated with a plastic pestle in Chromium Nuclei Isolation Kit Lysis Buffer (10X Genomics, Pleasanton, CA). Samples were incubated on ice for 10 minutes, and the tissue suspension was pipetted onto the Nuclei Isolation Column and Collection Tube and centrifuged at 16,000 rcf for 20 seconds at 4°C. Flow-through in the Collection Tube was centrifuged again at 1,000 rcf for 3 minutes to pellet the nuclei and resuspended in 300 μl of Wash and Resuspension Buffer before staining with 1 μl  DRAQ5 (5 mM solution, ThermoFisher Scientific) and incubating on ice for 10 minutes. DRAQ5+ nuclei were sorted into Nuclei Wash Buffer on a BD FACS Melody (BD Biosciences) using a 100 μm nozzle. Recovered nuclei were centrifuged again under the above parameters and were gently resuspended in Nuclei Wash Buffer to a target concentration of 1,000 nuclei μl−1. Nuclei were counted on a hemocytometer and concentration was adjusted as necessary. Following flow cytometry, sorted DRAQ5+ nuclei were centrifuged at 2,000 rcf for 5 minutes at 4°C, and nuclei were resuspended in 100 µl 1X PBS, prior to loading into the Chromium or ICELL8 cx systems as described below.  After manually counting nuclei using Trypan Blue, based on the concentration, samples were loaded at a volume to target 20,000 nuclei. Libraries were prepared using the manufacturers’ instructions-10X 3’ v4 GEM-X (Chromium GEM-X Single Cell 3’ Reagent Kits v4 Document Number CG000731 Rev B August 2024) or 10X 5’ v3 GEM-X (Chromium GEM-X Single Cell 5’ Reagent Kits v3 Document Number CG000733 Rev A March 2024) kits (10X Genomics)- and sequenced on a NovaSeqX (Illumina, San Diego, CA) instrument following manufacturers’ guidelines: 28 X 10 X 10 X 90 [read1 X i7 index X i5 index X read2].

first: Urvi

middle: N

last: Shroff

first: Xianglong

last: Zhang

first: Tracy

last: Yamawaki

first: Daniel

last: Lu

first: Junedh

last: Amrute

first: Andrea

last: Bredemeyer

first: Amrita

last: Das

first: Brandon

last: Ason

first: Simon

last: Jackson

first: Chi-Ming

last: Li

first: Kory

middle: J

last: Lavine


title: Gene expression profiling of Escherichia coli isolates from aging colonies (Series 1)

city: Paris

first: Claude

last: Saint-Ruf

address: 24 rue du faubourg Saint-Jacques

department: Institut Cochin

country: France

institute: Université paris Cité

summary: This series presents normalized gene expression profiles of three Escherichia coli K-12 MG1655-derived strains (pygYFP, Y5, and Y6). Strains Y5 and Y6 were originally isolated from 3-week-old aging colonies, but RNA for transcriptomic profiling was extracted from 7-day-old colonies regrown under identical conditions. All samples were hybridized to the NimbleGen GPL9088 one-color microarray platform, and processed values correspond to RMA-normalized, non-log2 intensities. These data compare the parental strain to two evolved isolates carrying distinct regulatory and stress-response mutations, and can be used to study diversification and adaptive strategies emerging in aging bacterial colonies.

platform_organism: Escherichia coli str. K-12 substr. MG1655

overall_design:
Three RNA samples were analyzed in this series: one parental control strain (pygYFP_B1) and two evolved isolates (Y5_B1 and Y6_B1) that were originally recovered from 3-week-old aging colonies. For transcriptome profiling, all strains were regrown under identical conditions and RNA was extracted from 7-day-old colonies. Each sample was hybridized once to the NimbleGen GPL9088 platform.
*** This submission (Series 1 of BioProject PRJNA1355247) does not include raw NimbleGen files because the original *.pair* files from this older experiment are no longer available. ***

first: Adrien

last: Launay

first: Olivier

last: Tenaillon

first: Ivan

last: Matic


title: Gene expression profiling of Escherichia coli isolates from aging colonies (Series 2)

summary: This series presents normalized gene expression profiles of three Escherichia coli   K-12 MG1655-derived strains (pygYFP, Y5, and Y6). Strains Y5 and Y6 were originally   isolated from 3-week-old aging colonies, but RNA for transcriptomic profiling was   extracted from 7-day-old colonies regrown under identical conditions.    All samples were hybridized to the NimbleGen GPL9088 one-color microarray platform.   Processed values correspond to RMA-normalized, non-log2 intensities. Raw .pair files   and the processed matrix for these samples are provided.    These data complement Series 1 and constitute a second biological replicate   (B2) for transcriptomic analysis of the parental strain and two evolved isolates   carrying distinct mutations affecting regulatory and stress-response pathways   in aging bacterial colonies.

overall_design: Three RNA samples were analyzed in this series: one parental control strain   (pygYFP_B2) and two evolved isolates (Y5_B2 and Y6_B2) originally recovered   from 3-week-old aging colonies. For transcriptome profiling, all strains   were regrown under identical conditions and RNA was extracted from 7-day-old   colonies. Each sample was hybridized once to the NimbleGen GPL9088 platform   (one-color Cy3).


title: Ribosome recruitment assays in a synthetic, m¹A methylated pool of mRNA

first: Sigrid

last: Nachtergaele

address: 260 Whitney Ave

department: Molecular, Cellular, and Developmental Biology

summary: The transcriptome-wide distribution of N¹-methyladenosine (m¹A) in mRNA remains contested, hindering a clear understanding of regulation of mRNA metabolism by m¹A. Additionally, the enzyme(s) that installs the majority of reported mRNA m¹A sites has yet to be identified. In this study, we characterized TRMT61B, an m¹A methyltransferase known to methylate mitochondrial RNAs, but whose exact sequence preferences have been underexplored. We performed ribosome recruitment assays on a synthetic RNA pool of 5UTR and CDS sequences in vitro methylated by TRMT61B.

overall_design: An RNA pool containing >35,000 human and >500 viral 5′UTR and CDS regions (300 nt total length, including adapter sequences, full 5'UTRs, and truncated endogenous CDS was generated through in vitro transcription of a synthetic DNA oligonucleotide library. RNA was then untreated (no enzyme, NE) or methylated with low (LE) or high (HE) amounts of TRMT61B in a large (640-680 uL) reaction. NE, LE, HE samples were then pooled together and four replicates of Direct Analysis of Ribosome Targeting (DART) was carried out. Sequencing was then carried out on input and 80S-bound RNA fractions, using InduroRT to generate cDNA. DESeq2 was then carried out to compare abundance of individual transcripts in the input vs. 80S-bound RNA samples.

first: Dorthy

last: Fang

first: John

last: Babich

first: Ryan

last: Stanton

first: Isaac

last: Vock

first: Kyrillos

last: Abdallah

first: Mingyi

last: Zhu

first: Richard

first: Matthew

last: Simon

first: Wendy V

last: Gilbert