title: Co-transcriptomic analysis of group A Streptococcus (GAS; Streptococcus pyogenes) invasions of human endothelial cells

city: Taipei City

first: Shu-Wei

middle: Marcia

last: Su

address: No. 155, Sec. 2, Linong St, Beitou District

country: Taiwan

institute: National Yang Ming Chiao Tung University

summary: Group A Streptococcus (GAS, aka Streptococcus pyogenes) causes an array of human diseases from mild pharyngitis to life-threatening necrotizing fasciitis. Invading host cells is a strategy for GAS to avoid antibiotic killing and immune system clearance. Our previous study showed that GAS is able to multiply in human microvascular endothelial cell line-1 (HMEC-1), and has higher survival in HMEC-1 cells due to insufficient acidification in lysosomes. GAS peroxide response regulator (PerR) modulates not only peroxide stress, but also metal homeostasis and bacterial virulence. Therefore, we aimed to investigate the role of PerR during GAS invading endothelial cells. First, we found that ΔperR mutant was more tolerant to H2O2 in vitro using hydrogen peroxide sensitivity assay. We further used cDNA-qPCR analysis to clarify the resistance mechanism of ΔperR mutant. The gene expressions of dpr, ahpC, and ahpF were up-regulated, explaining the enhanced resistance of the ΔperR mutant against peroxide stress. However, the proliferation of the ΔperR mutant in HMEC-1 cells was significantly lower than wild type strain after 5 h post-infection. To explore the underlying mechanisms of ΔperR mutant during infection, we performed dual RNAseq analysis to identify differentially expressed genes, and validated them using cDNA-qPCR analysis. The resulting up-regulated genes of iron efflux pump pmtA, iron/zinc chelating protein dpr, and zinc acquisition system (adcA, lmb/adcAII, phtD), and down-regulated zinc efflux pump czcD gave rise to impaired metal homeostasis in the ΔperR mutant. Furthermore, in vitro growth curve assays showed that the ΔperR mutant was sensitive to zinc deficiency and resistant to zinc toxicity. Taken together, this study has demonstrated the critical roles that GAS PerR plays in protecting from peroxide stress of host innate immune responses and zinc sequestration of nutritional immunity. Our novel findings open a new avenue of strategies in the development of antimicrobial agents. Our study demonstrates the importance of PerR. It aids GAS virulence in immune evasion during the LC3-associated phagocytosis in endothelial cells.

sample_organism:
Streptococcus pyogenes
Homo sapiens

platform_organism:
Streptococcus pyogenes
Homo sapiens

overall_design: For dual RNA-seq analysis, human microvascular endothelial cell line-1 (HMEC-1) cells containing intracellular GAS were collected after 1 hour of infection. We isolated the total RNA samples from HMEC-1 cells only, GAS strain NZ131 wild-type-infected HMEC-1 cells, and GAS strain NZ131 ∆perR-infected HMEC-1 cells from four independent infection experiments.

first: Marcia

last: Shu-Wei Su

first: Jiunn-Jong

last: Wu


title: THAP1 is a maternal effect factor required for the first cell cycle  via Rrm1 in early mouse embryos

status: Public on Dec 19 2025

city: Beijing

first: xi

last: wu

address: Tsinghua univesity

country: China

institute: Tsinghua university

type: Expression profiling by high throughput sequencing

summary: Maternal effect genes (MEGs) produce factors that accumulate in oocytes and play critical roles in embryo development. Mutations of MEGs are frequently linked to reproductive and congenital disorders. The majority of identified mammalian MEGs encode epigenetic factors and RNA regulators. Here, we identify a MEG encoding the transcription factor Thanatos-associated protein 1 (Thap1). Thap1 is highly expressed in mouse oocytes and early embryos. Oocyte-specific deletion of Thap1 results in delayed progression of mouse embryos from the 1-cell to the 2-cell stage and 2-cell arrest, accompanied by defective zygotic genome activation (ZGA) and strongly impaired female fertility. Mechanistically, THAP1 activates a critical subset of genes in oocytes, including Rrm1, which produces ribonucleotide reductase required for generating deoxynucleotide triphosphates (dNTPs). Low-input metabolome profiling across 7 stages during the oocyte-to-embryo transition shows gradual, THAP1-dependent dNTP accumulation that peaks in MII oocytes. Overexpression of Rrm1 in zygotes almost fully restores the 2-cell progression and ZGA in Thap1 maternal-knockout embryos. Our findings identify THAP1 as a key maternal effector critical for the earliest stage of mammalian development.

overall_design: To investigate the function of THAP1 in mouse oocytes and early development, we conducted a Thap1 maternal knockout mouse. RNA-seq was used to examine the transcriptome in control and Thap1 mKO mouse oocytes and embryos.

first: Qiang

last: Fan

first: Xi

first: Yanna

last: Dang

first: Shuyan

last: Ji

first: Wei

last: Xie


title: Agouti integrates environmental cues to regulate paternal behaviour

status: Public on Dec 18 2025

city: Princeton

first: Forrest

middle: Dylan

last: Rogers

state: NJ

address: 40 Woodlands Way

department: Princeton Neuroscience Institute

country: USA

institute: Princeton University

summary: Paternal care is rare among mammals and the neural mechanisms governing its emergence are poorly understood. We leveraged the natural paternal behaviour of African striped mice (Rhabdomys pumilio), and natural variation therein, to explore the neurobiological mechanisms that subserve male parental care. To support this aim, we performed single-nucleus RNA-sequencing (snRNA-seq) using the 10X Genomics platform in MPOA samples from sexually naïve alloparental (n=3), infanticidal (n=3), or control males (n=3), as well as sires (n=4), and dams (n = 4).

overall_design: We performed single-nucleus RNA-sequencing using the 10X Genomics platform in MPOA samples from sexually naïve alloparental (n=3), infanticidal (n=3), or control males (n=3), as well as sires (n=4), and dams (n = 4). Following euthanasia via rapid decapitation, we isolated brain tissue from the medial preoptic area of each animal. This tissue was flash frozen. Frozen samples were processed to isolate nuclei prior to sequencing. One sample corresponds to one animal. We did not pool samples.

middle: D

first: Sehee

last: Kim

first: Sarah

middle: A

last: Mereby

first: Anna

middle: M

last: Kasper

first: Anastasios

middle: B

last: Callanan

first: Ricardo

last: Mallarino

first: Catherine

middle: J

last: Pena


title: Microglia replacement by ER-Hoxb8 conditionally immortalized macrophages provides insight into Aicardi-Goutières Syndrome neuropathology

city: Philadelphia

first: Frederick

middle: Chris

last: Bennett

state: PA

address: 3501 Civic Center Blvd

department: Neurology

institute: University of Pennsylvania

summary: Microglia, the brain’s resident macrophages, can be reconstituted by surrogate cells - a process termed “microglia replacement.” To expand the microglia replacement toolkit, we here introduce estrogen-regulated (ER) homeobox B8 (Hoxb8) conditionally immortalized macrophages, a cell model for generation of immune cells from murine bone marrow, as a versatile model for microglia replacement. We find that ER-Hoxb8 macrophages are highly comparable to primary bone marrow-derived (BMD) macrophages in vitro, and, when transplanted into a microglia-free brain, engraft the parenchyma and differentiate into microglia-like cells. Furthermore, ER-Hoxb8 progenitors are readily transducible by virus and easily stored as stable, genetically manipulated cell lines. As a demonstration of this system’s power for studying the effects of disease mutations on microglia in vivo, we created stable, Adar1-mutated ER-Hoxb8 lines using CRISPR-Cas9 to study the intrinsic contribution of macrophages to Aicardi-Goutières Syndrome (AGS), an inherited interferonopathy that primarily affects the brain and immune system. We find that Adar1 knockout elicited interferon secretion and impaired macrophage production in vitro, while preventing brain macrophage engraftment in vivo - phenotypes that can be rescued with concurrent mutation of Ifih1 (MDA5) in vitro, but not in vivo. Lastly, we extended these findings by generating ER-Hoxb8 progenitors from mice harboring a patient-specific Adar1 mutation (D1113H). We demonstrated the ability of microglia-specific D1113H mutation to drive interferon production in vivo, suggesting microglia drive AGS neuropathology. In sum, we introduce the ER-Hoxb8 approach to model microglia replacement and use it to clarify macrophage contributions to AGS.

overall_design: Myeloid cells (progenitors, monocytes, or macrophages) sequenced in vitro or in vivo (brain-engrafted), with or without CRISPR-editting. A minimum of 3 replicates per sample.

first: Nemec

last: Kelsey M

first: Uy

last: Genevieve

first: Chaluvadi

last: V Sai

first: Purnell

last: Freddy S

first: Elfayoumi

last: Bilal

first: Byerly

last: Leila

first: O'Reilly

last: Micaela L

first: O'Brien

last: Carleigh A

first: Aisenberg

last: William H

first: Lombroso

last: Sonia I

first: Guo

last: Xinfeng

first: Blank

last: Niklas

first: Oon

last: Chet Huan

first: Yaqoob

last: Fazeela

first: Temsamrit

last: Brian

first: Rawat

last: Priyanka

first: Thaiss

last: Christoph A

first: Bailis

last: Will

first: Williamson

last: Adam P

first: Wang

last: Qingde

first: Bennett

last: Mariko L

last: F Chris


title: Pathogenic Role of MIF Receptor (CD74) Expressing T cells in Inflammatory Arthritis

city: New Haven

first: Edward

last: Doherty

state: CT

address: 300 Cedar street

department: Internal Medicine

institute: Yale School of Medicine

summary: High expression alleles of the innate cytokine, macrophage migration inhibitory factor (MIF), are associated with the development or the severity of autoimmune inflammatory diseases, including rheumatoid arthritis. Numerous studies support MIF’s role in activating inflammatory pathways and MIF inhibition reduces joint pathology in different experimental models of arthritis.  We examined the impact of gene deletion of MIF or its cognate receptor CD74 in the T cell-dependent model of collagen-induced arthritis (CIA) and observed the complete absence of arthritis development, suggesting an unforeseen role for MIF/CD74 signaling in the development of arthritogenic T cells.  While MIF has been shown in model systems to contribute to T cell activation by augmenting innate responses, fewer than 1% of T lineage cells express CD74 in naïve spleens and lymph nodes, and its functional consequences in pathogenic T cell subpopulations have not been studied.  We found CD74+ T cells to expand during CIA and to increase in number within joint synovium, where they express an effector memory phenotype and recapitulate CIA development upon transfer into naïve mice. We further found evidence for the presence of CD74+ T cells in the circulation and joint synovium of patients with rheumatoid arthritis.  MIF-dependent, CD74+ T cells may contribute to the chronicity of rheumatoid synovitis and to disease relapse in previously inflamed joints.

overall_design: Single cell suspensions were prepared from the 3 spleens of mice either naïve or 28 days after injection with CFA + Chicken Type II collagen.  Samples were then barcoded using TotalSeq Ahashtags (BioLegend, San Diego, CA). The samples were processed at Yale’s W.M. Keck Biotechnology Resource Laboratory with 10x Genomics (Pleasanton, CA) microfluidics for single cell isolation with RNA sequencing performed on the resulting plates.

first: Lais

last: Osmani

first: Joshua

last: Bilsborrow

first: Jennefer

last: Par-Young

first: Susanna

last: Choi

first: Pathricia

last: Tilstam

first: Min

last: Shin

first: Marta

last: Piecychna

first: Helen

last: Cai

first: Lin

last: Leng

first: Wan-Uk

last: Kim

first: Insoo

last: Kang

first: Richard

last: Bucala


title: Single-cell multi-omic and multi-region atlas of the Aging Male Mouse Brain

city: La Jolla

first: Joseph

middle: R

last: Ecker

state: CA

address: 10010 North Torrey Pines Road

department: Genomic Analysis Laboratory

institute: HHMI-Salk-Institute

type: Methylation profiling by high throughput sequencing

summary: Aging is the primary risk factor for many neurodegenerative diseases, primarily attributed to progressive changes in cellular epigenomes. Despite this known association, the complexity of the mammalian brain complicates the identification of brain regions and cell types that are most susceptible to aging effects. While alterations in gene expression have been observed in aging neural cells, comprehension of the epigenetic regulatory mechanisms underlying the transcriptomic changes remains largely elusive.  To unveil these complexities in a comprehensive manner, we utilized single-nucleus methylome sequencing (snmC-seq3) and multi-omic sequencing (snm3C-seq) to produce extensive datasets comprising 132,551 methylomes and 72,666 chromatin conformation-methylome joint profiles from eight distinct brain regions of C57BL/6J mice aged 2, 9, and 18 months. These regions include the anterior hippocampus (AHC), posterior hippocampus (PHC), frontal cortex (FC), amygdala (AMY), nucleus accumbens (NACB), PAG/PCG, entorhinal cortex (ENT), and caudate putamen (CP). We utilized this dataset to investigate age-related changes in DNA methylomes and transcription factor motifs potentially affected by methylation alterations. We also examined methylation changes in transposable elements, observing locus-specific retrotransposon activation. At the chromatin conformation level, we analyzed aging-related changes in compartmentalization, topologically associating domains (TADs), and chromatin loops. Furthermore, we explored regional heterogeneity during aging within identical cell types and validated our findings using the MERFISH dataset.

overall_design:
C57BL/6J (males and females) mice of different ages were purchased from Jackson Laboratories and maintained in the Salk animal barrier facility on 12-hour dark-light cycles with food ad-libitum for up to 4 weeks (Housing condition: Temperature: 21-23 °C, relative humidity: 61-63%). Brains were extracted, sliced, and dissected in an ice-cold dissection buffer as previously described (Liu et. al. 2021). Females were synchronized for their estrous cycle by exposing them to male bedding for three days, and collected during diestrus. For snmC-seq3 and snm3C-seq samples, brains were sliced coronally at 600 μm intervals from the frontal pole across the whole brain, yielding 18 slices, and dissected brain regions were obtained according to the Allen Brain Reference Atlas Common Coordinate Framework version 3 (CCFv3) as previously described (Liu et. al. 2023). For nuclei isolation, each region was dissected from 2-7 animals and pooled to make 2-3 biological replicates. No statistical methods were used to predetermine sample size. We empirically determined to use two to three biological experiments for all single-cell epigenomic experiments to achieve minimum reproducibility for this large-scale project. Blinding and randomization is not performed during handling the tissue samples. Additionally, all dissected regions were digitally registered into CCFv3 using ITK-SNAP60 (v4.0.0) at a 25 μm resolution.

Additional raw data for this project are included in the following series:
https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE169182
https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE169236
https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE169251
https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE169273
https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE169345
https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE169384
https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE169467
https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE181116
https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE181141
https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE181359

The file "Additional_GSMs_analyzed.tsv" contains the mapping of the GSM associated with raw data to the GSM in this series that contains the quantified processed data of that raw data.

first: Qiurui

last: Zeng

first: Wei

last: Tian

first: Anna

last: Bartlett

last: Nery

first: Rosa

last: Gomez Castanon

first: Julia

last: Osteen

first: Jordan

last: Altshul

first: Nicholas

last: Johnson

first: Mia

last: Kenworthy

first: Cynthia

last: Valadon


title: Ectopic expression of cytosolic DHODH uncouples de novo pyrimidine biosynthesis from mitochondrial electron transport

city: San Francisco

first: Andrea

last: Curtabbi

address: 140 Dolores Street APT 208

institute: UCSF

summary: Dihydroorotate dehydrogenase (DHODH) is a rate-limiting enzyme of de novo pyrimidine synthesis. In most eukaryotes, this enzyme is bound to the inner mitochondrial membrane, where it couples orotate synthesis to ubiquinone reduction. As ubiquinone must be regenerated by respiratory complex III, pyrimidine biosynthesis and cellular respiration are tightly coupled. Consequently, inhibition of respiration suppresses DNA synthesis and cell proliferation. Here, we show that expression of Saccharomyces cerevisiae URA1 gene (ScURA) in mammalian cells uncouples pyrimidine biosynthesis from mitochondrial electron transport. ScURA forms a homodimer in the cytosol that uses fumarate as electron acceptor instead of ubiquinone, enabling respiration-independent pyrimidine biosynthesis. Cells expressing ScURA are resistant to drugs that inhibit complex III and the mitochondrial ribosome. Additionally, ScURA enables the growth of mtDNA-lacking ρ0 cells in uridine-deficient medium and ameliorates the phenotype of cellular models of mitochondrial diseases. Overall, this genetic tool uncovers the contribution of pyrimidine biosynthesis to the phenotypes arising from electron transport chain defects.

overall_design: RNAseq analysis of 143B WT cells (CONTROL) or expressing ScURA (ScURA) treated with antimycin/myxothiazol (INHIBITOR for WT and SCURINH for ScURA) or veichle

first: Sara Natalia

last: Jaroszewic

first: Rocío

last: Sanz Cortés

first: Rebeca

last: Acín-Pérez

first: Dimitrios

last: Prymidis

first: Maksym

last: Cherevatenko

first: Raquel

last: Martínez-de-Mena

first: María Jesús

last: Esteban-Amo

first: Miguel

last: de la Fuente

first: Christian

last: Frezza

first: José Antonio

last: Enríquez


title: Spatially distinct cellular and molecular landscapes define prognosis in triple negative breast cancer [1]

city: Pasadena

first: David

middle: Eugene

last: Frankhouser

address: 1500 E Duarte Rd

department: Population Sciences

institute: City of Hope

summary: Triple-negative breast cancer is a prevalent breast cancer subtype with the lowest 5-year survival. Several factors contribute to its treatment response, but the inherent molecular and cellular tumor heterogeneity are increasingly acknowledged as crucial determinants. Here we report on the spatial and molecular heterogeneity underlying a retrospective, treatment-naïve group of women with differential prognoses (17 with >15 years survival- good prognosis (GPx) and 15 with <3 years survival-poor prognosis (PPx)) profiled using GeoMX® DSP. Analyses revealed that the state of epithelia and its microenvironment (TME) are transcriptionally distinct between the two groups. Invasive epithelia in GPx shows a significant increase in immune transcripts, with the TME exhibiting increased immune cell presence (via IF). PPx epithelia, in contrast, are more metabolically and translationally active, with the TME being more mesenchymal/fibrotic. Notably, pre-cancerous epithelia in PPx display a prescience of aggressiveness, as evidenced by increased EMT-signaling. We identify distinct epithelial gene signatures for PPx and GPx, that can, with high accuracy, classify samples at the time of diagnosis and are likely to inform therapy.

sample_organism:
Homo sapiens
synthetic construct

platform_organism:
Homo sapiens
synthetic construct

overall_design: GeoMx human whole transcriptome atlas was performed on ROIs that were selected from 32 FFPE sections from good prognosis (GPx) and poor prognosis (PPx) triple negative breast cancer samples.

first: Kavitha

last: Mukund

first: Darya

last: Veraksa

middle: E

first: Lixin

last: Yang

first: Jerneja

last: Tomsic

first: Raju

last: Pillai

first: Srijan

last: Atti

first: Zahra

last: Mesrizadeh

first: Daniel

last: Schmolze

first: Xio-Cheng

last: Wu

first: Mary-Anne

last: LeBlanc

first: Lucio

last: Miele

first: Augusto

last: Ochoa

first: Victoria

last: Seewaldt

first: Shankar

last: Subramaniam


title: High-dimensional profiling uncovers heterogeneity, lineage-specific precursors and inflammation-induced changes in the mononuclear phagocyte compartment of the human intestine

city: Copenhagen

first: William

middle: Winston

last: Agace

address: Blegdamsvej 3B

department: Skin Immunology Research Center

country: Denmark

institute: Copenhagen University

summary: With use of single-cell omics this study explore mononuclear phagocyte complexity of the human intestinal lamina propria in health and IBD. The study identifies distinct dendritic cell and macrophage subsets, lineage specific precursors and state specific signatures.

overall_design:
Healthy controls consist of resection material from colorectal cancer patients undergoing surgery, samples are taken away from cancer site. IBD samples are similarly taken from patients undergoing surgery, material is from inflamed areas in this case.

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

first: Thomas

last: Fenton

first: Line

last: Wulff

first: Venla

last: Väänänen

first: Gareth-Rhys

last: Jones

first: Camilla

middle: K

last: Lemvigh

first: Lene

last: Riis

first: Mads

last: Wewer

first: Julien

last: Vandamme

first: Peter

last: Jørgensen

first: Calum

middle: C

last: Bain

first: Julie

last: Lee

first: Kirstine

middle: G

last: Belling

first: Gwo-Tzer

last: Ho

first: Tune

middle: H

last: Pers

first: Anja

last: Poulsen

first: Gorm

last: Madsen

first: Ole

last: Nielsen

first: Henrik

middle: L

last: Jakobsen

first: Jose

last: Izarzugaza

first: Flemming

last: Bendtsen

first: Søren

last: Brunak

first: Allan

last: Mowat

first: Lars

last: Olsen

first: Urs

last: Mörbe

middle: W


title: Single cell and isoform-specific translational profiling of the mouse brain

first: Brian

last: Yee

address: 9500 Gilman Dr

department: Health

institute: UCSD

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

overall_design: Refer to individual Series


title: DeCAF redefines fibroblast states uncovering multidimensional tumor-stroma relationships driving clinical tumor progression and immunotherapy response

city: Chapel Hill

first: Jen Jen

last: Yeh

state: North Carolina

address: 450 West drive

institute: The University of North Carolina at Chapel Hill


title: Simultaneous Targeting of KRAS and CDK4 Synergistically Induces Durable Growth Arrest in Pancreatic Cancer Cells

city: Göttingen

first: Adi

last: Mackay

address: Robert-Koch-Straße 40

department: Pathology

country: Germany

institute: University Medical Center Göttingen

summary: Mutant Ras oncoproteins, particularly KRAS, are among the most prevalent drivers of cancer. Small-molecule KRAS inhibitors have emerged as promising cancer therapeutics, yet resistance development remains a major hurdle. To overcome this challenge, we explored rational combination strategies aimed at enhancing therapeutic efficacy and durability. We show that the KRAS-G12C inhibitor Sotorasib synergizes with the CDK4/6 inhibitor Palbociclib to eliminate pancreatic ductal adenocarcinoma (PDAC) cells and organoids harboring KRAS-G12C mutations. This synergy was especially pronounced following drug washout, indicating a durable cellular response. Similar synergistic effects were observed in non-small-cell lung cancer (NSCLC) cells. Additionally, the KRAS-G12D inhibitor MRTX1133 cooperated with Palbociclib to suppress KRAS-G12D-mutant PDAC cells. Mechanistically, the combinations induced sustained cell cycle arrest, marked by reduced RB phosphorylation, decreased E2F1 expression, and increased levels of CDKN1B/p27. Deletion of CDKN1B largely reversed the growth-inhibitory effect, highlighting its essential role in mediating the observed synergy. In an orthotopic, immunocompetent mouse model of PDAC, MRTX1133 significantly delayed tumor growth and extended survival; however, despite its ability to suppress RB phosphorylation, Palbociclib failed to enhance these effects. This might be due to increased tumor vascularization upon Palbociclib treatment, as revealed by single cell RNA sequencing. In summary, our findings demonstrate the therapeutic potential of enhancing cell cycle restriction point activation in KRAS inhibitor-based therapies, while emphasizing the importance of precisely adjusting the drug combinations to the in vivo situation.

overall_design: 1.5 x 10^5 KrasG12D;Trp53R172H/+ (KPC) cells in culture medium and 50 % matrigel were injected into the pancreatic tail of C57BL/6J mice aged 4 to 6 months. Upon successful tumor detection, mice were assigned to treatment groups: Vehicle-control, Palbociclib (25 mg/kg, PZO199, Sigma-Aldrich) and/or MRTX1133 (30 mg/kg, HY-134813, Medchem Express). The treatments were administered i.p. once a day within weekly cycles of three consecutive treatmens followed by four recovery days until humane endpoint.


title: A Cellular Basis for Heightened Gut Sensitivity in Females

first: Holly

last: Ingraham

address: 1550 4th Street Rock Hall 284F

department: Cellular and Molecular Pharmacology

institute: University of California, San Francisco

summary: Visceral pain disorders, such as irritable bowel syndrome, exhibit a marked female prevalence. Enhanced signaling between enterochromaffin (EC) cells in the gut epithelium and mucosal sensory nerve fibers likely contributes to this sex bias. Here, we identify a novel estrogen-responsive paracrine pathway in which two enteroendocrine cell types, PYY-expressing L-cells and serotonergic EC cells, communicate to increase gut sensitivity in females. We demonstrate that ERα estrogen signaling upregulates the bacterial metabolite SCFA receptor Olfr78 on colonic L-cells, increasing PYY release and their sensitivity to acetate. Elevated PYY acts on neighboring EC cells via NPY1R, thereby enhancing serotonin release and gut pain. We propose that hormonal fluctuations, in conjunction with internal (stress) or environmental (diet) factors, amplify this local estrogen-responsive colonic circuit, resulting in maladaptive gut sensitivity.

overall_design: RNA-seq was performed on L-cells and EC cells isolated from colons of 10–14-week-old ovariectomized (OVX) female mice. L-cells were obtained from OVX Cck-Cre;R26LSL-tdTomato mice treated with sesame oil vehicle (VEH) or estradiol benzoate (EB) 4 hours prior to isolation. Approximately 7,000 L-cells were collected per treatment group (OVX + VEH and OVX + EB). EC cells were obtained from OVX ECFLTG mice treated with VEH or EB, with at least 500 EC cells isolated per sample and processed in the same manner.

first: Archana

last: Venkataraman

first: Eric

last: Figueroa

first: Joel

last: Castro

first: Fernanda

last: Castro Navarro

first: Deepanshu

last: Soota

first: Stuart

last: Brierley

first: David

last: Julius


title: Sperm RNA landscape during sexual maturation in Duroc boars

city: Hamar

first: Asmita

last: Shrestha

state: Innlandet

address: Holsetgata 22

department: Department of Biotechnology

country: Norway

institute: University of Inland Norway

type: Non-coding RNA profiling by high throughput sequencing

summary:
Sexual maturation in boars impacts reproductive efficiency in swine production, yet the molecular mechanisms underlying this developmental transition remain poorly understood. This study aimed to investigate the transcriptomic changes in sperm from Duroc boars during sexual maturation, conducting a longitudinal analysis. The total RNA and miRNA profiles from the same individuals (n = 6) at puberty (7.24 ± 0.39 months) and sexual maturity (10 ± 0.40 months) were compared, identifying molecular signatures associated with reproductive development. Total RNA sequencing (Illumina NovaSeq-6000) and miRNA sequencing (Illumina NextSeq-500) were performed on all 12 paired samples (6 boars at 2 time points), followed by differential expression analysis using a paired statistical model in DESeq2 to account for repeated measures.
Differential expression analysis identified 60 differentially expressed genes using stringent criteria (adj P < 0.05, |log₂FC| ≥ 0.5), with 65% upregulated in sperm of boars 10-months versus 7-months of age. Key upregulated genes included NCLN, RGS12, CIB2, FOXP4, PHC1, CDC25B and AKAP1, while HSP90AA1, EVI5, FSIP2, VDAC3, and ALMS1 were key downregulated genes. Furthermore, Gene ontology analysis revealed significant enrichment of 11 biological processes, mostly related to reproductive development, and four molecular functions (transferase activity, transferring phosphorus-containing groups, protein serine/threonine kinase activity, protein kinase activity and signal sequence binding). Additionally, our miRNA analysis identified six differentially expressed miRNAs (adj P < 0.05 & |log2FC| ≥ 0.5); ssc-miR-193a-5p, ssc-miR-574-3p, ssc-miR-126-3p, ssc-miR-196a, ssc-miR-210 were upregulated, and ssc-miR-338 showed downregulation in 10-months age. Integrated analysis of differentially expressed mRNA and predicted miRNA targets identified 18 miRNA-mRNA regulatory pairs, enriched in pathways related to cell cycle processes and chromatin binding, suggesting coordinated regulation across RNA biotypes in sperm during sexual maturation.
Our study reveals coordinated transcriptomic shifts in boar sperm during sexual maturation. Most genes show increased abundance at 10 months of age, with enrichment in terms of reproductive development. Several miRNAs appear to regulate these changes through targeted mRNA interactions. These findings expand our understanding of the

overall_design: Total RNA-seq and miRNA-seq profiling of sperm samples from six Duroc boars collected at pubertal (~7 months) and mature (~10 months) ages using a longitudinal paired design.

first: Maren

middle: V

last: Son

first: Adnan

last: Hashim

first: Soudabeh

last: Rouzbehani

first: Gregor

last: Gilfillan

first: Urszula

last: Berge

first: Elisabeth

last: Kommisrud

first: Anne

last: Alm-Kristiansen


title: Specific SLC25 carriers regulate mitochondrial protein synthesis

city: Nedlands

first: Danielle

middle: Louise

last: Rudler

state: WA

address: 6 Verdun Street

department: Mitochondrial Medicine and Biology

country: Australia

institute: The Kids Research Institute

summary: A genome-wide knock out screen identified members of the SLC25 family of mitochondrial carrier proteins as important regulators of the rate of de novo mitochondrial protein synthesis. To elucidate this relationship, we generated human cell knockouts for SLC25A25, SLC25A44, SLC25A45, and SLC25A48, which have been shown to exchange ATP and magnesium, branched-chain amino acids, methylated basic amino acids, and choline, respectively.  Multi-omic and functional analyses identified that these four carriers are crucial for mitochondrial translation, biogenesis and function of the oxidative phosphorylation system, as well as mitochondrial morphology. Thermostability screens showed that SLC25A48 is specifically stabilised by choline and changes in the mitochondrial metabolome and lipidome indicated defects in choline biosynthetic pathways and remodelling of mitochondrial membranes, both consistent with SLC25A48 being a choline transporter. These results highlight the essential roles of specific SLC25 transporters in maintaining mitochondrial structure and function and show that of mitochondrial translation is sensitive to local concentrations of branched chain amino acids, methylated basic amino acids, ATP, magnesium, and choline.

overall_design: Total RNA from CAL51 cells were isolated from three control cells (WT) and three of each knockout (SLC25A44KO, SLC25A45KO, SLC25A48KO) and was sequenced and analysed for differential expression.

first: Aleksandra

last: Filipovska


title: DeCAF redefines fibroblast states uncovering multidimensional tumor-stroma relationships driving clinical tumor progression and immunotherapy response [scRNA-Seq]

first: Jen Jen

last: Yeh

summary: Cancer-associated fibroblast (CAF) heterogeneity within pancreatic ductal adenocarcinoma (PDAC) has been previously assessed through single-cell RNA sequencing (scRNAseq) to discover distinct CAF populations such as myCAFs and iCAFs. While useful as a biological framework, no studies have conclusively and robustly demonstrated a correlation of CAF subpopulations with clinical prognosis or therapy response. We define restraining (rest) and promoting (pro) CAFs in patient samples that are both prognostic and predictive of therapy response in disparate tumor types. Using a single sample classifier, DeCAF, we uncover distinct clinical and spatial interactions between pro- and restCAF subtypes and basal-like and classical tumor subtypes that support tumor stroma crosstalk where basal-like tumors are enriched in proCAFs and classical tumors are enriched in restCAFs. Finally, we find striking differences in the immune contexture of pro- and restCAF tumors where restCAF dominant tumors are enriched in patients who respond to immune checkpoint inhibition (ICI) in two large clinical trials, and proCAF dominant tumors are enriched in patients who respond to myeloid inhibition. This work is the first to define CAF subtypes that are clinically robust, prognostic, predictive of immunotherapy response, and associate with tumor molecular subtype.

overall_design:
129 de-identified primary PDAC patient samples were collected from the IRB-approved University of North Carolina Lineberger Comprehensive Cancer Center Tissue Procurement Core Facility and were flash frozen in liquid nitrogen.  FFPE samples were prepared, hematoxylin and eosin stained.

***************************************************************
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: Xianlu

last: Peng

first: Ian

last: McCabe


title: DeCAF redefines fibroblast states uncovering multidimensional tumor-stroma relationships driving clinical tumor progression and immunotherapy response [Spatial Transcriptomics]


title: Activating mutations in ESR1 contribute to an immunosuppressive breast tumor microenvironment.

city: Montreal

first: Alain

last: Pacis

state: QC

address: 1010 Sherbrooke #1800

country: Canada

institute: Canadian Centre for Computational Genomics (C3G)

summary: Estrogen receptor (ER, ESR1)-positive breast cancer patients are most at risk of long-term relapse, where activating somatic mutations in ESR1 promote metastasis and therapeutic resistance. ER+ patients are also disadvantaged in receiving and responding to immunotherapies, the mechanisms of which remain to be elucidated. Here, we engineered a novel transgenic mouse model carrying either Y541S or D542G point mutation in ESR1, mirroring the two most common mutations in ER+ metastatic breast cancer patients. End-stage ESR1mut tumors do not differ in the total number of immune cells yet display downregulation in immune-mediated pathways and decreased immune-modulatory cytokines, including IL-17a and IL-1b. Furthermore, T cells and macrophages have lower IFNγ activity and antigen presentation, respectively. Mechanistically, ESR1mut negatively regulates immune modulator expression compared to wild-type ESR1 and upregulates Stat5 to further dampen cytokine expression. In concordance, clinical validation on ESR1mut patient tumors shows decreased IL-17a and IL-1b. Collectively, our findings reveal that activating mutations in ER contribute to an immunosuppressive tumor microenvironment by dampening cytokine secretion and immune cell activity, supporting clinical observations where hormone receptor patients, especially ER mutant patients, respond poorly to immunotherapies.

overall_design: RNA-seq profiling MIC WT, MIC ESR1-Y541S, and MIC ESR1-D542G endpoint mammary tumors.

first: Yu

last: Gu

first: William

last: Muller


title: Hypoxia-activated scleraxis a mediates epicardial progenitor differentiation into a unique cardiac perivascular cell type

city: New York

first: Jingli

last: Cao

state: NY

department: Cell & Developmental Biology

institute: Weill Cornell Medical College

type: Genome binding/occupancy profiling by high throughput sequencing

summary: The epicardium is a crucial source of progenitor cells and paracrine signals that support heart development and regeneration. However, the molecular mechanisms that guide epicardial cell fate decisions remain incompletely understood. Here, we identify the transcription factor Scleraxis a (encoded by scxa) as a key regulator of epicardial progenitor differentiation in zebrafish. Through single-cell transcriptomics, genetic lineage tracing, and cardiac injury models, we show that scxa is transiently induced in activated epicardial progenitor cells (aEPCs) during both heart regeneration and developmental coronary angiogenesis. scxa+ epicardial cells primarily give rise to a previously uncharacterized cardiac population of perivascular cells marked by col18a1a, molecularly distinct from classical pericytes and vascular smooth muscle cells. We refer to this population as epicardial-derived perivascular mesenchymal cells (Epi-PMCs). These Epi-PMCs closely associate with coronary vessels and may contribute to vascular stabilization and remodeling, potentially through the anti-angiogenic but vessel-stabilizing activity of endostatin derived from collagen XVIII. Loss of scxa increases coronary vessel density. Mechanistically, we identify hypoxia and Hif1a signaling as upstream regulators of scxa, with systemic hypoxia or Hif factor stabilization robustly inducing scxa expression in the epicardium. Together, these findings uncover a hypoxia-responsive Scxa-Col18a1a axis that drives epicardial differentiation toward a vascular-supportive fate, offering new insight into the regulation of coronary vessel development and the regenerative potential of the epicardium.

overall_design: CUT&Tag of histone modification H3K4me1 of epicardial progenitor cells. Heart amputation surgery was performed on adult fish (6 months of age) carrying tcf21:nucEGFP and ptx3a:mScarlet reporters. Hearts were isolated at 3 days post injury together with the uninjured control (2 biological replicates each). Ventricles were digested and EGFP+ cells (uninjured) and EGFP+mScarlet+ (3 dpa) cells were isolated for CUT&Tag processing.

first: Jun

last: Yao

last: Xia


title: DeCAF redefines fibroblast states uncovering multidimensional tumor-stroma relationships driving clinical tumor progression and immunotherapy response [RNA-Seq]


title: The WNK-OXSR1 osmosensing pathway mediates intestinal regeneration via Hippo-YAP signaling

city: Xiamen

first: Xiawei

last: Huang

state: Fujian

address: Xiang an nan lu 4221

institute: Xiamen university

summary: Animals activate regenerative processes to repair injuries and restore homeostasis following tissue damage. A central question in regeneration is how damage signals are sensed and translated into regenerative growth. Tissue injuries lead to the release of intracellular contents and bodily fluids and disturb the osmotic balance. However, the role of osmolarity in regeneration remains largely unexplored. Using Drosophila and mouse intestine, as well as samples from inflammatory bowel disease (IBD) patients, we identify a key role for the osmolarity-sensing WNK-OXSR1 kinase cascade in intestinal regeneration. Mechanistically, OXSR1 phosphorylates the RhoB GTPase at threonine 37 upon intestinal injury, thereby disrupting its interaction with ARHGAP17 and increasing the levels of GTP-bound RhoB. RhoB activation in turn leads to enhanced F-actin polymerization and YAP activation, thus promoting tissue regeneration. We further show that pharmacological inhibition of WNK or OXSR1 reduces the oncogenic potential of intestinal regeneration. These findings reveal osmolarity as a critical damage signal in regeneration and position WNK-OXSR1 as a potential therapeutic target for stimulating intestinal repair.

overall_design: RNA was extracted using Trizol Reagent (Invitrogen) and purified using RNeasy Mini Kit (Qiagen) following the kit manual. Library construction, sequencing and bioinformatics analysis were performed by MegaGenomics Inc. (Beijing, China). The P value was calculated by Deseq2 (v1.26.0). The differentially expressed genes (DEGs) were defined as adjusted p-value < 0.05 and |log2(FoldChange)|>0.5 Three biological replicates were performed for WT and Oxsr1 KO HEK293T cells.

first: Bo

last: Liu

first: He ming

first: Xiaobing

last: Jiang

first: Jingrong

last: Deng

first: Jiahui

last: Wang

first: Chengfang

last: Wu

first: Minhuang

last: Hu

first: Bei

last: Zeng

first: Zhihao

first: Huimin

last: Pan

first: Yuxia

last: Yang

first: Kewei

last: Zheng

first: Rui

last: Shen

first: Mingqing

last: Zhang


title: Spaceflight induces systemic effects on extracellular matrix and immune system in different age-stages

city: Saarbrücken

first: Friederike

last: Grandke

address: Saarbrücken Campus

institute: Saarland University

summary: Spaceflights significantly impacts astronaut health, causing muscle atrophy, bone loss, cardiovascular deconditioning and immune system dysregulation, especially during long missions. To explore molecular changes, we generated a single-cell RNA sequencing dataset, of 216 samples from 28 mouse organs and tissues under spaceflight and control conditions. Since spaceflight effects closely resemble aspects of aging, we included two age groups (3 and 8 months) to assess age-related influences. Our data revealed that spaceflight drives age-independent systemic changes, primarily affecting tissue remodeling and alterations in cytoskeleton, membrane and extracellular matrix (ECM), particularly in endothelial cells. In contrast, aging has a weaker impact, mainly altering immune regulation in an age-dependent manner. These findings suggest that cytoskeletal and ECM changes may contribute to immune Dysregulation.

overall_design: We performed single-cell RNA-sequencing (scRNA-seq) on organ samples from four female Balb/C mice exposed to 40 days of spaceflight on the ISS, alongside four age-matched controls housed under identical conditions on earth (Figure 1A, 1B). Our study design incorporated two age groups, 3-months-old (young adult) and 8-months-old (mature adults), capturing the typical age range of astronauts. Three days after splashdown, the mice were taken down, and the tissues were immediately processed for single-cell RNA sequencing.

first: Nicholas

last: Schaum

first: Fabian

last: Kern

first: Jérémy

last: Amand

first: Thomas

last: Ambrosi

first: Teni

last: Anbarchian

first: Karl

last: Annusver

first: Reza

last: Ashrafi

first: Kruti

last: Calcuttawala

first: Shubhangi

last: Das Barman

first: Antoine

last: de Morree

first: Angela

last: Detweiler

first: Hala

middle: S

last: Dhowre

first: Annika

last: Engel

first: Tobias

last: Fehlmann

first: Virginia

last: Ferguson

first: Matthias

last: Flotho

first: Astrid

last: Gillich

first: Michael

last: Haney

first: Yan

last: Hang

first: Pascal

last: Hirsch

first: Taichi

last: Isobe

first: Joanna

last: Kalucka

first: Maria

last: Kasper

first: Seung

last: Kim

middle: A

last: Kluiver

first: Davis

last: Lee

first: Song

first: Olivia

last: Leventhal

first: Ling

first: Catriona

middle: Y

last: Logan

middle: F

last: Lugo-Fagundo

first: Luca

last: Mannino

first: Maxim

last: Markovic

first: Maurizio

last: Morri

first: Shravani

last: Mukherjee

first: Norma

last: Neff

first: Patricia

last: Nguyen

first: Róbert

last: Pálovics

first: Weng

last: Peng

first: Maksim

last: Plikus

first: Ermelinda

last: Porpiglia


title: Prenatal and postnatal effects of gestational immune activation on synaptic and neurodevelopmental pathways via epigenetic mechanisms (RNA-seq)

city: Blacksburg

first: Chang

last: Lu

state: VA

address: 235 Goodwin Hall, 635 Prices Fork Road, Virginia Tech

department: Chemical Engineering

institute: Virginia Tech

summary: Previous epidemiological research suggests that maternal immune activation (MIA) during early pregnancy is a significant risk factor for neurodevelopmental and psychiatric disorders in offspring. Epigenetic factors and chromatin-related phenomena remain largely plastic during the course of prenatal and early postnatal development, allowing a dynamic impact of environmental effects on access to genes and regulatory elements. In this study, we tested the effects of maternal infection with the mouse-adapted influenza A/WSN/33 (H1N1) virus as a MIA model. Additionally, to assess the prenatal and postnatal effects of MIA, we cross-fostered half of the neonatal mice immediately after birth.This study includes RNA-seq and ChIP-seq profiling (H3K27ac, H3K4me3) from the prefrontal cortex of MIA-exposed and control mice. RNA-seq and ChIP-seq data are submitted under separate Series and linked via sample annotations.

overall_design: This study examined transcriptomic and epigenomic changes in offspring following maternal immune activation (MIA) via prenatal influenza infection. Timed pregnant CD1 mice were intranasally infected with 5 × 10³ pfu of influenza A/WSN/33 (H1N1) or mock-treated on embryonic day 9.5. At birth, ~50% of pups were cross-fostered to surrogate dams (MIA or control) to separate prenatal and postnatal influences. Offspring were weaned at P21, housed by sex, and assigned to one of seven groups based on birth and rearing conditions: Group A (Mock-Mock): Offspring born to control mother and raised by their birth mother, together with stepsiblings born to MIA-exposed mother. Group B (MIA-Mock): Offspring born to MIA-exposed mother but raised by control surrogate mother, together with stepsiblings born to this control mother. Group C (Mock-MIA): Offspring born to control mother but raised by MIA-exposed surrogate mother, together with stepsiblings born to this MIA mother. Group D (MIA-MIA): Offspring born to MIA-exposed mother and raised by their birth mother, together with stepsiblings born to control mother. Group E (Mock-Mock_1): Offspring born to control mother and raised by their birth mother, together with stepsiblings born to a different control mother. Group F (Mock-Mock_2): Offspring born to control mother but raised by control surrogate mother, together with stepsiblings born to the control surrogate mother. Group G (Mock-Mock_3): Offspring born to control mother and raised by their birth mother with no stepsiblings. Mice were sacrificed in adulthood. Frontal cortex (bregma 1.90–1.40) was collected for RNA-seq. Neuronal nuclei were isolated and sorted via FACS (NeuN⁺);  ~6,000 nuclei per RNA-seq replicate. Libraries were prepared using SMART-seq v4 and Nextera XT (RNA-seq), and sequenced on an Illumina HiSeq 4000 (single-end, 50 bp), yielding ~11M per RNA-seq library.

first: Bohan

last: Zhu

first: Gaoshan

last: Li


title: MicroRNAs shape mouse age-independent tissue adaptation to spaceflight via ECM and developmental pathways

first: Friederike

last: Grandke

summary: As human space exploration accelerates, understanding the organism-wide molecular effects of longer spaceflight in mammals becomes increasingly critical. Non-coding RNAs like miRNAs are key to regulating this landscape. We thus analyzed 686 small RNA samples of mice from 13 solid organs at 3 and 8 months of age, after at least 3 weeks on the ISS and compared them to earth-bound controls. We observed significant spaceflight effects in systemic tissue remodeling pathways along the Fat-Liver-Pancreas axis and in heart, brain, spleen and thymus. The MIR-17/92 and MIR-1/133 families drive distinct molecular changes through specific gene targeting. Age-dependent changes, smaller in magnitude compared to age-independent changes, primarily involved tissue remodeling through MIR-8, MIR-154 and MIR-15 families in MAT, pancreas, and diaphragm. Our findings provide evidence on how spaceflight regulates mammalian gene expression in preparation for interplanetary spaceflight.

overall_design: We sequenced 686 samples across 13 organs of young (3 months) and middle-aged (8 months) mice that were sent to the ISS (Flight). We compared them against mice living in standard conditions (Vivarium Ground Control) and mice living in an environment matched to ISS conditions (Habitat Ground Control). We euthanized mice at two time-points (matching timelines for controls and flight mice), one before returning to earth (TERM) and one after (LAR) in order to distinguish spaceflight-induced effects from the reentry-induced stress

first: Shusruto

last: Rishik

first: Viktoria

last: Wagner

first: Annika

last: Engel

first: Nicole

last: Ludwig

first: Kruti

last: Calcuttawala

first: Fabian

last: Kern

first: Verena

last: Keller

first: Marcin

last: Krawczyk

first: Louis

last: Stodieck

first: Virginia

last: Ferguson

first: Amanda

last: Roberts

first: Eckart

last: Meese

first: Nicholas

last: Schaum

first: Steven

last: Quake

first: Tony

last: Wyss-Coray

first: Andreas


title: The ADAR G1007R mutation in Aicardi-Goutières Syndrome causes neuroinflammation through RNA sensing signaling pathways in mice

city: Pittsburgh

first: Shuchang

last: Liu

address: 203 Lothrop Street

institute: University of Pittsburgh

summary: The ADARG1007R mutation is one of the most frequent mutations found in type six Aicardi-Goutières Syndrome (AGS), a progressive inflammatory encephalopathy in pediatric patients, typically begins to manifest  in newborns or infants. Due to the rarity of this disease and the limited accessibility of the human brain tissues, the pathologic features and the underlying mechanism of this mutation has not been well understood.  In this study, we generated a genetic mutant mouse model which successfully recapitulated the genetic and pathologic features of human ADARG1007R AGS, including the early-onset brain inflammatory reaction in heterozygous mutant mice and the deep gray matter area ISG expression. Through analysis of the homozygous ADARG1007R/G1007 mice, we found that the G1007R mutation interrupts the RNA splicing donor site in the RNA transcripts that alters ADAR1 RNA splicing, leading to depletion of ADAR1 protein and embryonic death. Furthermore, we demonstrated that the brain inflammation could be reversed by deletion of the cellular RNA receptor MDA5 which blocks the cellular RNA sensing signaling pathway. Collectively, this study generated a clinically relevant AGS animal model, revealed the molecular mechanism of the ADARG1007R mutation in AGS, and demonstrated that the MDA5-dependent RNA sensing signaling pathway plays a critical role in neuroinflammation.

overall_design: All mice used in this study were kept in a specific pathogen-free animal facility and all studies were carried out following protocols approved by our Institutional Animal Care and Use Committee. The G1007R mutation was introduced into the mouse genome using our CRISPR/Cas9 protocol as described previously. Considering the genetic diversity of human diseases of ADAR1 mutations, mice used in this study, including AdarG1007R/G1007R and AdarG1007R/G1007R; Ifih1-/- mice, were purposely maintained in C57/BL6J and DBA2 mixed background. Three repeats for wild-type (WT), three repeats for G1007R mutant, and three repeats for G1007R mutatant + MDA5 retreat

first: XinFeng

last: Guo

first: Jia-Jun

first: Chaowei

last: Shang

first: Christopher

middle: J

last: Guerriero

first: Clayton

last: Wiley

first: Richard

last: Steinman

first: Yi

last: Sheng

first: Jeffrey

last: Brodsky

first: Timothey

last: Billiar

first: Silvia

first: Qingde

last: Wang


title: Autophagy is required for the development and functionality of lacrimal gland-like organoids

city: Birmingham

first: Gamze

last: Kocak

address: Edgbaston

country: United Kingdom

institute: University of Birminghham

summary: Lacrimal glands (LGs) serve as pivotal exocrine glands crucial for protecting the ocular surface. Dysfunction in LG cell composition or secretion is implicated in dry eye disease (DED). While autophagy plays a vital role in tissue homeostasis in many organs, how it affects LG development and secretory function is not known. Here, we have undertaken a genetic study by utilizing autophagy-deficient human embryonic stem cells (hESCs) and differentiating them into LG-like organoids. Autophagy-deficient LG-like organoids exhibited improper development and secretion, along with increased protein aggregation, proliferation, and cell death. These phenotypes were associated with an accumulation of PAX6, a transcription factor crucial for brain and eye development, which we identified as an autophagy substrate. Pharmacological interventions with nicotinamide mononucleotide (NMN) and melatonin were able to rescue the cellular dysfunction in autophagy-deficient LG-like organoids. Together, our study highlights the role of autophagy in LG along with potential therapeutic interventions for DED.

overall_design:
To investigate the role of autophagy in LG-like organoids, we used multi-zonal eye cells (SEAM) differentiation from ATG5 WT and KO hESCs.
We then performed gene expression profiling analysis using data obtained from bulk RNA-seq of  ATG5 WT and KO hESC-derived SEAM.
Comperative gene expressing profiling of RNA-seq data for  ATG5 WT and KO hESC-derived SEAM.


title: Transcriptome of Hepato-Hematopoietic Lineages in Fetal Liver Organoids from Human Pluripotent Stem Cells

city: Cincinnati

first: Kentaro

last: Iwasawa

state: OH

address: 3333 BURNET AVENUE

department: Division of Gastroenterology, Hepatology and Nutrition

institute: Cincinnati Children's Hospital Medical Center

summary: To investigate the functional difference between different cell lineages in highly hematopoietic fetal liver organoids (hFLOs) from induced pluripotent stem cell, we FACS-sorted cell lineages from hFLOs and compared them to conventional iPSC-monocellular or primary cell controls by bulk RNA-sequencing.

overall_design: We then performed gene expression profiling analysis using data obtained from RNA-seq of FACS-isolated lineages of hematopoietic fetal liver organoids (HLOs) from two different iPSC lines, and controls (PBMC-derived macrophages, primary hepatocytes, primary postnatal liver macrophages, iPSC-derived monocytes and -macrophages from a third iPSC-line

first: Milad

last: Rezvani

first: Kyle

last: Lewis

first: Konrad

last: Thorner

first: Takanori

last: Takebe


title: RAB3GAP2 is a Regulator of Skeletal Muscle Endothelial Cell Proliferation and Associated with Capillary Density

status: Public on Dec 17 2025

city: Malmo

first: Dmytro

last: Kryvokhyzha

address: Jan Waldenströms gata 35

department: Clinical Sciences

country: Sweden

institute: Lund University

summary: Skeletal muscle fiber composition and capillary density influence physical performance and whole-body metabolic properties. ~45% of the variance in fiber type is heritable, which motivated us to perform a genome-wide association study of skeletal muscle histology from 656 Swedish men. Four independent variants were associated (p < 5x10− 8) with proportion of type IIx fibers or capillary-to-fiber ratio (C:F). The strongest signal localized to the rs115660502 variant, where the G-allele corresponded with increased C:F and reduced skeletal muscle expression of the proximal gene, RAB3 GTPase Activating Non-Catalytic Protein Subunit 2 (RAB3GAP2). The G-allele was less frequent in elite short-track sprinters and more frequent in endurance athletes than in matched non-athlete (population) controls; RAB3GAP2 expression was reduced by high-intensity intermittent training. RAB3GAP2 protein was not uniformly expressed in muscle tissue but localized to the endothelium and capillaries. Experimental reduction of RAB3GAP2 in human endothelial cells led to increased tube formation in vitro, to regulation of secreted factors promoting angiogenesis and T-cell activation, to reduced intracellular levels of von Willebrand factor (VWF) and, post-implantation, to increased endothelial cell density in vivo in mice. The amount of RAB3GAP2 in skeletal muscle was positively associated with exercise-induced release of VWF in vivo in humans. By regulating the release of protein factors (VWF, CD70, TNC, TNXB, MCP1, IGFBP3, COL1A1, TFPI2 and tPA), RAB3GAP2 influences fitness adaptation after exercise by improving muscle healing and promotion of capillary formation.

overall_design: HMEC cells were sequenced with RNAseq to identify an angiogenic factor that is secreted more from the KD-cells vs controls.


title: Tenascin C+ myofibroblasts exacerbate vascular neointimal hyperplasia by propagation of nerve-macrophage interactions

city: Shanghai

first: Guangzheng

last: Shi

state: Shanghai

address: No. 197 Ruijin 2nd Road, Huangpu District, Shanghai

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

summary: Neointimal hyperplasia (NIH) is a major pathological feature of vascular injury, yet the underlying cellular and molecular drivers remain poorly understood. Here, we combine lineage tracing, single-cell transcriptomics, genetic mouse models and live imaging to identify a population of Tenascin C-expressing (Tnc+) myofibroblasts (MFs) that emerges in both a ligation-induced mouse model of carotid artery NIH and clinical NIH specimens.

overall_design: Carotid arteries from Tnc-cKO mice were harvested two weeks post vascular ligation. Arteries were enzymatically digested to single-cell suspension, and then scRNA-seq libraries were built with the 10x Genomics platform.

first: Qun

last: Li


title: Non-hematopoietic erythropoietin splice variants are produced in the diseased human brain and confer neuroprotection

city: Berlin

first: January

last: Weiner

address: Charitéplatz 1

department: CUBI

institute: Berlin Institute of Health, Charité Medical University of Berlin

summary: Erythropoietin (EPO) is a pleiotropic cytokine with important functions in neuronal development and neuroprotection, but hematopoietic effects limit the therapeutic application of EPO in neurological diseases. We discovered human endogenous EPO splice variants that are non-hematopoietic but cytoprotective. Here, we demonstrate at the single-cell level that an alternative splice variant lacking exon 3 (hS3) is expressed in the human brain and is upregulated above EPO mRNA levels in ischemic and inflammatory neurological diseases. Conversely, hS3 mRNA expression is reduced below EPO levels in neurodegenerative disease. In an oxygen-glucose deprivation (OGD) model of ischemia, a single dose of cell-free synthesized constant glycosylated active hS3 protects neuronal cultures derived from human induced pluripotent stem cells (hiPSC) and human embryonic stem cells (hESC) more effectively than EPO. We identify the D-helix as a key functional domain of hS3 and demonstrate that the neuroprotective effect is enhanced by PD29, a novel small peptide derived from the D-helix of hS3. Long-term hS3 administration increases the neuroprotective effects in the OGD model by dose-dependent differential expression of apoptosis-related protein-coding genes and long non-coding RNAs (lncRNAs). In addition, our results suggest that hS3 induces early cell cycle inhibition without impairing differentiation of hiPSC and hESC into neuronal subtypes. In conclusion, EPO splice variants are part of the endogenous neuroprotective system in the human brain with significant therapeutic potential.

overall_design: Two cell lines (MDC_P4_H1 and BIH005_A) were subjected to two types of treatment (hS31 or hS32) or left untreated. RNA was collected at one of four different time points.

first: Theresa

last: Hartung

first: Andreas

last: Meisel

first: Josef

last: Priller


title: Sequence and chemical specificity define the functional landscape of intrinsically disordered regions [MNase-seq]

city: Garching

first: Frauke

last: Huth

address: James-Franck Str. 1

department: Department of Biosciences

institute: Technical University Munich

summary: Intrinsically disordered regions (IDRs) pervasively engage in essential molecular functions, yet they are often poorly conserved as assessed by sequence alignment. To explore the seeming paradox of how sequence variability is compatible with persistent function, we examined the functional determinants for a poorly conserved but essential IDR. We show that IDR function depends on two distinct but related properties: sequence and chemical specificity. While sequence-specificity operates via binding motifs and depends on the precise order and identity of residues, chemical specificity reflects the sequence-encoded chemistry of multivalent interactions across an IDR and depends on local and global chemical properties. Unexpectedly, a binding motif essential in the wild- type IDR can be removed when compensatory changes to the sequence chemistry are introduced, highlighting the orthogonality and interoperability of these properties and expanding the sequence space compatible with function. Our results provide a general framework for the functional constraints on IDR evolution.

overall_design: The effects of WT Abf1, Abf1 variants and no Abf1 on nucleosome positioning and gene regulation were investigated through an Anchor-Away approach in vivo, read out via methylation-aware Nanopore-Sequencing or total RNA-Sequencing as well as through an in vitro Remodeller assay, read out using MNase-Sequencing.

first: Iris

last: Langstein-Skora

first: Andrea

last: Schmid

first: Drin

last: Shabani

first: Lorenz

last: Spechtenhauser

first: Mariia

last: Likhodeeva

first: Franziska

last: Kunert

first: Felix

last: Metzner

first: Ryan

last: Emenecker

first: Mary

middle: O

last: Richardson

first: Wasim

last: Aftab

first: Maximilian

last: Götz

first: Sarah

last: Payer

first: Niccoló

last: Pietrantoni

first: Valentina

last: Sjeničić

first: Sakthi

last: Ravichandran

first: Till

last: Bartke

first: Karl-Peter

last: Hopfner

first: Ulrich

last: Gerland

first: Philipp

last: Korber

first: Alex

last: Holehouse


title: Sequence and chemical specificity define the functional landscape of intrinsically disordered regions [RNA-seq]

first: Frauke

last: Huth

first: Iris

last: Langstein-Skora

first: Andrea

last: Schmid

first: Drin

last: Shabani

first: Lorenz

last: Spechtenhauser


title: Rational redesign of antigen binding domain improves in vivo efficacy of the CD22-CAR

city: Aurora

first: Terry

last: Fry

state: CO

institute: University of of Colorado Denver and Children's Hospital Col

summary: Chimeric antigen receptor (CAR) T cells targeting CD19 are highly effective against B-lineage malignancies. However, about half of patients either fail to achieve complete remission (in the case of lymphoma) or relapse (in the case of acute lymphoblastic leukemia). CD22 represents an alternative highly B-lineage-restricted target. Although CD22-targeted CAR T cells are clinically active, targeting this antigen has proven difficult relative to CD19, attributable, in part, to lower expression levels. Commonly, patients relapse or progress with reduced CD22 expression compared to pre-treatment, contrasting with the loss of CD19 expression typically observed after CD19-CAR therapy. Prior work demonstrated that an antigen-independent “tonic” signaling CD22-CAR has enhanced clinical efficacy relative to other CD22-CARs tested. However, tonic signaling has been shown to be detrimental to long-term CAR T cell function. Here, we demonstrate a balance between binding affinity and antigen-independent signaling (determined by length of the linker between fragment variable regions) in determining CAR function against CD22. We show that CAR function against both CD22Lo and WT leukemia can be augmented by boosting binding affinity without shortening the linker to induce tonic signaling, establishing rational combinatorial modification of antigen binding domain as an important approach for modulating the function of cellular therapeutics.

overall_design: Human donor PBMC-derived CD3+ T cells were activated and virally transduced with unique chimeric antigen receptor (CAR) constructs targeting the CD22 antigen, enriched for CAR+ cells, then cryopreserved. Following thaw, cells either stimulated on plate-bound CD22 or left at rest for 24 hours, followed by dead cell depletion and analysis by scRNA-seq.

first: Kole

middle: R

last: DeGolier

first: Catherine

last: Pham-Danis

first: Samuel

middle: D

last: Burciaga

first: Zachary

last: Walsh

first: Christine

last: Brzezinski

first: Amanda

last: Novak

first: Lillie

last: Leach

first: Jennifer

last: Cimons

first: Wei

last: Li

first: Zhu

last: Zhongyu

first: Dimiter

last: Dimitrov

first: James

middle: P

last: Scott-Browne

first: Mark

last: Kohler


title: Sequence and chemical specificity define the functional landscape of intrinsically disordered regions [ONT]

first: Frauke

last: Huth

first: Iris

last: Langstein-Skora

first: Andrea

last: Schmid

first: Mariia

last: Likhodeeva

first: Franziska

last: Kunert

first: Felix

last: Metzner

first: Ryan

last: Emenecker

first: Mary

last: Richardson

first: Wasim

last: Aftab

first: Maximilian

last: Götz

first: Sarah

middle: K

last: Payer

first: Niccoló

last: Pietrantoni

first: Valentina

last: Sjeničić

first: Sakthi

last: Ravichandran

first: Till

last: Bartke

first: Karl-Peter

last: Hopfner

first: Ulrich

last: Gerland

first: Philipp

last: Korber

first: Alex

last: Holehouse


title: Responders to low-dose ATG induce CD4+ T cell exhaustion in type 1 diabetes

city: Seattle,

first: Stephanie

last: Osmond

address: 1201 9th Ave

institute: Benaroya Research Institute

summary: Low-dose anti-thymocyte globulin (ATG) transiently preserves C-peptide and lowers HbA1c in individuals with recent-onset type 1 diabetes (T1D); however, the mechanisms of action and features of the response remain unclear. Here, we characterized the post hoc immunological outcomes of ATG administration and their potential use as biomarkers of metabolic response to therapy (i.e., improved preservation of endogenous insulin production).METHODSWe assessed gene and protein expression, targeted gene methylation, and cytokine concentrations in peripheral blood following treatment with ATG (n = 29), ATG plus granulocyte colony-stimulating factor (ATG/G-CSF, n = 28), or placebo (n = 31).RESULTSTreatment with low-dose ATG preserved regulatory T cells (Tregs), as measured by stable methylation of FOXP3 Treg-specific demethylation region (TSDR) and increased proportions of CD4+FOXP3+ Tregs (P < 0.001) identified by flow cytometry. While treatment effects were consistent across participants, not all maintained C-peptide. Responders exhibited a transient rise in IL-6, IP-10, and TNF-α (P < 0.05 for all) 2 weeks after treatment and a durable CD4+ exhaustion phenotype (increased PD-1+KLRG1+CD57- on CD4+ T cells [P = 0.011] and PD1+CD4+ Temra MFI [P < 0.001] at 12 weeks, following ATG and ATG/G-CSF, respectively). ATG nonresponders displayed higher proportions of senescent T cells (at baseline and after treatment) and increased methylation of EOMES (i.e., less expression of this exhaustion marker).CONCLUSIONAltogether in these exploratory analyses, Th1 inflammation-associated serum and CD4+ exhaustion transcript and cellular phenotyping profiles may be useful for identifying signatures of clinical response to ATG in T1D.TRIAL REGISTRATIONClinicalTrials.gov NCT02215200.FUNDINGThe Leona M. and Harry B. Helmsley Charitable Trust (2019PG-T1D011), the NIH (R01 DK106191 Supplement, K08 DK128628), NIH TrialNet (U01 DK085461), and the NIH NIAID (P01 AI042288).

overall_design: RNA-sequencing was performed on whole blood samples from 88 participants in the trial (31 placebo treated, 29 ATG treated, and 28 ATG-GCSF) treated. A total of 496 samples were profiled. Samples for each donor were collected at up to six visits measured in weeks following the start of therapy (visits 0, 2, 12, 24, 48, and 96).

first: Laura

middle: M

last: Jacobsen

first: Kirsten

last: Diggins

first: Lori

last: Blanchfield

first: James

last: McNichols

first: Daniel

last: Perry

first: Jason

last: Brant

first: Xiaoru

last: Dong

first: Rhonda

last: Bacher

first: Vivian

last: Gersuk

first: Desmond

last: Schatz

first: Mark

last: Atkinson

first: Clayton

middle: E

last: Mathews

first: Michael

last: Haller

first: S

middle: Alice

first: Peter

last: Linsley

first: Todd

last: Brusko


title: Joint single-cell profiling of Cas9 edits and transcriptomes reveals on- and off-target effects on gene expression (rhAmpSeq)

city: La Jolla

first: Graham

last: McVicker

address: 10010 N Torrey Pines Rd

institute: Salk Institute for Biological Studies

summary: A longstanding barrier in genome engineering with CRISPR-Cas9 has been the inability to characterize editing outcomes at single-cell resolution. We present "Superb-seq", a new scRNA-seq method that measures on-target and off-target genome edits and associated transcriptomes. In contrast to Perturb-seq that captures single-cell guide information, Superb-seq directly captures single-cell edit profiles by T7 in situ transcription. We demonstrate and validate this method through two Superb-seq experiments in 30,000 cells of three cell types, amplicon sequencing (rhAmpSeq), and bulk RNA-seq. Superb-seq uses off-the-shelf kits, standard laboratory equipment, and requires no virus, which will improve CRISPR screens in diverse tissues and functional characterization of CRISPR therapeutics.

overall_design: RhAmpseq was performed on two sets of eight genomic DNA samples. The first sample of each set was a control sample of unedited (electroporation only) K562 cells. The subsequent seven test samples were K562 cells edited with CRISPR-Cas9 and T7 promoter, using seven guides targeting chromatin remodeler genes ARID1A, SMARCA4, CHD3, and CHD4, one guide per sample. These eight samples were processed into rhAmpSeq libraries using one of two primer sets: either a multiplex panel of 52 primer pairs targeting Superb-seq detected and in silico predicted edit sites, or a singleplex primer pair targeting ADSS1. Sixteen total libraries, one replicate per sample and primer set.

middle: H

last: Lorenzini

first: Brad

last: Balderson


title: A Glucocorticoid-Fas axis controls immune evasion during metastatic seeding [CUT&Run_4T1-Dex]

city: Boston

first: Judith

last: Agudo

state: MA

address: 1 Jimmy fund way

institute: Dana Farber Cancer Institute

summary: Metastasis is the major cause of death for patients with Triple Negative Breast Cancer (TNBC) and other solid malignancies. Metastases arise from cancer cells that disseminate from the original tumour, survive systemic immune surveillance, and colonize new organs. Little is known about how initial disseminated tumour cells (DTCs) overcome anti-tumour immunity upon seeding a new organ. Here we used a visible antigen in a model of TNBC with cognate CD8+ T cells to interrogate mechanisms of immune evasion in early metastatic seeding. Analysis of surviving DTCs revealed Glucocorticoid Receptor (GR) activation as a key driver of resistance to both CD8+ T cells and Natural Killer (NK) cells. Niche profiling using an optimized labelling tool uncovered Fas-FasL as a key pan-cytotoxic pathway against DTCs, which is repressed by GR activation. Pharmacologic inhibition of GR in combination with immunotherapy reduced metastatic burden and expanded lifespan in mice. Thus, we discovered a novel mechanism of immune evasion that operates specifically in DTCs, illustrating the unique immune-cancer interactions at this stage in the metastatic cascade. Our findings suggest that there are therapeutic opportunities to eliminate DTCs, separately from treatments aimed at primary tumours, with GR inhibition as one promising target.

overall_design: 4T1 breast tumour cells were treated with Dexamethasone or vehicle control for one week in vitro. Tumour cells were collected and ChIP was performed using a Glucocorticoid Receptor (GR) antibody. CUT&Run was performed on the resulting DNA. Details are included under Fig. 4.

first: Monica

last: Cassandras

first: Adam

last: Getzler


title: Joint single-cell profiling of Cas9 edits and transcriptomes reveals on- and off-target effects on gene expression (RNA-seq)

overall_design: RNA-seq was performed on two sets of three human nuclei samples, one set per target site and cell type pair (CTLA4 in Jurkat, B2M in K562). For each set, first was a control sample with mock in situ transcription (no T7 RNA polymerase). Second was another control sample with CRISPR-Cas9 RNP only (no T7 promoter) and in situ transcription. Third was a test sample with both CRISPR-Cas9 RNP and T7 promoter, and in situ transcription. Two technical replicate libraries per sample.


title: A Glucocorticoid-Fas axis controls immune evasion during metastatic seeding [scRNAseq_4T1-tdTomato-cTAT-zsGreen]

overall_design: 4T1-shGR (Glucocorticoid Receptor, GR, knockdown) and 4T1-shControl breast tumour cells were transduced to express tdTomato and secreted lipid-soluble cTAT-zsGreen for niche labelling. Tumour cells were injected into primary tumour-bearing BALB/c mice. Tumour cells and labelled niche cells were FACS-sorted from the lungs of mice at D7 post-injection. Details are included under Fig. 3.


title: RNA sequencing on  liver samples from wild type C57BL/6J mice exposed to the vandetanib

city: Hangzhou

first: Wu

middle: Wen

last: Tong

state: Zhejiang

address: 866 Yuhangtang Road

institute: Zhejiang University

summary: Purpose: The aim of this study was to  demonstrate the hepatotoxicity mechanism induced by vandetanib. Methods: Liver mRNA profiles of C57BL/6J mice exposed to the vandetanib  were generated by RNA sequencing. Results: We analyzed the results of high-throughput transcriptome sequencing on liver samples from C57BL/6J mice exposed to the vandetanib and identified that vandetanib increased apoptosis-related genes expression apoptosis and  the JAK-STAT signaling pathway was significantly enriched.

overall_design: RNA was isolated from mice administered with CMC-Na or 100 mg/kg/day vandetanib using the TRIzol reagent (15596026, Invitrogen). The subsequent RNA sequencing (RNA-seq) was performed by LC-Bio.

first: Peihua

last: Luo

first: Hao

last: Yan

first: Wentong

last: Wu


title: An inflammatory and quiescent HSC subpopulation expands with age in humans

first: Ksenia

last: Safina

address: 4 Blackfan St

institute: Brigham and Women's Hospital

summary: Aging of the blood system impacts systemic health and can be traced to hematopoietic stem cells (HSCs). Despite multiple reports on human HSC aging, a unified map detailing their molecular age-related changes is lacking. We developed a consensus map of gene expression in HSCs by integrating seven single-cell datasets. This map reveals previously unappreciated heterogeneity within the HSC population. It also links inflammatory pathway activation (TNF/NFκB, AP-1) and quiescence within a single gene expression program. This program dominates an inflammatory HSC subpopulation that increases with age, highlighting a potential target for further experimental studies and anti-aging interventions.

overall_design:
Sternum bone marrow samples from young (<40 y.o.) and aged (>60 y.o.) individuals were analysed with scRNA-seq.
Bone marrow cells from nine donors were collected from the iliac crest of patients undergoing cardiac surgery under an excess sample banking and sequencing protocol that covers all study procedures and was approved by the Institutional Review Board (IRB) of Mass General Brigham. Donors were confirmed negative for common CHIP mutations using targeted sequencing. Mononuclear cells were isolated using Ficoll or lymphoprep and cryopreserved in liquid nitrogen storage. Cells were thawed using standard procedures, and viable (DAPI negative) cells were sorted on a Sony SH800 flow cytometer. Next, 10,000-15,000 cells were loaded onto a 10x Genomics chip. Further processing was done using the recommended procedures for the 10x Genomics 3’ v3.0, v3.1, or v4 chemistry. Libraries were sequenced on the NovaSeq SP 100 cycle with the following parameters (Read 1: 28 + Read 2: 75 + Index 1 (i7): 10 + Index 2 (i7): 10). Count matrices were generated using CellRanger v.7.1.0 with default settings and GRCh38 as the reference genome.

first: Jonathan

last: Good

last: van Galen


title: Granzyme B-mimicking nanozyme-containing nanovesicles for tumour-targeting delivery

city: tinajin

first: qiqi

last: liu

state: other

address: nankai university

institute: nankai university

summary: Cytotoxic T lymphocytes play a crucial role in anti-tumour immunity, with granzyme B (GrB) being a decisive factor in this process. Developing a GrB-based delivery system to mimic T cell-based immunotherapy holds promise but remains challenging. Here, we present an artificial metalloenzyme (nanozyme) with GrB-like protease activity capable of inducing caspase-dependent cell apoptosis. The nanozyme is based on the site-specific binding of Pd(II) ions to recombinant human heavy chain apo-ferritin nanocages to generate a binuclear catalytic centre consisting of two Pd atoms with bridging cysteine ligands, monodentate methionine and histidine residues, and two water molecules. We show that encapsulation of the Pd-ferritin complex within membrane-fused lipid nanoparticles comprising surface-displayed single-chain antibodies affords GrB-mimicking nanozyme-based nanovesicles capable of receptor-mediated delivery of the nanozyme into the cytoplasm of tumour cells and induction of caspase-dependent apoptosis. This study provides valuable insights into the construction of nano-delivery systems with artificial GrB activity and presents a promising therapeutic option for solid tumours.

overall_design: Tumour cells were treated with FNP-antiHER2 for 24 h. RNA was extracted and sequenced using the Illumina platform.

first: Xueyan

last: Hu

first: Qiqi

last: Liu

first: Helong

last: Kang

first: Ming

last: Zhao

first: Zheng

last: Zhou

first: Yuanke

last: Li

first: Weicheng

last: Peng

first: Tianyi

last: Qi

first: Ruming

first: Li

last: Jiao

first: Jie

last: Zhuang

first: Yijin

first: Stephen

last: Mann

first: Xinglu

last: Huang


title: Multiomic ATAC + Gene Expression sequencing on E8.75 wild type and Nr6a1 null mouse embryos

city: Kansas City

first: Andrew

middle: Joseph

last: Price

state: MO

address: 1000 E. 50th Street

department: Computational Biology

institute: Stowers Institute for Medical Research

summary: Mouse embryos from 2 different litters were collected at E8.75. 3-4 Nr6a1 null embryos and 3-4 wild type embryos were pooled into one sample per genotype. Nuclei was then isolated using the 10X Genomics Nuclei Isolation Kit. Samples were then processed and analyzed using 10X Genomics Multiomic ATAC + Gene Expression single nuclei profiling.

overall_design: Two samples, each containing 3-4 embryos, were collected from 2 litters of E8.75 mouse embryos according to genotype and analyzed with 10 Multiomic ATAC + Gene Expression single cell profiling.

first: Emma

middle: L

last: Zajic

first: Paul

last: Trainor


title: E. coli CFT073 gene expression at t= 4 hours after a temperature upshift from 23˚C to 37˚C (M9 minimal glycerol medium, exponential phase growth)

city: Northampton

first: Christine

middle: Ann

last: White-Ziegler

address: Clark Science Center

department: Biological Sciences

institute: Smith College

summary: We investigated the effect of a temperature shift from 23˚C to 37˚C, mimicking the temperature transition experienced by a microbe as it enters a human host.  This strategy allowed the exploration of gene expression changes in a short term response to a temperature upshift and to characterize the suite of genes required to respond to 37˚C in the first few hours.  We demonstrate that a significant number of genes are rapidly altered in expression within hours after a temperature shift.   Many of these genes are related to adhesion, immune evasion, and stress responses, suggesting that temperature may serve as a sentinel cue to predict and prepare for niche adaptation in the host.

overall_design:
An aerobic, mid-exponential phase (A600= 0.2-0.6) culture was grown at 23˚C in M9 minimal medium containing glycerol as a carbon source.  Aliquots of the initial culture were shifted to 37˚C to allow harvest of cells at time points after the shift.  RNA-Seq results compare the T=4 hours timepoint after the shift to 37˚C to a culture retained at 23˚C, T=4 hours.
Analysis used 23˚C (T=4 hours) RNA as control for comparison to the experimental samples harvested at T=4 hours after a temperature upshift to 37˚C

first: Daniel

last: Stoebel


title: Regulation of INSM1 Gene Expression and Neuroendocrine Differentiation in High-Risk Neuroblastoma

city: New Orleans

first: Chiachen

last: Chen

state: Louisiana

address: 533 Bolivar Street

department: Genetics

institute: LSUHSC

summary: Neuroblastoma (NB), a pediatric cancer of sympatho-adrenal (SA) lineage, is marked by disrupted differentiation and cellular heterogeneity. INSM1, a zinc-finger transcription factor, is highly expressed in NB and developing SA tissues, where it regulates neuroendocrine differentiation, especially in chromaffin cells. We investigated INSM1’s role in maintaining an undifferentiated, progenitor-like state in NB and its regulation via metabolic and epigenetic mechanisms. Transcriptomic profiling, promoter assays, and metabolic flux analysis revealed that INSM1 expression correlates with methionine cycle activity, particularly the S-adenosylmethionine (SAM)/S-adenosylhomocysteine (SAH) ratio. Disruption of SAM/SAH balance altered INSM1 promoter activity and histone methylation, implicating epigenetic control in NB cell fate. Retinoic acid-induced differentiation downregulated INSM1 and N-Myc, linking INSM1 to tumor cell immaturity. INSM1 overexpression in SH-SY-5Y cells upregulated neuroendocrine and thyroid hormone-related genes (CHGA, CHGB, DDC, NCAM1, DIO3, TH), while suppressing genes involved in cell cycle (MYBL2, RRM, CDC25A), methionine metabolism (AHCY, MAT2A), transcriptional regulation (EZH2), and oncogenic signaling (ALK, LINC011667). These findings suggest that INSM1 promotes NB aggressiveness by sustaining a neuroendocrine progenitor-like phenotype through metabolic-epigenetic coupling.

overall_design: SH-SY-5Y cells  transduced with INSM1 adenoviral vectors for INSM1 overexpression. The effects of INSM1 on neuroblastoma differentiation  and growth were analyzed by RNA-seq

first: Siyuan

last: Cheng

first: Xiuping

last: Yu

first: Michael

middle: S

last: Lan


title: OR7A10 GPCR engineering boosts CAR-NK therapy against solid tumors

city: West Haven

first: Sidi

state: CT

address: 850 West Campus Drive

institute: Yale University

summary: Chimeric antigen receptor (CAR)-natural killer (NK) cell therapies hold promise for solid tumors but remain limited by poor tumor infiltration, persistence, and resistance within the tumor microenvironment (TME). To identify gain-of-function (GOF) targets that enhance CAR-NK efficacy, we performed an unbiased in vivo Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) activation (CRISPRa) screen, followed by a barcoded targeted in vivo open reading frame (ORF) screen in primary human CAR-NK cells. We identified, and robustly validated OR7A10, a G protein-coupled receptor (GPCR), as the top candidate. Engineering CAR-NKs with OR7A10 cDNA, a CRISPR-independent method with simple manufacturing strategy, enhanced proliferation, activation, degranulation, cytokine production, death ligand expression, chemokine receptor expression, cytotoxicity, persistence, metabolic fitness, and TME resistance, while reducing exhaustion in primary human NK cells derived from multiple peripheral blood and cord blood donors. OR7A10-GOF CAR-NKs displayed robust in vivo efficacy across multiple solid tumor models, achieving a 100% complete response in an orthotopic breast cancer model with long term tumor control and survival benefit. These findings establish OR7A10-engineered CAR-NKs as a highly potent and scalable off-the-shelf therapeutic for solid tumors.

overall_design: Bulk RNA-seq profiling of NK cells transduced with different CAR constructs (OR7A10, OR7A10Stop, truncatedHER2CAR_OR7A10, truncatedHER2CAR_OR7A10Stop, HER2CAR_OR7A10, HER2CAR_OR7A10Stop) under 0hr and 24hrs stimulation conditions.

first: Luojia

last: Yang

first: Kaiyuan

last: Tang


title: A Glucocorticoid-Fas axis controls immune evasion during metastatic seeding [RNAseq_4T1-tdTomato]

first: Judith

last: Agudo

overall_design: 4T1-tdTomato breast tumour cells were injected into BALB/c or NSG female mice. Tumour cells were FACS-sorted (DAPI-/CD45-/tdTomato+) from the mammary tumor or lungs of mice at indicated tmepoints. Details are included under Fig. 1a.

first: Monica

last: Cassandras


title: Epigenetic Signatures of Brain Aging by Single Nucleus Methylation Sequencing [DHC-9mo-m-001]

first: Joseph

last: Ecker

state: CA

address: 10010 North Torrey Pines Road

department: Genomic Analysis Laboratory

institute: HHMI-Salk-Institute

summary: Illustrating aging effects on the epigenomic features of brain is critical to understand brain functional decline and neurodegeneration. Single nucleus methylation sequencing was applied to brain regions of mouse at the age of 2, 9 and 18 months. We use these data to identify the epigenomic changes with aging in mouse brains.

overall_design:
Apply snmC-seq3 to mouse brain regions to identify aging effects on epigenomics

Processed data for these samples are available at:

https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE313770

first: Q

last: Zeng

first: Wei

last: Tian

first: Anna

last: Bartlett

first: J

last: Nery

first: Rosa

last: Gomez Castanon

first: Julia

last: Osteen

first: Jordan

last: Altshul

first: N

last: Johnson

first: M

last: Kenworthy

first: C

last: Valadon

first: Huaming

last: Behrens


title: A Glucocorticoid-Fas axis controls immune evasion during metastatic seeding [scRNAseq_4T1-GFPThy11]

overall_design: A mix of 4T1-GFP and 4T1-Thy1.1 breast tumour cells were intravenously injected into BALB/c female mice following adoptive transfer of activated Jedi anti-GFP CD8+ T cells. Tumour cells were FACS-sorted (DAPI-/CD45-/GFP+ or Thy1.1+) from the lungs of mice at D7 post-injection. Details are included under Fig. 1g.


title: A Glucocorticoid-Fas axis controls immune evasion during metastatic seeding [RNAseq_4T1-GFP]

overall_design: 4T1-GFP breast tumour cells were intravenously injected into Jedi or NSG female mice. Tumour cells were FACS-sorted (DAPI-/GFP+) from the lungs of mice at D7 post-injection. Details are included under Fig. 1e.


first: Sidi

last: Chen

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

overall_design: Refer to individual Series


title: OR7A10 GPCR engineering boosts CAR-NK therapy against solid tumors [Primary Screen]

overall_design: α-HER2-CAR-NK92 cell were transduced with dCas9-VP64-Blast and MS2-P65-HSF1-Hygro (Addgene)  at MOI < 1 and selected with Hygromycin and Blastcidin for 5-7 days. After complete selection, 1.8e8 CAR-NK92 cells were transduced with the sgRNA library (Addgene) at MOI = 0.2 (0.2 infectivity * 1.8e8 cells / 70,290 sgRNAs = 512x coverage). After 7 days of selection with Zeocin, > 4e7 Library-infected CAR-NK92 cells were maintained in culture (4e7 cells / 70,290 sgRNAs = 569x, although the coverage was limited by the bottleneck of the smaller in the process, e.g. 512x coverage). Eight-to-twelve-week-old female NSG mice were inoculated with 1e6 HT29-GL cells through subcutaneous injection. On day 8, 5e6 library-infected CAR-NK92 cells were adoptively transferred into each tumor-bearing mouse via intravenous injection (5e6 / 70,290 sgRNAs = 71x coverage). To support CAR-NK92 cell activity, mice received daily intraperitoneal injections of 2.5 μg human IL-2 beginning on the day of NK cell transfer. On 13-day post-transfer, mice were euthanized, and tumors were harvested for the subsequent experiments.

first: Paul

last: Renauer

first: Luojia

last: Yang


title: OR7A10 GPCR engineering boosts CAR-NK therapy against solid tumors [mini screen]

overall_design: This experimental data is a primary human CAR-NK barcoded ORF-UMI mini-screen. An ORF library was generated for the top 35 gene targets (cDNA length ≤ 1 kb) from the initial CRISPRa screen, along with five fluorescent reporter genes as null controls. The ORF library was synthesized as individual eBlocks (IDT), pooled at equimolar ratios, and amplified with primers containing four-nucleotide random barcodes (UMIs; NNNN) to introduce molecular redundancy targeting the same gene. The ORF expression vector plasmid was bought from Addgene (#145026). The barcoded ORF library was pool-cloned into the expression vector using Gibson assembly.  Primary human peripheral blood NK cells from two healthy donors were first transduced with HER2-CAR lentivirus and then with the ORF library lentivirus at an MOI of ~0.2. A total of 20 million library-transduced CAR-NK cells were injected into each HT29 tumor-bearing mouse (n = 5 mice for Donor 0957 and n = 3 for Donor 0958 for each time point). Genomic DNA was extracted from tumors at day 3 and 6 post-treatment, as well as from pre-injection cells, and subjected to NGS to quantify ORF and UMI barcode representations.  Note that each set of fastq files is demultiplexed by the reverse

first: Kaiyuan

last: Tang


title: Epigenetic Signatures of Brain Aging by Single Nucleus Methylation Sequencing [CP-9mo-m-023]

summary: Illustrating aging effects on the epigenomic features of brain is critical to understand brain functional decline and neurodegeneration. Single nucleus methylation sequencing was applied to brain regions of both male and female mouse at the age of 2, 9 and 18 months. We use these data to identify the epigenomic changes with aging in mouse brains.


title: OR7A10 GPCR engineering boosts CAR-NK therapy against solid tumors [scRNA-Seq]

overall_design: Single-cell transcriptomics was performed by 10X Genomics sequencing on OR7A10-OE and OR7A10(STOP) control CAR-PBNK-hIL-15 cells that were extracted from in vivo xenograft models of HT29-GL in NSG mice after 3 days.


title: Gut microbiota promote immune tolerance at the maternal-fetal interface

city: New York

last: Brown

state: New York

address: 413 E 69th Street, 1252A

institute: Weill Cornell Medicine

summary: Immune tolerance at the maternal-fetal interface is required for fetal development. Excessive maternal interferon gamma (IFNγ) and interleukin-17 (IL-17) is linked to pregnancy complications, but the regulation of maternal IFNγ and IL-17 at the maternal-fetal interface (MFI) is poorly understood. Here we demonstrate a gut-placenta immune axis in pregnant mice in which the absence or perturbation of gut microbiota dysregulates maternal IFNγ and IL-17 responses at the MFI, resulting in fetal resorption. Microbiota-dependent tryptophan derivatives suppress IFNγ+ and IL-17+ T cells at the MFI by priming myeloid-derived suppressor cells (MDSCs) and gut-derived RORγt+ Tregs, respectively. The tryptophan derivative indole-3-carbinol, or tryptophan-metabolizing Lactobacillus murinus, rebalances the T cell response at the MFI and reduces fetal resorption in germ-free mice. Furthermore, MDSCs, RORγt+ Tregs, and microbiota-dependent tryptophan derivatives are dysregulated at the MFI in human recurrent miscarriage cases. Together, our findings identify microbiota-dependent immune tolerance mechanisms that promote fetal development.

overall_design: scRNA-seq of CD45+ cells from blood, placenta, and uterus of E16.5 pregnant SPF and germ-free mice

middle: A

first: Mohammed

last: Amir

first: Shui

last: Yu

first: Daniel

last: Wong

first: Jinghua

last: Gu

first: Uthra

last: Balaji

first: Christopher

last: Parkhurst

first: Seunghee

last: Hong

first: Lucy

last: Hart

first: Hannah

middle: C

last: Carrow

first: Mamadou

last: Bah

first: Aparna

last: Ananthanarayanan

first: Katherine

middle: Z

last: Sanidad

first: Mengze

last: Lyu

first: Anisa

last: Siddikova

first: Marina

last: Lima Silva Santos

first: Inna

last: Serganova

first: Gretchen

last: Diehl

first: Josef

last: Anrather

first: Naohiro

last: Inohara

first: Gregory

last: Sonnenberg

first: Virginia

last: Pascual

first: Melody

middle: Y


title: Urinary Clusterin as a Biomarker of Human Kidney Disease Progression and Response to the Endothelin Receptor Antagonist Atrasentan: An Exploratory Analysis from the SONAR Trail

city: Ann Arbor

first: Jennifer

last: Harder

state: Michigan

address: 1150 W. Medical Center Drive

department: Division of Nephrology

institute: The University of Michigan

summary: The endothelin receptor antagonist atrasentan improved kidney outcomes in the SONAR trial for type 2 diabetes and chronic kidney disease (NCT01858532), though individual responses varied. To identify molecular biomarkers of atrasentan response and outcome, we conducted a nested case-control proteomics study (N=180) within the SONAR trial population and identified urinary clusterin (uCLU) as the top candidate. Transcriptomic analyses of human kidney biopsies at tissue and single cell level from independent cohorts revealed higher CLU mRNA levels associated with worse kidney function and outcomes. An endothelin signaling activation score derived from pathway genes was reduced by atrasentan in mice with diabetic kidney disease. In the SONAR trial (N=3,060), higher uCLU predicted worse outcomes, while atrasentan reduced uCLU by 42.6% over six weeks. Early uCLU changes independently predict improved kidney outcomes. In summary, uCLU is associated with kidney disease progression and response to atrasentan treatment, supporting its potential as a pharmacodynamic biomarker to target therapy.

overall_design: Bulk RNAseq data obtained from kidney glomerular tissues from wild type (n=5), BTBR ob/ob mice with 6 wk of saline (n=5) or atrasentan (n=7) treatment, respectively

first: Wenjun

last: Ju

first: Viji

last: Nair

first: Priya

last: Vart

first: J

last: Smeijer

first: Kelly

last: Hudkins

first: Maria

last: Larkina

first: Paul

last: Perco

first: Fred

last: Burdet

first: Kerby

last: Shedden

first: Michael

last: Hwang

first: Edmond

last: Lee

first: Christopher

last: O’Connor

first: John

last: Hartman

first: Markus

last: Bitzer

first: Mark

last: Ibberson

first: Kevin

last: Duffin

last: Gomez

first: Charles

last: Alpers

first: Matthias

last: Kretzler

first: Hiddo

last: Heerspink


title: LINE-1 locus transcription nucleates oncogenic chromatin architecture [RNA-Seq]

city: Memphis

first: Jian

last: Xu

state: Tennessee

address: 262 Danny Thomas Place, MS 345

institute: St. Jude Children's Research Hospital

summary: Retrotransposons are a prolific class of genomic parasites frequently upregulated in human cancers, where their spontaneous mobility can generate insertional mutations and structural variations. However, whether retrotransposon-derived gene products contribute to cancer beyond retrotransposition remains unclear. Here, we identify an oncogenic, chromatin-associated function of RNAs produced by Long Interspersed Element-1 (LINE-1), the only autonomous human retrotransposon. Sequencing analysis of subcellular fractions revealed that LINE-1 RNAs primarily exist as nascent transcripts expressed from full-length, cell type-specific genomic copies of evolutionarily young LINE-1 subfamilies. Through developing a long-read sequencing-based chromosome conformation assay, we uncovered a class of highly interactive LINE-1 loci essential for gene expression across cancer subtypes, establishing an unexpected regulatory link between LINE-1 locus transcription and oncogenic gene expression via long-range chromatin interactions. Depleting LINE-1 RNAs disrupted LINE-1-centric chromatin interactions, leading to the downregulation of associated genes, whereas inserting a LINE-1 element into an endogenous genomic locus induced de novo chromatin interactions in a transcription-dependent manner. Therefore, retrotransposons not only mutagenize linear genetic content but also regulate cancer gene expression through transcription-dependent nucleation of chromatin architecture.

overall_design: Chromatin RNA-sequencing (Chromatin RNA-seq) was performed on ten cancer cell lines, as well as on ER-positive and triple-negative breast cancer (TNBC) tissue samples. RNA-seq was conducted on MCF7 cells treated with either LINE-1-targeting ASO1, ASO2, or a scrambled ASO control. Both Chromatin RNA-seq and Cytosolic RNA-seq were performed on three cell lines. Additionally, mRNA-sequencing was carried out on MCF7 cells stably expressing Tet-inducible Cas9 and sgRNAs targeting either L1HS_175, L1HS_274, or the AAVS1 safe harbor locus.

last: Lee Jr.

first: Yuannyu

last: Zhang


title: LINE-1 locus transcription nucleates oncogenic chromatin architecture [ChIRP-seq]

overall_design: Chromatin Isolation by RNA Purification (ChIRP)-sequencing was peformed in MCF7 cells with biotinylated probes targeting the LINE-1 5’UTR, ORF1 or ORF2.


title: RNA-seq of ESCC Cells  Overexpressed TFAP2β or its LLPS-deficient Mutants

city: Chengdu

first: Lu

last: Pu

state: Sichuan

address: #37 Guoxue Alley, Wuhou District

institute: west china hospital

summary: This dataset contains raw RNA-seq data from esophageal squamous cell carcinoma (ESCC) cells overexpressing GFP-TFAP2β, its LLPS-deficient mutants (ΔIDR3 and Mut_IDR3) or GFP control. The experiment aims to explore the downstream genes of TFAP2β and its phase separation-deficient mutants to uncover the role of TFAP2β's LLPS activity in regulating gene expression in ESCC. Data were collected from the genomic binding of TFAP2β and its LLPS-deficient mutants, providing insights into the regulatory networks controlled by TFAP2β in cancer progression.

overall_design: RNA-seq of TFAP2β, its LLPS-deficient mutants (ΔIDR3 and Mut_IDR3) or GFP control in ESCC cells KYSE150 to understand the downstream genes and regulatory activity associated with TFAP2β’s phase separation mechanism.