title: Effect of Periplocymarin on Gene Expression in Gastric Cancer Cell Lines HGC27 and MKN45

city: Shenyang

first: Angting

last: Ke

state: Liaoning

address: No. 4, Chongshan East Road, Huanggu District

department: Department of Surgical Oncology

institute: Fourth Affiliated Hospital of China Medical University

summary: Periplocymarin (PPM) is an extract isolated from the traditional plant Periploca sepium with a cardiac glycoside-like structure. Previous studies have gradually revealed the anticancer potential of PPM. PPM exhibits a stronger inhibitory effect on tumor cell viability compared to ouabain, and it can both induce and sensitize TRAIL-induced apoptosis in cancer cells, while showing relatively low toxicity to PBMCs. The anticancer effects of PPM are influenced by several cellular processes it induces, such as autophagy and metabolism, although the exact mechanisms remain unclear. We have observed that PPM inhibits cell viability, proliferation, and the cell cycle in gastric cancer cells, but the process of inducing apoptosis in gastric cancer cells is moderate and slow. Therefore, there is an urgent need to elucidate the changes in the gene expression profile of tumor cells following PPM treatment, to better understand how PPM affects cellular biological processes and signaling pathways, thus maximizing its anticancer potential.

overall_design: RNA-Seq Profiling of HGC27 and MKN45 Cells After 0.3 μM PPM Treatment For 24h

first: Dongqiu

last: Dai


title: Chemotranscriptomic Profiling with a Thiamine Monophosphate Photoaffinity Probe

status: Public on Feb 17 2025

city: Nijmegen

first: Daphne

middle: Ai Lin

last: van den Homberg

address: Heyendaalseweg

department: Physical Organic Chemistry

country: Netherlands

institute: Radboud University

type: Expression profiling by high throughput sequencing

summary: RNA is a multifaceted biomolecule with numerous biological functions and can interact with small molecule metabolites as exemplified by riboswitches. Here, we profile the Escherichia Coli transcriptome on interactions with the metabolite Thiamine Monophosphate (TMP). We designed and synthesized a photoaffinity probe based on the scaffold of TMP and applied it to chemotranscriptomic profiling. Using next-generation RNA sequencing, several potential interactions between bacterial transcripts and the probe were identified. A remarkable interaction between the TMP probe and the well-characterized ribB riboswitch was validated by RT-qPCR, and further verified with competition assays. Localization of the photocrosslinked nucleotides using reverse transcription and docking predictions of the probe suggested binding to the riboswitch aptamer. After examining binding of unmodified TMP to the riboswitch using SHAPE, we found selective yet moderate binding interactions, potentially mediated by the phosphate group of TMP. Lastly, TMP appeared to enhance gene expression of a reporter gene that is under riboswitch control, while the natural ligand Flavin Mononucleotide (FMN) displayed an inhibitory effect, hinting at a potential biological role of TMP. This work showcases the possibility of chemotranscriptomic profiling to identify new RNA-small molecule interactions.

overall_design: Transcriptome-wide profiling of interactions of the TMP photoaffinity probe with E. Coli RNA extracts using next-generation RNA sequencing

first: Stefan

last: Crielaard

first: Casper

last: Peters

first: Alexander

last: Slivkov

first: Willem

middle: A

last: Velema


title: Mesenchymal stem cell with enhanced antioxidant capacity integrates into smooth muscle cells in diabetic detrusor underactivity

city: Seoul

first: DONG-MYUNG

last: SHIN

address: Pungnap-2 dong, Songpa-gu

department: Department of Biomedical Sciences

country: South Korea

institute: University of Ulsan College of Medicine

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

sample_organism:
Homo sapiens
Rattus norvegicus

platform_organism:
Homo sapiens
Rattus norvegicus

overall_design: Refer to individual Series


title: Transcriptional and epigenetic rewiring by the NUP98::KDM5A fusion oncoprotein directly activates CDK12 (ATAC-seq)

city: Vienna

first: Thomas

last: Eder

address: Veterinaerplatz 1

department: Department for Biological Sciences and Pathobiology

country: Austria

institute: University of Veterinary Medicine, Vienna

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Nucleoporin 98 (NUP98) fusion oncoproteins are strong drivers of pediatric acute myeloid leukemia (AML) with poor prognosis. Here we show that NUP98 fusion-expressing AML harbors an epigenetic signature that is characterized by increased accessibility of hematopoietic stem cell genes and enrichment of activating histone marks. We employ an AML model for ligand-induced degradation of the NUP98::KDM5A fusion oncoprotein to identify epigenetic programs and transcriptional targets that are directly regulated by NUP98::KDM5A through CUT&Tag and nascent RNA-seq. Orthogonal genome-wide CRISPR/Cas9 screening identifies 12 direct NUP98::KDM5A target genes, which are essential for AML cell growth. Among these, we validate cyclin-dependent kinase 12 (CDK12) as a druggable vulnerability in NUP98::KDM5A-expressing AML. In line with its role in the transcription of DNA damage repair genes, small-molecule-mediated CDK12 inactivation causes increased DNA damage, leading to AML cell death. Altogether, we show that NUP98::KDM5A directly regulates a core set of essential target genes and reveal CDK12 as an actionable vulnerability in AML with oncogenic NUP98 fusions.

overall_design: ATAC-seq:

first: Selina

last: Troester

first: Ben

last: Haladik

first: Raúl

middle: J

last: Heredia

first: Kaan

last: Boztug

first: Florian

last: Grebien


title: Transcriptional and epigenetic rewiring by the NUP98::KDM5A fusion oncoprotein directly activates CDK12 (RNA-Seq)

overall_design: RNA-seq: a dTAG-NUP98::KDM5A AML cell line was treated with dTAG13 for 8h and 24h, or with BSJ-4-116 for 4h to investigate transcriptional changes upon fusion protein degrdation or CDK12 degradation.

first: Nadja

last: Wukowits


title: Transcriptional and epigenetic rewiring by the NUP98::KDM5A fusion oncoprotein directly activates CDK12 (Cut&Tag)

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Cut&Tag: H3K27ac and H3K4me3 CUT&Tag was perfromed in primary material from a NUP98::KDM5A patient-derived xenograft model and in a dTAG-NUP98::KDM5A AML cell line that was treated with dTAG13 for 8h to investigate changes in histone mark distribution upon fusion protein degradation.


title: Transcriptional and epigenetic rewiring by the NUP98::KDM5A fusion oncoprotein directly activates CDK12 (nascent RNA-Seq)

overall_design: Nascent RNA-seq: a dTAG-NUP98::KDM5A AML cell line was treated with dTAG13 for 1h, followed by 4-thiouridine treatment for 1h to investigate changes in nascent mRNA expression upon fusion protein degradation via SLAM-seq.

first: Johannes

last: Schmoellerl

last: Zuber


title: Transcriptional and epigenetic rewiring by the NUP98::KDM5A fusion oncoprotein directly activates CDK12 (CRISPR screen)

overall_design: Genome-wide CRISPR screen: a single cell-derived Cas9 clone of a NUP98::KDM5A-expressing AML cell line was transduced with a genome-wide sgRNA library and a loss-of-function screen was performed over 14 cell population doublings to investigate functional genetic dependencies.


title: DNMT3A regulates megakaryocyte-biased hematopoietic stem cell fate decisions [EMseq]

city: Houston

first: Margaret

last: Goodell

state: TX

address: One Baylor Plaza, N1030

country: USA

institute: Baylor College of Medicine

type: Methylation profiling by high throughput sequencing

summary: Hematopoietic stem cells (HSCs) are defined by their capacity to regenerate all main components of the peripheral blood, but individual HSCs exhibit a range of preferences for generating downstream cell types. Their propensities are thought to be epigenetically encoded, but no differential regulatory mechanisms have been identified. Here, we considered whether DNA methyltransferase 3A (DNMT3A) has a role in determining the differentiation choice of HSCs. We find that the most primitive megakaryocyte-biased HSCs depend most on DNMT3A for efficient hematopoietic regeneration, particularly of the lymphoid lineages. Reduced DNMT3A amplifies the biased HSC behavior. DNMT3A also regulates megakaryocyte and platelet output in part through DNA methylation at key megakaryocyte lineage loci. Together, our findings establish the role of epigenetic regulation in the fate of megakaryocyte-biased HSCs and their downstream progeny and suggest that the outcomes of DNMT3A-mutant clonal hematopoiesis and malignancies might vary depending on the identity of the HSC that acquires the mutation.

overall_design: To compare the genome-wide methylation patterns in WT and Dnmt3a+/– megakaryocyte-erythroid progenitors and megakaryocytes: Megakaryocyte-erythroid progenitors were sorted by FACS from mouse bone marrow using the markers Lineage–CD117+Sca1–CD16/32loCD34lo. Megakaryocytes were cultured ex vivo from mouse bone marrow.

first: Sarah

last: Waldvogel

first: Virginia

last: Camacho

first: Dandan

last: Fan

first: Anna

middle: G

last: Guzman

first: Alejandra

last: Garcia-Martell

first: Elmira

last: Khabusheva

first: Josephine

last: De La Fuente

first: Rachel

last: Rau

first: Ashlyn

last: Laidman

first: Maria

middle: N

last: Barrachina

first: Estelle

last: Carminita

first: Justin

last: Courson

first: Michael

middle: R

last: Williamson

first: Joanne

middle: I

last: Hsu

first: Chun-Wei

last: Chen

first: Jaime

last: Reyes

first: Subhashree

last: Pradhan

first: Rolando

middle: E

last: Rumbaut

first: Alan

last: Burns

first: Jianzhong

last: Su

first: Benjamin

last: Deneen

first: Kellie

last: Machlus


title: A pontine center in descending pain control

city: Bethesda

first: Tianming

last: Li

state: MD

address: 35A Convent Drive

department: NIDCR

institute: National Institutes of Health

summary: Our perception of pain changes according to expectation, context and mood illustrating how top-down-circuits affect sensory processing. Here we developed an intersectional platform for identifying supraspinal descending neurons that are engaged when an animal experiences pain. Amongst these, we identified a cluster of cells in the pontine micturition center (Barrington’s nucleus), expressing corticotropin-releasing-hormone (BarCrh) that detect painful but not innocuous stimuli. When activated, BarCrh-neurons attenuate nocifensive responses as well as tactile neuropathic pain. Mechanistically, pain related input from the ventrolateral periaqueductal gray activates BarCrh-neurons, which in turn project to the spinal dorsal horn to mediate analgesia. In combination, our data demonstrate that Barrington’s nucleus is not just a relay station dedicated to triggering micturition but also powerfully controls painful sensory input to the brain.

overall_design: AAV-retro-H2B-GFP was injected into the lumbar spinal cord of the TRAP2 mice strain. Upon pain activation, hindbrain descending neurons activated by different types of pain were captured by FACS and subject for 10x single nucleus RNA sequencing.

first: Liu

last: Yuanyuan

first: Hayashi

last: Lauren

first: Huang

last: Wenxi


title: Perturbing local steroidogenesis to improve breast cancer immunity

city: Cambridge

first: Qiuchen

last: Zhao

state: UK

address: Downing College, Regent St, Room45

department: Department of Pathology, CRUK Cambridge Centre

country: United Kingdom

institute: University of Cambridge

summary: Breast cancer, particularly triple-negative breast cancer (TNBC), evades the body's immune defences, in part by fostering an immunosuppressive tumour microenvironment. We present evidence that suppressing local steroidogenesis can enhance anti-tumour immunity against TNBC. Through targeted metabolomics of steroids alongside immunohistochemistry techniques, we initially profiled the presence of various steroids in TNBC patient tumours and identified steroidogenic activity in regions with immune infiltration. In mice, genetic inhibition of immune cell steroidogenesis limited TNBC tumour progression, significantly reducing immunosuppressive components such as tumour-associated macrophages, as revealed by single-cell RNA sequencing. Crucially, inhibiting steroidogenesis seems to strengthen anti-tumour immune responses in dendritic cells and T cells by obstructing glucocorticoid signalling, according to RNA sequencing insights. By undertaking metabolic modelling of the single-cell transcriptomics and targeted tumour steroidomics, we identified mast cells and basophils as the primary steroidogenic agents in TNBC, offering pathways for enhanced therapeutic precision.

overall_design: The immunosuppressive tumour microenvironment (TME) of triple-negative breast cancer (TNBC) undermines the efficacy of immunotherapies. The role of steroids in this immunosuppression remains incompletely understood. In this study, we employed single-cell RNA sequencing (scRNA-seq) of CD45+ immune cells from TNBC tumours in Cyp11a1^fl/fl Vav1Cre and control mice, investigating how genetic inhibition of immune cell steroidogenesis impedes tumour progression by notably reducing immunosuppressive elements. Further, RNA sequencing of glucocorticoid-treated and untreated human dendritic cells (DCs) revealed that glucocorticoids inhibit immune-activating factors in DCs, thereby weakening their capacity to stimulate T cells. By undertaking metabolic modelling of the single-cell transcriptomics from CD45+mCherry+/- immune cells in TNBC tumours from Cyp11a1-mCherry reporter mice, we identified mast cells as the primary steroidogenic contributors in TNBC, offering avenues for increased therapeutic precision.

first: Jhuma

last: Pramanik

first: Bidesh

last: Mahata


title: Transcriptional Heterogeneity and Differential Response of Rod Photoreceptor Pathway Uncovered by Single Cell RNA Sequencing of the Aging Mouse Retina

status: Public on Feb 16 2025

first: Matthew

last: Brooks

address: 6 Center Dr Bldg 6, Rm 303

department: NEI

institute: NIH

summary: Gene expression of single cells from mouse superior and inferior retina during aging.

overall_design: Mouse retinas from 3, 8, 12, and 24 months of age were used to generate scRNA profiles. For each age a 1mm punch from both inferior ans superior dissociate populations were used as input into 10x Genomics Single Cell 3' v2.0.

first: Laura

last: Campello

first: Benajmin

last: Fadl

first: Soumitra

last: Pal

first: Anand

last: Swaroop


title: Patterning effects of FGF17 and cAMP on generation of dopaminergic progenitors for cell replacement therapy in Parkinson’s disease. [scRNA-Seq]

city: Copenhagen

first: Erno

last: Hänninen

address: Blegdamsvej 3B

department: reNEW, Novo Nordisk Foundation Center for Stem Cell Medicine

country: Denmark

institute: University of Copenhagen

summary: Cell replacement therapies using human pluripotent stem cell-derived dopaminergic (DA) progenitors are currently in clinical trials for treatment of Parkinson’s disease (PD). Recapitulating developmental patterning cues, such as fibroblast growth factor 8 (FGF8), secreted at the midbrain-hindbrain boundary (MHB), is critical for the in-vitro production of authentic midbrain DA progenitors. Here, we explore the application of alternative MHB-secreted FGF-family members, FGF17 and FGF18, for DA progenitor patterning. We show that while FGF17 and FGF18 both recapitulate DA progenitor patterning events, FGF17 induced expression of key DA progenitor markers at higher levels than FGF8 and transplanted FGF17-patterned progenitors fully reversed motor deficits in a rat PD model. Early activation of the cAMP pathway mimicked FGF17-induced patterning, although strong cAMP activation also came at the expense of EN1 expression. In summary, we identified FGF17 as a promising candidate for more precise and robust DA progenitor patterning, with potential to improve cell products for treatment of PD.

overall_design: To compare the ability of different MHB-associated FGFs from the FGF8 subfamily to fine-tune hPSC-derived ventral midbrain (VM) DA progenitors, we added either FGF8 or FGF17 from day 9 to 16 for the production of DA progenitors. On day 9 the cells were treated either with FGF8 or FGF17 for 15 min, 1 h, 4 h, and 24 h (n=3). We performed bulk RNA sequencing for these treatments to investigate immediate differences in FGF response pathways. On differentiation day 16 we performed scRNA-seq on hashtag-labeled (hto) DA progenitors patterned with either FGF8 or FGF17 (n=4).

first: Agnete

last: Kirkeby


title: DNMT3A regulates megakaryocyte-biased hematopoietic stem cell fate decisions [RNASeq]

first: Margaret

last: Goodell

overall_design: To compare the transcriptomic profiles of platelets in this study: Whole blood was collected from 4 Dnmt3a+/– mice and 4 WT littermates via cardiac puncture, and platelets were pelleted from platelet-rich plasma by differential centrifugation. Anti-Ter119 and Anti-CD45 bead depletion was performed to removed contaminating leukocytes and RBCs.

first: Sarah

last: Waldvogel

first: Virginia

last: Camacho

first: Dandan

last: Fan

first: Anna

last: Guzman

first: Alejandra

last: Garcia-Martell

first: Elmira

last: Khabusheva

first: Josephine

last: De La Fuente

first: Rachel

last: Rau

first: Ashlyn

last: Laidman

first: Maria

last: Barrachina

first: Estelle

last: Carminita

first: Justin

last: Courson

first: Michael

last: Williamson

first: Joanne

last: Hsu

first: Chun-Wei

last: Chen

first: Jaime

last: Reyes

first: Subhashree

last: Pradhan

first: Rolando

last: Rumbaut

first: Alan

last: Burns

first: Jianzhong

last: Su


title: The potential therapeutic application of ds-cRNA delivery to neurons on microglia  5xFAD mouse

city: Shanghai

first: Li

last: Yang

address: 131 Dong-An Road

department: Institutes of Biological Sciences

country: China

institute: Fudan University

summary: To investigate the effect of circPOLR2A(9,10) delivery to neurons on microglia gene expression in 5xFAD mice

overall_design: Ribosomal RNA (rRNA) depleted (ribo-) RNA-seq for microglia: WT mice treated with AAV PHP.eB-EGFP (n = 3), 5xFAD mice treated with AAV PHP.eB-EGFP (n = 2), 5xFAD mice treated with AAV PHP.eB-circPOLR2A(9,10) (n = 4) and 5xFAD; PKR KO mice (n=4) .

first: Ling-Ling

first: Bo-Wen

last: Jiang

first: Xin

last: Feng

first: Si-Nan

last: Zhai


title: Increased burden of rare risk variants across gene expression networks predisposes to sporadic Parkinson’s disease

city: Piscataway

first: Eleanna

last: Kara

state: New Jersey

address: 683 Hoes Lane West

institute: Rutgers University

summary: Alpha-synuclein (αSyn) is an intrinsically disordered protein that accumulates in the brains of patients with Parkinson’s disease and forms inclusions in neurons called Lewy Bodies. While the mechanism underlying the dysregulation of αSyn in Parkinson’s disease is unclear, it is thought that prionoid cell-to-cell propagation of αSyn has an important role. Through a high throughput screen, we recently identified 38 genes whose knock down modulates αSyn propagation. Follow up experiments were undertaken for two of those genes, TAX1BP1 and ADAMTS19, to study the mechanism with which they regulate αSyn homeostasis. We used a recently developed M17D neuroblastoma cell line expressing triple mutant (E35K+E46K+E61K) “3k” αSyn under doxycycline induction. 3k αSyn spontaneously forms inclusions that show ultrastructural similarities to Lewy Bodies. Experiments using that cell line showed that TAX1BP1 and ADAMTS19 regulate how αSyn interacts with lipids and phase separates into inclusions, respectively, adding to the growing body of evidence implicating those processes in Parkinson’s disease. Through RNA sequencing, we identified several genes that are differentially expressed after knock-down of TAX1BP1 or ADAMTS19. Burden analysis revealed that those differentially expressed genes (DEGs) carry an increased frequency of rare risk variants in Parkinson’s disease patients versus healthy controls, an effect that was independently replicated across two separate cohorts (GP2 and AMP-PD). Weighted gene co-expression network analysis (WGCNA) showed that the DEGs cluster within modules in regions of the brain that develop high degrees of αSyn pathology (basal ganglia, cortex). We propose a novel model for the genetic architecture of sporadic Parkinson’s disease: increased burden of risk variants across genetic networks dysregulates pathways underlying αSyn homeostasis, thereby leading to pathology and neurodegeneration.

overall_design: We determined which genes are differentially expressed after knocking down two genes of interest, TAX1BP1 or ADAMTS19, in comparison to scrambled (non-targeting) control. Three biological (independent) replicates were completed for this experiment.

first: Kara

last: Eleanna


title: Mesenchymal stem cell with enhanced antioxidant capacity integrates into smooth muscle cells in diabetic detrusor underactivity (RNA-Seq)

first: DONG-MYUNG

last: SHIN

summary: Diabetic cystopathy, affecting half of individuals with diabetes, often progresses to detrusor underactivity (DUA), a late and irreversible stage of bladder dysfunction. Despite its prevalence, the etiology of diabetic DUA remains elusive, with no established treatments. Mesenchymal stem cell (MSC) transplantation, demonstrated as a promising therapeutic avenue in preclinical studies, lacks comprehensive analyses of its mechanisms, tumorigenic risks, and optimal protocols for diabetic DUA. This study addresses these gaps, exploring transcriptome changes post-MSC treatment in a diabetic DUA preclinical model. Oxidative injury emerged as a significant pathological mechanism, with N-acetylcysteine (NAC) enhancing MSC synergistic effects, potentially reducing the amount of required MSCs . Additionally, human umbilical cord-derived MSCs with high antioxidant capacity (PFO-MSCs) exhibited improved and sustained therapeutic efficacy, integrating into myocytes within bladder muscle bundles or neighboring pericytes. Single-cell transcriptome analysis of engrafted cells revealed expression changes in pathways related to muscle development and immune responses, emphasizing the pivotal roles of cMET and PD-L1 in muscle regeneration and immune modulation within diabetic DUA. This study provides a comprehensive overview of the pathogenesis and repair processes following MSC therapy for diabetic DUA, enhancing our understanding of its mode of action, efficacy, and safety for future clinical applications.

overall_design: Female Sprague-Dawley rats (8 weeks old) were used in this study. streptozotocin (STZ) was used to induce type I diabetes. After fasting overnight, rats were injected intraperitoneally with STZ (50 mg/kg) dissolved in 0.1 M citrate acid buffer (pH 4.5) solution. For the non-diabetic control group, rats were injected with an equivalent volume of citrate buffer solution. After 4 weeks, gene expression analysis

first: Dong-Myung

last: Shin

first: Chae-Min

last: Ryu

first: YongHwan

last: Kim


title: 5-Aminolevulinic acid suppresses the effector function of feline lymphocytes by reducing the mitochondrial membrane potential

city: Ube

first: Yoichi

last: Mizukami

country: Japan

institute: Yamaguchi　University

summary: 5-Aminolevulinic acid (ALA) is an endogenous amino acid in mammalian cells; it is the first amino acid in the heme biosynthesis pathway occurring in the mitochondria. ALA possesses anti-inflammatory properties by inducing heme oxygenase (HO)-1 expression and releasing heme metabolites in humans and mice. We previously showed that ALA enhances the production of interferon-gamma (IFN-γ) and interleukin-17A in concanavalin A (ConA)-stimulated canine lymphocytes. However, the effects of ALA on feline lymphocytes remain to be investigated. In this study, we demonstrated that ALA-induced HO-1 expression and decreased IFN-γ production in ConA-stimulated feline lymphocytes. Comprehensive RNA sequencing analysis revealed that the Activating transcription factor 4 (ATF4) signaling pathway was inhibited by the addition of ALA. Moreover, we confirmed that ALA decreased ATF4 protein expression. Furthermore, the addition of ALA inhibited the phosphorylation levels of AKT and induced mitochondrial dysfunction in activated lymphocytes. Thus, ALA may suppress the effector function of feline lymphocytes via mitochondrial metabolism, thereby representing a novel mechanism in ALA research.

overall_design: RNA was extracted from feline peripheral blood lymphocytes (PBLs) treated as follows: control, ALA/SFC alone, ConA alone, and a combination of ALA/SFC and ConA for 24 h of incubation, followed by RNA sequencing.

first: Shoma

last: Nishibori

first: Aika

last: Chijiwa

first: Satoshi

last: Kambayashi

first: Nao

last: Iwatani

first: Aki

last: Sakai

first: Keishiro

last: Isayama

first: Nobuyuki

last: Shiranaga

first: Takuya

last: Mizuno

first: Masaya

last: Igase


title: Gene expression profile at single cell level of mouse liver in whole and fibrotic niche of the liver [FiNi-seq]

status: Public on Feb 15 2025

city: Daejeon

first: Kwon Yong

last: Tak

address: Gu Sung Street

institute: KAIST

summary: Age-dependent fibrosis accumulated in the liver includes driver cell types of aging. We used single cell RNA sequencing (scRNA-seq) from specially digested liver to analyze the age-related fibrosis.

overall_design: The liver was digested to enriched the fibrotic niche and was sequenced in parallel with control data. Experiment was done in CCl4-treated mouse to see early fibrosis niche

first: Juyeon

first: Myungsun

last: Park

first: Chuna

first: Jong-eun


title: RNA-Seq analysis of IL-15 stimulated Tfh cells of kidney transplant recipients

city: Boston

first: Hengcheng

last: Zhang

state: Boston

address: 221 Longwood Ave

department: Medicine/Renal

institute: Brigham and Women's Hospital

summary: Assessment of transcriptional programs of Tfh isolated from lymph nodes recipients post-transplantation to investigate the effect of IL-15 on Tfh cells.

overall_design: Tfh cells were sorted from the lymph nodes of kidney transplant recipients and cultured with IL-15 for 6 hours. Bulk RNA-Seq was performed to profile transcriptional changes in response to IL-15 stimulation.

first: Peter

middle: T

last: Sage


title: Transcriptional and epigenetic rewiring by the NUP98::KDM5A fusion oncoprotein directly activates CDK12

first: Thomas

last: Eder

overall_design: Refer to individual Series


title: RNA-Seq analysis of germinal center B cells from Tfr-depleted and control kidney transplant recipients

summary: Assessment of transcriptional programs of germinal center B cells isolated from lymph nodes of Tfr-depleted (Tfr-DTR) and control (Tfr-Con) recipients post-transplantation to investigate the role of Tfr cells in kidney transplant rejection.

overall_design: Lymph node GC B cells were isolated from Tfr-depleted (Tfr-DTR) and control (Tfr-Con) kidney transplant recipients 20 days post-transplantation. Bulk RNA-Seq was performed to identify transcriptional differences.


title: Tfr Cells Restrain Rejection of Kidney Allografts by Limiting Pro-Inflammatory Cytokines in B cells [scRNA-seq]

summary: Pathogenic antibodies produced by alloreactive B cells mediate antibody-mediated rejection (ABMR) after kidney transplantation, but the mechanisms controlling these antibodies remain poorly understood. Follicular Regulatory T (Tfr) cells modulate follicular helper T (Tfh) cell-mediated B cell responses but their roles in controlling alloreactive antibody are unknown. Here, we studied the developmental signals and functions of Tfr cells in allogeneic kidney transplantation.  We show that costimulatory blockade alters the development of Tfr cells disproportionately by decreasing germinal center (GC)-like Tfr cells and increasing follicular-like Tfr cells. Functionally, Tfr cell deletion resulted in accelerated rejection and increases in donor-specific B cells in both draining lymph nodes and kidney allografts. Mechanistically, Tfr cell deletion increased GC B cell expression of pro-inflammatory cytokines such as IL-15. Neutralization of IL-15 compensated for the loss of Tfr cells and prolonged kidney transplant recipient survival. Together these data demonstrate the signals for Tfr cell development and how these cells restrain rejection by limiting alloreactive B cell responses.

overall_design: Lymph node T follicular regulatory cells and Treg cells all in duplicate


title: The expression order determines the pioneer functions of Ngn3 and NeuroD1 in pancreatic endocrine differentiation -2

city: Beijing

first: Cheng-ran

last: Xu

address: NO.5 YIHEYUAN ROAD HAIDIAN DISTRICT, BEIJING, P.R.CHINA

department: School of Basic Medical Sciences

institute: Peking University

summary: This study investigates the roles of Ngn3 and NeuroD1 in pancreatic endocrine differentiation. Using a new mouse model, we mapped NGN3 binding sites by CUT&RUN and demonstrated its pioneering role in making chromatin accessible by scATAC-seq. According to conditional knock-out and over-expression experiments and scRNA-seq, we domonstrated that NGN3's pioneering function was dose tolerance, with low doses sufficing, while NeuroD1, as a conventional TF, has no significant pioneer effects under normal phenomena. But when NGN3 was absent, NeuroD1 can alternatively acted as a pioneer factor, and sequential expression of NeuroD1 ahead of Ngn3 predominantly drives α-cell generation.

overall_design: Six types of datasets were generated in this project. 1. CUT&RUN of Ngn3-flag, NeuroD1 or P300 in Ngn3low or Ngn3high cells isolated from E13.5 or E14.5 fetal pancreas. CUT&RUN of NeuroD1 in GFP+ cells isolated from E15.5 Pdx1CreER;Neurod1OE or Ngn3KI;Ngn3Cre;Neurod1OE mice. 2. 10x genomics snATAC-seq of endocrine or pancreatic cells collected from Ngn3CreER;RossaRFP or Pdx1Cre;RossaRFP mice. 10x genomics snATAC-seq of Epcam+ cells collected from Neurod1KO or Ngn3Cre;Ngn3fl mice and their corresponding WT groups (CKO/OE and WT were distinguished by gender). 3. ATAC-seq of Ngn3low or Ngn3high cells in E13.5 fetal pancreas. ATAC-seq of the E13.5 GFP+ cells isolated from Ngn3KI mice, E15.5 or E17.5 Venus+ cells isolated from Neurod1KO mice, E15.5 GFP+ cells isolated from Pdx1CreER;Ngn3fl;Neurod1OE, Pdx1CreER;Neurod1OE or Ngn3KI;Ngn3Cre;Neurod1OE mice. 4. 10x genomics scRNA-seq of the Epcam+ cells isolated from E14.5 Ngn3Cre;Ngn3fl, E14.5 or E17.5 Ngn3Cre;Ngn3OE and E15.5 Neurod1KO mice and their corresponding control WT groups  (CKO/OE and WT were distinguished by gender). 5. Smartseq3 scRNA-seq of the E12.5 or E15.5 GFP+ cells isolated from Pdx1CreER;Ngn3fl;Neurod1OE, Pdx1CreER;Neurod1OE or Ngn3KI;Ngn3Cre;Neurod1OE mice, and Epcam+ cells isolated from WT mice. 6. Smartseq2 scRNAseq of Ngn3flagGFPlow, Ngn3flagGFPhigh and GFP+ cells isolated from Ngn3KI mice cells.

first: Liu

last: Yang

first: Xin-Xin

last: Yu

first: Xin

last: Wang

first: Chen-Tao

last: Jin

first: Cheng-Ran


title: Tfr Cells Restrain Rejection of Kidney Allografts by Limiting Pro-Inflammatory Cytokines in B cells.


title: Mesenchymal stem cell with enhanced antioxidant capacity integrates into smooth muscle cells in diabetic detrusor underactivity (Affymetrix)

first: DONG-MYUNG

last: SHIN

overall_design: Human umbilical cord-derived mesenchymal stem cells (UC-MSCs) were harvested in the normal cultured UC-MSCs (Cultured #) and the Resovist+/GFP+ engrafted cells (Engrafted #) from bladders of rats in which type I diabetes at 7 DAT. Total RNA from single UC-MSC engrafted rat bladder by MACS-FACS sorting was isolated using the cell lysis buffer, including treatment with NP40, Rnase Inhibitors, and V1(dT)24 primer(5 ng/ul). One ug of total RNA was used for the Affymetrix GeneChip Human Genome U133 Plus 2.0 Array (Affymetrix, Santa Clara, CA), in accordance with the manufacturer’s instructions.

first: Dong-Myung

last: Shin

first: Chae-Min

last: Ryu

first: YongHwan

last: Kim


title: Transcriptional Heterogeneity and Differential Response of Rod Photoreceptor Pathway Uncovered by Single‐Cell RNA Sequencing of the Aging Mouse Retina

first: Matthew

last: Brooks

summary: Gene expression of single cells from mouse retina during aging.

overall_design: Mouse retinas from 3, 12, 18, and 24 months of age were used to generate scRNA profiles. For each age both whole retina and CD73 depleted retina dissociate populations were used as input into 10x Genomics Single Cell 3' v3.1.

first: Laura

last: Campello

first: Benajmin

last: Fadl

first: Soumitra

last: Pal

first: Anand

last: Swaroop


title: A dihydrochalcone-specific O-methyltransferase from leaf buds of Populus trichocarpa implicated in bud resin formation

city: Victoria

first: Dawei

last: Ma

state: BC

address: 3800 Finnerty Road

department: Biology

country: Canada

institute: University of Victoria

summary: We used transcriptomics and differential gene expression analyses of P. trichocarpa and P. balsamifera leaf buds to identify a gene encoding a P. trichocarpa dihydrochalcone-specific O-methyltransferase, which we named PtDOMT1. Detailed enzymatic analyses demonstrated PtDOMT1 to be a highly selective and regiospecific O-methyltransferase which methylates 4 and 4' positions of dihydrochalcones using S-adenosyl-L-methionine as a methyl donor.

sample_organism:
Populus trichocarpa
Populus balsamifera

platform_organism:
Populus trichocarpa
Populus balsamifera

overall_design: Dormant cuttings of P. balsamifera and P. trichocarpa, harvested in February, were induced to flush in a growth chamber. The cutting were ~30 cm long and were stored at 4 °C in the dark until the controlled experiment. The twigs were placed in water in a growth chamber at 22 °C under 16 h of light and sampled at 2 d intervals. For each time point, leaf buds were collected from one individual cutting as a replicate to prevent wound-induced effects during the time course. Samples were collected from three replicates of P. balsamifera and two replicates of P. trichocarpa. Each sampling consisted of 10 leaf buds, and samples were collected over 12 d. Samples collected at day 0 (D0), day 4 (D4), and day 10 (D10) were selected to represent dormant, pre-bud break, and bud break phases of the lateral leaf buds, respectively. Three biological replicates of P. balsamifera and two biological replicates of P. trichocarpa were used to prepare RNA-seq libraries as previously described using the NEBNext® Ultra™ RNA Library Prep Kit for Illumina® (New England Biolabs)

first: Eerik-Mikael

last: Piirtola

first: Peter

last: Constabel


title: TRAF3 regulates TFH cell differentiation during infections (ATAC-Seq)

city: Iowa City

first: Noah

middle: S

last: Butler

state: IA

address: 4-350 BSB, 51 Newton Road

department: Department of Microbiology and Immunology

institute: University of Iowa

summary: Comparison of chromatin accessibility between WT and tamoxifen-inducible deletion of TRAF3 (iTraf3−/−)  SMARTA TFH cells during LCMV armstrong infection

overall_design: Tamoxifen-inducible deletion of TRAF3 (iTraf3−/−) and WT SMARTA TCR transgenic CD4+ T cells were transferred to recipient mice, followed by  LCMV armstrong infection. On day 5 post-infection, WT or iTraf3−/− SMARTA CXCR5+SLAMlo TFH cells were sorted into RP10 medium. ATAC-seq was performed to compare the differential chromatin accessibility among the iTraf3−/− and WT SMARTA TFH cells.

first: Peng

last: Shao


title: TRAF3 regulates TFH cell differentiation during infections

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


title: GPR37L1 identifies spinal cord astrocytes and protects neuropathic pain after nerve injury

status: Public on Feb 14 2025

city: DURHAM

first: Jing

last: Xu

state: NC

address: 816 Louise Circle

institute: Duke university

summary: Astrocytes in the spinal cord dorsal horn (SDH) play a pivotal role in synaptic transmission and neuropathic pain. However, the precise classification of SDH astrocytes in health and disease remains elusive. Here we reveal Gpr37l1 as a marker and functional regulator of spinal astrocytes. Through single-nucleus RNA sequencing, we identified Gpr37l1 as a selective GPCR marker for spinal cord astrocytes. Notably, SDH displayed reactive astrocyte phenotypes and exacerbated neuropathic pain following nerve injury combined with Gpr37l1 deficiency. In naïve animals, GPR37L1 knockdown in SDH astrocytes induces astrogliosis and pain hypersensitivity, while Gpr37l1-/- mice fail to recover from neuropathic pain. GPR37L1 activation by maresin-1 increased astrocyte GLT-1 activity and reduced spinal EPSCs and neuropathic pain. Selective overexpression of Gpr37l1 in SDH astrocytes reversed neuropathic pain and astrogliosis after nerve injury. Our findings illuminate astrocyte GPR37l1 as an essential negative regulator of pain, which protects neuropathic pain through astrocyte signaling in SDH.

overall_design: We performed single-nucleus RNA sequencing (snRNA-seq) on WT and Gpr37l1-/- mice with or without 1-day spared nerve injury (SNI)

first: Zihan

last: Yan

first: Sangsu

last: Bang

first: Dmitry

last: Velmeshev

first: Ru-Rong

last: Ji


title: Mammary intraepithelial lymphocytes promote lactogenesis and offspring fitness

first: Eduard

last: Ansaldo

address: 4 Memorial Drive

department: LHIM

institute: NIH/NIAID

summary: To test the role of intraepithelial lymphocytes in mammay grand epithelial function during pregnancy we performed single cell RNA sequencing of epithelial and myoepithelial cells from the mammary gland in WT mice, Raggc-KO mice,  Raggc-KO mice adaptively transferred with IEL precursors.

overall_design: For mammary epithelial and myoepithelial scRNA-seq, thymic IELp were adoptively transferred (+IELp) or not (-IELp) into pregnant Rag2-/-c-/- /- at day 10.5 of gestation. Mammary epithelial cells and myoepithelial cells (CD45-CD31-EPCAM+) from Rag2-/-c-/- mice with or without IELp were sorted 8 days post-transfer (G18.5) from samples labeled with TotalSeqC hashtags antibodies (Biolegend) with a Sony MA900 sorter. Sorted cells were pooled together and 45,000 cells were loaded to a Chromium Single Cell Controller (10X Genomics) to encapsulate the cells into droplets (1 lanes).

first: Dan

last: Corral

first: Cihan

last: Oguz


summary: T cells accumulate in the mammary gland during pregnancy and lactation. In order to understand the diversity, transcriptional states, TCR repertoire, and function of mammary gland T cells, and how these change during pregnancy and lactation compared to homeostasis, we performed single cell RNA sequencing on mammary gland T cells in mice.

overall_design: Mammary T cells from nulliparous,  pregnant (G18.5), and lactating (L5) mice were sorted from samples labeled with TotalSeqC hashtags antibodies (Biolegend) using a Sony MA900 sorter. Sorted cells were pooled and 42000 cells per lane (total of 8 lanes) were loaded on a Chromium Single Cell Controller (10X Genomics) to encapsulate the cells into droplets. The gene expression (GEX),  hashtag oligonucleotide (HTO) and VDJ (TCR) sequencing libraries were prepared using Chromium Single Cell 5’ v2 reagent kits following the manufacturer’s instructions, and sequenced on an Illumina Nextseq1000 platform (NextSeq 1000/2000 P2 reagents (100 cycles)).

first: Mahnoor

last: Abid


title: Itaconate modulates immune responses via inhibition of peroxiredoxin 5

city: St Louis

first: Maxim

middle: N.

last: Artyomov

state: Missouri

address: 660 S Euclid Ave

department: Pathology & Immunology

institute: Washington University in St Louis

summary: Gene expression in WT BMDMs pretreated with itaconic acid, malonic acid or methylsuccinic acid.

overall_design: BMDMs were pretreated with media supplemented with metabolites (5 mM final concentration) itaconic acid (IA), malonic acid(MA) or methylsuccinic acid (MetSA) for 16 h. Control cells were treated in media (M) alone. Each condition is performed in 3 biological replicates.

first: Tomas

last: Paulenda

first: Barbora

last: Echalar

first: Lucie

last: Potuckova

first: Veronika

last: Vachova

first: Denis

last: Kleverov

first: Johannes

last: Mehringer

first: Ekaterina

last: Potekhina

first: Alex

last: Jacoby

first: Devashish

last: Sen

first: Chris

last: Nelson

first: Rick

last: Stegeman

first: Vladimir

last: Sukhov

first: Danielle

last: Kemper

first: Cheryl

middle: F

last: Lichti

first: Nicholas

last: Day

first: Tong

last: Zhang

first: Monika

last: Bambouskova

first: Daved

middle: H

last: Fremont

first: Wei-jun

last: Qian

first: Sergej

last: Djuranovic

first: Slavica

last: Pavlovic-Djuranovic

first: Vsevolod

middle: V

last: Belousov

first: Andrzej

middle: M

last: Krezel


title: Gene expression analysis of human esophageal cancer cells co-cultured with Fusobacterium nucleatum

city: Fukushima City

first: Shotaro

last: Nakajima

state: Fukushima

address: 1 Hikariga-oka

department: School of Medicine

institute: Fukushima Medical University

summary: This study analyzed changes in gene expression caused by infection with Fusobacterium nucleatum (Fn) in human esophageal cancer (EC) cells. The hypothesis was that Fn infection affects the activation of the stimulator of interferon genes (STING) signaling pathway and the expression of its downstream genes in EC cells. Results revealed interesting upregulated genes related to the STING signaling pathway in response to Fn infection.

overall_design: To investigate the effect of Fn infection on gene expression in esophageal cancer (EC) cells, EC cell lines were co-cultured with Fn in RPMI-1640 without fetal bovine serum (FBS) and penicillin/streptomycin for 6 hours in a 5% CO₂ atmosphere at 37°C.

first: Kyoko

last: Nishiyama


title: Identification of human T cells that recognize a mycobacterial adjuvant, trehalose monomycolate (TMM)

city: Suita

first: Daisuke

last: Motooka

state: Osaka

address: 3-1, Yamadaoka

institute: NGS core facility, Research Institute for Microbial Diseases, Osaka University

summary: Mycobacterium tuberculosis (M. tb) causes tuberculosis (TB) which remains one of the world’s deadliest diseases. As M.tb are protected by its thick lipid cell wall, the host has developed specific immune receptors that recognize their unique lipids as antigens. Among them is a mycolipid-specific T cells restricted by common antigen-presenting molecules, such as CD1. However, complete understanding of their subsets, clonotypes and antigen-specificity has not been fully achieved. Here, we identified novel mycolipid-specific T cells by comprehensive screening using lipid biochemical ´ single cell-TCR-RNA-sequencing-based clonotypic analysis. When human PBMCs were co-cultured with mycobacterial crude lipid extract, a CD4+ CTL-like clone, Y-50, was highly proliferated. Indeed, NFAT-GFP reporter cells reconstituted TCRαβ chains expressed by Y-50 strongly reacted with crude lipids in the presence of monocyte-derived dendritic cells from syngenic/allogenic donors. Activity-based purification of crude lipids identified trehalose monomycolate (TMM) as an antigen. CD1b was necessary and sufficient for the presentation of TMM to be recognized by Y-50. TMM-loaded CD1b tetramer-positive T cells were detected in PBMCs and cord blood cells. The frequency of tetramer+ T cells in PBMCs was higher in active TB patients than uninfected donors. These results demonstrate the presence of novel unconventional T cells in humans that recognize TMM.

overall_design: Reconstituted CDR3 mutant library cells were stimulated with TMM for 20 h and then sorted by GFP negative/positive populations. Unstimulated library cells were uesed as a control group. Each sorted cells was analyzed by bulk TCR-sequencing

first: Yuki

last: Sakai

first: Wakana

last: Kusuhara

last: Okuzaki

first: Sho

last: Yamasaki


title: Multilevel Stressors and Systemic and Tumor Immunity in Black and White Women With Breast Cancer

first: Stefan

last: Ambs

address: Building 37, Room 3050B

department: CCR

institute: NCI

summary: A cross-sectional study was conducted between 2012 and 2023 to collect breast tumor and tumor-adjacent normal tissue (N=48 tumor, 41 tumor-adjacent normal) from women with breast cancer. Participants were recruited through the University of Maryland Medical Center and other hospital sites within the area of Baltimore, Maryland. RNA-sequencing and whole exome sequencing was performed on these fresh-frozen tissues. Genomic DNA was extracted from frozen breast tumors and adjacent normal tissue using the Qiagen DNeasy Blood & Tissue kit and checked for quality using the Agilent Genomic DNA Screen tape assay. WES was performed by the service provider Psomagen and data processed as in Tang, et al, 2023.

overall_design: RNA was isolated from frozen breast tumors using TRIzol method. The mRNA profiles of the above breast tumors were generated by deep sequencing using Illumina NovaSeq 6000.

first: Alexandra

last: Harris

first: Catherine

last: Pichardo

first: Jamirra

last: Franklin

first: Huaitian

last: Liu

first: Wayne

last: Lawrence

first: Brittany

middle: D

last: Jenkins-Lord

first: Amanda

last: Acevedo

first: Tiffany

last: Dorsey

first: Tanya

last: Agurs-Collins


title: Identification of CD1b-restricted human T cells recognizing a mycobacterial adjuvant, trehalose monomycolate (TMM)

summary: Mycobacterium tuberculosis causes tuberculosis (TB) which remains one of the world’s deadliest diseases. As mycobacteria are protected by its thick lipid cell wall, the host has developed immune systems targeting their unique lipids. Among them is an unconventional T cell subset called mycolipid-specific T cells restricted by common antigen-presenting molecules, such as CD1. Here, we identified novel mycolipid-specific T cells using a method combining bulk T cell stimulation with crude lipids and single cell-TCR-RNA-sequencing-based clonotypic analysis. In human PBMCs, a CD4+ T cell clone with cytotoxic signature, Y-50, was proliferated in response to mycobacterial crude lipid extract. Activity-based purification of using reporter cells reconstituted with Y-50 TCR identified trehalose monomycolate (TMM) as an antigen. CD1b was necessary and sufficient for the presentation of TMM to be recognized by Y-50. Y-50 TCRab chains displayed characteristic features, such as positively-charged CDR3a and extra-long CDR3b. Mutagenesis and structural analysis of TCRab suggested that these unique CDR3 properties were required for the recognition of TMM presented by CD1b. CD1b tetramer loaded with TMM could detect T cells with similar characteristics to Y-50 in uninfected healthy donors, but their frequency was significantly higher in PBMCs from TB patients. In this study, we identified pre-set unconventional T cells specific for a mycobacterial adjuvant, TMM.

overall_design: PBMCs from healthy donors,TB active patients or LTBI were stnined with CD1b-TMM-tetramers. The tetramer posivie T cells were collected by cell sorter and analyzed by scRNA-TCR-seq.


title: Efficacy of Polo-life kinase 1 inhibitors in preclinical models of small cell lung cancer

city: Baltimore

first: John

middle: Christopher

last: Schmitz

address: 655 W. Baltimore Street

department: Medicine

institute: University of Maryland Baltimore

summary: Small cell lung cancer (SCLC) remains a significant health problem with more than 180,000 newly afflicted patients worldwide per year. Recurrence following initial frontline therapy is associated with resistance to salvage treatment and remains the area of greatest unmet need for SCLC patients. This study systematically examined the efficacy of three different polo like kinase 1 (PLK1) inhibitors in preclinical models of  SCLC. The study uncovered robust in vitro activity of PLK1 inhibitors against SCLC cell lines and confirmed the efficacy in patient-derived xenograft models of platinum sensitive and resistant SCLC.  A known challenge of targeted therapies in the clinic is the inevitable development of resistance and loss of efficacy. Therefore, understanding the potential mechanisms driving acquired resistance could offer additional strategies to enhance efficacy, delay the development of resistance or reverse the resistance. We generated lab-derived resistant SCLC cells by continuous exposure of parental cells to increasing concentration of onvansertib and performed RNA-seq to identify potential resistance mechanisms.

overall_design: To develop onvansertib-resistant SCLC H526 cells, H526 cells were grown with increasing onvansertib concentrations (10 nM up to 75 nM) over a one year period. Total RNA was isolated and submitted for RNA-seq.

first: Zhentao


title: Copper-enriched automotive brake wear particles perturb human alveolar cellular homeostasis

status: Public on Feb 13 2025

city: Southampton

first: James

middle: G. H.

last: Parkin

address: Level F, University Hospital Southampton

institute: University of Southampton

summary: Background Airborne fine particulate matter with diameter < 2.5 μm (PM2.5), can reach the alveolar regions of the lungs, and is associated with over 4 million premature deaths per year worldwide. However, the source-specific consequences of PM2.5 exposure remain poorly understood. A major, but unregulated source is car brake wear, which exhaust emission reduction measures have not diminished.AQ1   Methods We used an interdisciplinary approach to investigate the consequences of brake-wear PM2.5 exposure upon lung alveolar cellular homeostasis using diesel exhaust PM as a comparator. This involved RNA-Seq to analyse global transcriptomic changes, metabolic analyses to investigate glycolytic reprogramming, mass spectrometry to determine PM composition, and reporter assays to provide mechanistic insight into differential effects.   Results We identified brake- wear PM from copper-enriched non-asbestos organic, and ceramic brake pads as inducing the greatest oxidative stress, inflammation, and pseudohypoxic HIF activation (a pathway implicated in diseases associated with air pollution exposure, including cancer, and pulmonary fibrosis), as well as perturbation of metabolism, and metal homeostasis compared with brake wear PM from low- or semi-metallic pads, and also, importantly, diesel exhaust PM. Compositional and metal chelator analyses identified that differential effects were driven by copper.   Conclusions We demonstrate here that brake-wear PM may perturb cellular homeostasis more than diesel exhaust PM. Our findings demonstrate the potential differences in effects, not only for non-exhaust vs exhaust PM, but also amongst different sources of non-exhaust PM. This has implications for our understanding of the potential health effects of road vehicle-associated PM. More broadly, our findings illustrate the importance of PM composition on potential health effects, highlighting the need for targeted legislation to protect public health.

overall_design: We exposed alvolar type-II epithelical cells from a cell line to 5 different types of vehicle-derived PM

first: Lareb

last: Dean

first: Joseph

last: Bell

first: Natasha

last: Easton

first: Liam

last: Edgeway

first: Matthew

last: Cooper

first: Robert

last: Ridley

first: Franco

last: Conforti

first: Siyuan

last: Wang

first: Liudi

last: Yao

first: Juanjuan

last: Li

first: Helen

last: Vethakan Raj

first: Julian

last: Downward

first: Miriam

last: Gerlofs-Nijland

first: Flemming

last: Cassee

first: Yihua

first: Richard

middle: B

last: Cook

first: Mark

last: Jones

first: Donna

last: Davies

last: Loxham


title: Effects of type I IFN signaling on gene expression of Mycobacterium tuberculosis-infected bone marrow-derived macrophages (BMDMs)

city: New York

first: Ziwei

last: Yang

state: NY

address: 413 East 69th Street

institute: Weill Cornell Medical College

summary: Our previous work identified that type I IFNs mediate macrophage death after infection of Mycobacterium tuberculosis (Mtb) through a novel mechanism.Type I IFN signaling relies on its several hundred ISGs to execute various functions. In order to delineate the downstream mechanism of type I IFN-mediating cell death, we adopted RNA-seq experiments to identify which gene(s) are differentially expressed in WT or Ifnar1-/- bone marrow-derived macrophages (BMDMs). In the meantime, we're also interested in how genetic background affects transcriptomics in BMDMs infected with Mtb. Thus a transcriptomics analysis of C57BL/6, 129S2 and 129X1 infected with Mtb were also conducted.

overall_design: RNA-seq profiling of: 1) WT and Ifnar1-/- BMDMs (in the background of C57BL/6) infected with Mtb for 24h. 2) C57BL/6, 129S2, 129X1 BMDMs infected with Mtb for 24h.

first: Li

last: Zhang

first: Carl

last: Nathan


title: Fission yeast Caprin protein is required for efficient heterochromatin establishment

city: Edinburgh

first: Elizabeth

last: Bayne

address: Alexander Crum Brown Road

department: School of Biological Sciences

institute: University of Edinburgh

summary: Heterochromatin is a key feature of eukaryotic genomes that serves important regulatory and structural roles in regions such as centromeres. In fission yeast, maintenance of existing heterochromatic domains relies on positive feedback loops involving histone methylation and non-coding RNAs. However, requirements for de novo establishment of heterochromatin are less well understood. Here, through a cross-based assay we have identified a novel factor influencing the efficiency of heterochromatin establishment. We determine that the previously uncharacterised protein is an ortholog of human Caprin1, an RNA-binding protein linked to stress granule formation. We confirm that the fission yeast ortholog, here named Cpn1, also associates with stress granules, and we uncover evidence of interplay between heterochromatin integrity and ribonucleoprotein (RNP) granule formation, with heterochromatin mutants showing reduced granule formation in the presence of stress, but increased granule formation in the absence of stress. We link this to regulation of non-coding heterochromatic transcripts, since in heterochromatin-deficient cells, Cpn1 can be seen to colocalise with accumulating pericentromeric transcripts, and absence of Cpn1 leads to hyperaccumulation of these RNAs at centromeres. Together, our findings unveil a novel link between RNP homeostasis and heterochromatin assembly, and implicate Cpn1 and associated factors in facilitating efficient heterochromatin establishment by enabling removal of excess transcripts that would otherwise impair assembly processes.

overall_design: To investigate the potential function of  Schizosaccharomyces pombe  Cpn1 in transcriptome regulation, we performed RNA-seq on WT and cpn1∆ cells (3 biological replicates for each) for differential expression analysis.


title: An antisense oligomer conjugate with unpredicted bactericidal activity against Fusobacterium nucleatum

city: Würzburg

first: Jakob

middle: Jeremias

last: Jung

state: Bayern

address: Josef-Schneider-Str. 2/Bau D15

department: Institute of Molecular Infection Biology (IMIB)

country: Germany

institute: University of Würzburg

summary: Fusobacteria are commensal members of the oral microbiome that can spread from their primary niche and colonize distal sites in the human body. Their enrichment in colorectal and breast cancer tissue has been associated with tumor growth, metastasis, and chemotherapeutic resistance. The use of non-selective antibiotics to remove fusobacteria impairs tumor progression, but prolonged application risks side effects such as gastrointestinal problems and dysbiosis. Species-specific antisense antibiotics based on peptide nucleic acid (PNA) have shown efficacy in many Gram-negative species, suggesting that antisense PNAs may also enable a tailored depletion of fusobacteria. Here, we have investigated the antibacterial potential of cell penetrating peptide (CPP)-PNA conjugates targeting the mRNA of putative essential genes in Fusobacterium nucleatum. Unexpectedly, we observed no growth inhibition with any of the target-specific PNAs, but identified a non-targeting control CPP-PNA (FUS79) as a potent growth inhibitor of F. nucleatum. Our data suggest that the CPP and specific sequence features of FUS79 are responsible for its activity, rather than an antisense effect. Interestingly, FUS79 also inhibits the growth of five additional fusobacterial strains but not of Fusobacterium nucleatum ssp. vincentii (FNV). RNA-seq analysis suggests that FUS79 induces a membrane stress response in a vulnerable F. nucleatum strain but not in FNV. Collectively, our attempt at developing antisense antibiotics for fusobacteria discovers a potent growth inhibitor, whose bactericidal effect appears independent of target-specific mRNA inhibition.

overall_design: RNA-Seq of 2 strains of Fusobacterium nucleatum (FNN23 and FNV), treated with 10µM target PNA, scrambled PNA and water (control) over 2 time points (18 samples).  Illlumina NextSeq 500 was used

first: Valentina

last: Cosi

middle: J

first: Linda

last: Popella

first: Chandradhish

last: Ghosh

first: Lars

last: Barquist

first: Jörg

last: Vogel


title: Pou4f2/Brn3b Overexpression Promotes the Genesis of Retinal Ganglion Cell-like Projection Neurons from Late Progenitors

city: Salt Lake City

first: Monica

middle: L.

last: Vetter

state: UT

department: Neurobiology & Anatomy

institute: University of Utah

summary: Retinal ganglion cells (RGCs) are the projection neurons of the retina. In early retinal progenitor cells (RPCs), Atoh7 orchestrates the developmental RGC program and regulates the expression of critical downstream targets, including Pou4f factors. The absence of Pou4f2 or more POU4F family genes results in defects in RGC differentiation, aberrant axonal elaboration and ultimately RGC death, confirming the requirement of POU4F factors for RGC development and survival, with a critical role in regulating RGC axon outgrowth and pathfinding. Here, we investigated in vivo whether ectopic Pou4f2 expression in late retinal progenitor cells (late RPCs) is sufficient to induce the generation of cells with RGC properties, including projecting axons to the brain. Using a strong ubiquitous promoter to induce Pou4f2 overexpression in neonates, we observed a change in targeted cell distribution in the retinal tissue, including the presence of cells in the ganglion cell layer and inner plexiform layer. Single-cell RNA sequencing (scRNA-seq) analysis shows that several RGC-genes (Rbpms, Gap-43, Hs6st3, and Foxp2) are upregulated after Pou4f2 overexpression. Additionally, gene ontology analysis indicates the induction of genes related to axonogenesis and neuronal differentiation. Imaging throughout the visual pathway revealed high density of axons projecting toward the optic nerve head and extending to brain, such as the superior colliculus and geniculate nucleus. Thus, Pou4f2 induced neurons with specific RGC characteristics that share similarities with resident RGCs and notably project axons that reach brain targets. In conclusion, these results showed that POU4F2 alone was sufficient to promote aspects of RGC development and axon projection, inducing critical properties of projection neurons from retinal progenitors outside their developmental window.

overall_design: scRNA-seq of retinal neurons generated from late retinal progenitors treated with Pou4f2 or GFP alone

first: Viviane

last: Oliveira-Valença

first: Jacqueline

last: Roberts

first: Alejandra

last: Bosco

middle: L

first: Mariana

last: Silveira


title: Palmitoylation prevents B7-H4 lysosomal degradation sustaining tumor immune evasion

city: Ann Arbor

first: Weiping

last: Zou

state: Michigan

address: 109 Zina Pitcher Place

institute: University of Michigan

summary: To investigate the function of CDK4/6 inhibitor- Abemaciclib on lysosome function.

overall_design: We performed gene expression profiling analysis in human breast cancer cell line- MDA-MB-468  with or without  Abemaciclib for 12 hours.

first: Yijian

last: Yan

first: Jiali

last: Yu

first: Weichao

last: Wang


title: snRNAseq of mouse kidney after CACPR

city: Los Angeles

first: Jonathan

middle: William

last: Nelson

state: CA

address: 1333 San Pablo St.

institute: University of Southern California

summary: Acute kidney injury (AKI) is a lethal syndrome with many antecedents.  Transcriptomic and phenotypic investigations have demonstrated distinct patterns of renal injury which depend in part on the etiology of kidney injury. AKI following acute cardiac dysfunction is termed acute cardiorenal syndrome (CRS1). Cardiac arrest and cardiopulmonary resuscitation (CACPR) in the mouse models CRS1, and important translational insights have been generated in this model.  Here, we report the results of comparison between mRNA transcriptomics performed on CACPR-exposed mouse kidneys and those obtained from IRI-exposed mouse kidney. The IRI-exposed mouse kidney data comes from the Humphreys lab (PMID: 32571916)

overall_design: Kidney nuclei from Sham and CACPR mice isolated sequenced on the 10X Chromium Controller.

first: Michael

last: Hutchens

first: Kevin

last: Burfeind

first: Yoshio

last: Funahashi


title: Combining the CowPEAsy web application with in planta agroinfiltration for native promoter validation in Vigna unguiculata

city: Urbana

first: Lauriebeth

last: Leonelli

state: Illinois

address: 1206 W Gregory Dr.

department: Department of Agricultural and Biological Engineering

institute: University of Illinois Urbana-Champaign

summary: Cowpea (Vigna unguiculata) is an important protein source in Sub-Saharan Africa. Optimizing resilience and productivity through genetic engineering in cowpea has been slow due in part to a lack of defined species-specific regulatory elements and difficulty testing gene function within the native system. In many plant species, Agrobacterium-mediated transient gene expression is widely used to validate constructs before investing in transgenic lines, but its implementation in legumes has been challenging. In this study, we optimized an in planta agroinfiltration assay in trifoliate cowpea leaves using a betalain reporter. To demonstrate the “intact plant” aspect of this system, we used this assay to characterize drought-inducible promoters by challenging cowpea plants with drought stress. Subsequently, to identify and broaden the pool of native promoters known in cowpea, we developed a user-friendly web application, CowPEAsy, allowing users to interrogate gene expression from our canopy-level, developmental-series RNA-Seq dataset. Finally, using CowPEAsy, we identified six promoters that showed constitutive expression across all conditions and verified these promoters with our transient system. This work provides an in vivo platform for preliminary validation of regulatory elements in cowpea and other legumes and enhances current genetic resources by identifying a suite of physiologically relevant promoters of varying strengths.

overall_design: Leaf tissue punches for RNA-Seq were collected from greenhouse grown IT97K-499-35 cowpea plants at bottom, middle, and top canopy positions and at developmental stages R1 (5.5 weeks after planting) and R3 (6.5 weeks after planting).

first: Jake

last: Harris

first: Jooyeon

last: Jeong

first: Larissa

last: de Souza


title: ICOSL, OX40L, and CD30L Control Persistence of Asthmatic CD4 Trm Cells

city: La Jolla

last: Croft

address: 9420 Athena Circle

institute: La Jolla Institute for Immunology

summary: Tissue-resident memory T cells (Trm) are central to maintaining autoimmune and inflammatory disease. Modulation of their continued replenishment in tissues has significant implications for clinical treatment. In a murine model of lung inflammation, scRNA-seq revealed the complexity of antigen-responding CD4+ Trm, encompassing many pathogenic subpopulations including Th2, Th17, Th1 and CTL, but suggested several targets for therapeutic intervention, with significant co-expression of the costimulatory molecules OX40, ICOS, CD30, and CD30L throughout these T cells. Inhibiting ICOSL alone, or co-blocking OX40L and CD30L, with neutralizing antibodies, only partially suppressed the response of Trm to recall antigen, whereas inhibiting all three molecules strongly reduced the accumulation of Trm-derived effector memory T cells, and ablated all aspects of lung inflammation. Most importantly, transient therapeutic inhibition of these molecules together prevented the continued accumulation and long-term persistence of induced Trm, leading to a state of tolerance such that subsequent exposure to antigen failed to re-establish a pathogenic inflammatory response. These data show that costimulatory molecules are critical for reactivation and persistence of pathogenic CD4+ Trm, and reveal several therapeutic combinations that are applicable for treatment of lung inflammatory disease and potentially multiple autoimmune diseases.

overall_design: Mice were sensitized by injecting a mixture of 20 μg HDM (Greer Labs, Lenoir, NC) and 2 mg alum intraperitoneally (i.p.), followed by a resting period of 10 days. Sensitized mice were then exposed to 10 μgHDM via theintranasal (i.n.) route for four consecutive days (10 to 13) in a secondary response. Challenged mice were then rested for another period of 2 weeks and re-exposed to 10 μg HDM i.n. for four consecutive days (27-30) in a tertiary response. Secondary and tertiary challenged mice were rested and subjected a fourth time to 10 μg HDM on days 44 and 45 as a quaternary challenge. To monitor lung-localized memory T cells, mice were subjected to short-term labeling with APC eF780-labelled anti-CD90.2 (2 μg in 100ul of PBS; clone 30-H12; Invitrogen) injected via the retro-orbital sinus, 10 minutes before tissue collection. Memory T cells present within tissues protected from this antibody staining are referred to as extravascular tissue-resident cells. Lungs were analyzed at varying end-points for single-cell RNA sequencing of lung extravascular memory T cells.

first: Gurupreet

last: Sethi

first: Ashmitaa

last: Logandha Ramamoorthy Premlal


title: The expression order determines the pioneer functions of Ngn3 and NeuroD1 in pancreatic endocrine differentiation

city: Beijing

first: Cheng-ran

last: Xu

address: NO.5 YIHEYUAN ROAD HAIDIAN DISTRICT, BEIJING, P.R.CHINA

institute: Peking University

summary: This study investigates the roles of Ngn3 and NeuroD1 in pancreatic endocrine differentiation. Using a new mouse model, we mapped NGN3 binding sites by CUT&RUN and demonstrated its pioneering role in making chromatin accessible by scATAC-seq. According to conditional knock-out and over-expression experiments and scRNA-seq, we domonstrated that NGN3's pioneering function was dose tolerance, with low doses sufficing, while NeuroD1, as a conventional TF, has no significant pioneer effects under normal phenomena. But when NGN3 was absent, NeuroD1 can alternatively acted as a pioneer factor, and sequential expression of NeuroD1 ahead of Ngn3 predominantly drives α-cell generation.

overall_design: Six types of datasets were generated in this project. 1. CUT&RUN of Ngn3-flag, NeuroD1 or P300 in Ngn3low or Ngn3high cells isolated from E13.5 or E14.5 fetal pancreas. CUT&RUN of NeuroD1 in GFP+ cells isolated from E15.5 Pdx1CreER;Neurod1OE or Ngn3KI;Ngn3Cre;Neurod1OE mice. 2. 10x genomics snATAC-seq of endocrine or pancreatic cells collected from Ngn3CreER;RossaRFP or Pdx1Cre;RossaRFP mice. 10x genomics snATAC-seq of Epcam+ cells collected from Neurod1KO or Ngn3Cre;Ngn3fl mice and their corresponding WT groups (CKO/OE and WT were distinguished by gender). 3. ATAC-seq of Ngn3low or Ngn3high cells in E13.5 fetal pancreas. ATAC-seq of the E13.5 GFP+ cells isolated from Ngn3KI mice, E15.5 or E17.5 Venus+ cells isolated from Neurod1KO mice, E15.5 GFP+ cells isolated from Pdx1CreER;Ngn3fl;Neurod1OE, Pdx1CreER;Neurod1OE or Ngn3KI;Ngn3Cre;Neurod1OE mice. 4. 10x genomics scRNA-seq of the Epcam+ cells isolated from E14.5 Ngn3Cre;Ngn3fl, E14.5 or E17.5 Ngn3Cre;Ngn3OE and E15.5 Neurod1KO mice and their corresponding control WT groups  (CKO/OE and WT were distinguished by gender). 5. Smartseq3 scRNA-seq of the E12.5 or E15.5 GFP+ cells isolated from Pdx1CreER;Ngn3fl;Neurod1OE, Pdx1CreER;Neurod1OE or Ngn3KI;Ngn3Cre;Neurod1OE mice, and Epcam+ cells isolated from WT mice. 6. Smartseq2 scRNAseq of Ngn3flagGFPlow, Ngn3flagGFPhigh and GFP+ cells isolated from Ngn3KI mice cells.

first: Liu

last: Yang

first: Xinxin

first: Xin

first: Chen-Tao

last: Jin

first: Cheng-Ran


title: mRNA export factors store nascent transcripts within nuclear speckles as an adaptive response to transient global inhibition of transcription [SLAM-seq]

city: Melbourne

first: Mark

last: Dawson

state: VIC

address: 305 Grattan Street

country: Australia

institute: Peter MacCallum Cancer Centre

summary: Several transcription inhibitors have been developed as cancer therapies. However, they show modest clinical activity highlighting that our understanding of the cellular response to transcriptional inhibition remains incomplete. Here we report that potent inhibitors of transcription not only impact mRNA output, but also markedly impair mRNA transcript localisation and nuclear export. We demonstrate that retention of newly transcribed mRNA in nuclear speckles is an adaptive response to chemically distinct transcriptional inhibitors. Retained transcripts are fully processed and accumulate in proportion to the expression level of the genes from which they emanate. The TREX mRNA export complex plays an integral role in directing nascent transcripts to nuclear speckles where they are bound to NXF1, protected from degradation, and poised for rapid export following re-initiation of transcription. Our findings provide new insights into the crosstalk between transcription and mRNA export, with important implications for drugs aiming to inhibit transcription for therapeutic gain.

overall_design: full-length SLAMseq of MDA-MB-453 cells treated with JQ1-VHL

first: Ewa

last: Michalak

first: Enid

middle: Y

last: Lam

first: Sarah-Jane

middle: A


title: Bacterial skin infection primes bone marrow eosinophils to promote allergic skin-lung crosstalk

status: Public on Feb 12 2025

city: Salzburg

first: Natalia

middle: Rodrigues Nunes

last: da Silva

address: Steingasse 11,

institute: Paris Lodron Universität Salzburg

summary: Microbial exposure at barrier interfaces drives development and balance of the immune system, but the consequences of local infections for systemic immunity and secondary inflammation are unclear. Here, we show that dermal exposure to the bacterium Staphylococcus aureus persistently shapes the immune system of mice with specific impact on progenitor and mature bone marrow neutrophil and eosinophil populations. The infection-imposed changes in eosinophils were long-lasting with functional changes associated with epigenetic and metabolic reprogramming, indicative of trained immunity. Bacterial exposure enhanced dermal allergic sensitization and resulted in exacerbated allergen-induced lung inflammation. S. aureus-mediated reprogramming of bone marrow eosinophils was driven by the alarmin interleukin 33 and the complement cleavage fragment C5a. Our study highlights the systemic impact of skin inflammation and reveals eosinophil progenitor training and organ-crosstalk mechanisms that modulate systemic responses to allergens.

overall_design: Bone marrow-derived eosinophils were isolated from mice subjected to epicutaneous Staphylococcus aureus skin infection. ATAC-seq analysis was performed to assess changes in chromatin accessibility and epigenetic reprogramming. Differentially accessible regions were associated with immune memory and inflammatory pathways, contributing to the understanding of systemic immune training and eosinophil function.

first: Mariem

last: Radhouani

first: Asma

last: Farhat

first: Anna

last: Hakobyan

first: Sophie

last: Zahalka

first: Lisabeth

last: Pimenov

first: Alina

last: Fokina

first: Anastasiya

last: Hladik

first: Karin

last: Lakovits

first: Jessica

last: Broesamlen

first: Vojtech

last: Dvorak

last: Nunes

first: Andreas

last: Zech

first: Marco

last: Idzko

first: Thomas

last: Krausgruber

first: Jörg

last: Köhl

first: Ozge

last: Uluckan

first: Jiri

last: Kovarik

first: Kai

last: Hoehlig

first: Axel

last: Vater

first: Margret

last: Eckhard

first: Andy

last: Sombke

first: Nikolaus

last: Fortelny

first: Joerg

last: Menche

first: Sylvia

last: Knapp

first: Philipp

last: Starkl


title: Broad rim lesions are a novel pathological / morphological and imaging biomarker for rapid disease progression in Multiple Sclerosis (part 2)

city: Münster

first: Carolin

last: Walter

address: Domagkstraße 9

department: Medical Faculty of the WWU Münster

summary: Current Multiple Sclerosis (MS) treatments reduce relapse activity but have limited impact on disease progression. Clinical trials targeting progression often fail due to insufficient understanding of its underlying mechanisms. This study analyzes a clinically well-characterized MS autopsy cohort comparing selected individuals with opposite disease trajectories of slow versus rapid progression by extensive unbiased histology and spatial transcriptomics. It unveils a novel lesion type marked by an extensive myeloid cell rim with cellular and transcriptional signatures of inflammation, enhanced antigen presentation and lymphocyte activation that is strongly linked to rapid progression. Validation through an independent TSPO-PET study confirms the association between lesions with a broad myeloid cell rim and disease progression. These findings offer crucial insights into the mechanisms behind MS progression, identifying broad rim lesions as a biomarker for rapid disease progression and potentially guiding patient selection for future therapeutic trials targeting CNS intrinsic inflammation.

overall_design: Samples from broad rim lesions, mixed lesions, and active center lesions were investigated using spatial transcriptomics. For each sample, ROIs at the rim and at NAWM were selected and segmented into AOIs using CD68 as morphology marker.

first: Luisa

last: Klotz

first: Joost

last: Smolders

first: Jussi

last: Lehto

first: Markus

last: Matilainen

first: Luzia

last: Buchholz

first: Stefanie

last: Albrecht

first: Aletta M

middle: R

last: van den Bosch

first: Laura

last: Airas

first: Inge

last: Huitinga

first: Tanja

last: Kuhlmann


first: Michael

last: Croft

overall_design: Mice were sensitized by injecting a mixture of 20 μg HDM (Greer Labs, Lenoir, NC) and 2 mg alum intraperitoneally (i.p.), followed by a resting period of 21 days. Sensitized mice were then exposed to 10 μg HDM via the intranasal (i.n.) route for four consecutive days (21 to 24) in a secondary response. Challenged mice were then rested for another period of 4 weeks and re-exposed to 10 μg HDM i.n. for four consecutive days (45 to 48) in a tertiary response. To monitor lung-localized memory T cells, mice were subjected to short-term labeling with APC eF780-labelled anti-CD90.2 (2 μg in 100ul of PBS; clone 30-H12; Invitrogen) injected via the retro-orbital sinus, 10 minutes before tissue collection. Memory T cells present within tissues protected from this antibody staining are referred to as extravascular tissue-resident cells. Lungs were analyzed at day 45 and 47 for bulk sequencing.

first: Gurupreet

middle: S

last: Sethi

first: Ashmitaa

last: Ramamoorthy Premlal


title: mRNA export factors store nascent transcripts within nuclear speckles as an adaptive response to transient global inhibition of transcription.

first: Mark

last: Dawson

overall_design: ChIPseq for Pol II or BRD4 in MDA-MB-453 cells treated with JQ1-VHL

first: Ewa

last: Michalak

first: Enid

last: Lam

first: Sarah-Jane


title: Tracking and mitigating imprint erasure during induction of naïve human pluripotency at single-cell resolution [Bisulfite-Seq]

city: St. Louis

first: Thorold

middle: W.

last: Theunissen

state: MO

address: 4515 McKinley Ave, Room 3313

institute: Washington University in St. Louis

summary: Naïve human pluripotent stem cells (hPSCs) model the pre-implantation epiblast. However, parent-specific epigenetic marks (imprints) are eroded in naïve hPSCs, which represents an important deviation from the epiblast in vivo. To track the dynamics of imprint erasure during naïve resetting in real time, we established a dual-colored fluorescent reporter at both alleles of the imprinted SNRPN locus. During primed-to-naïve resetting, SNRPN expression becomes biallelic in most naïve cells and biallelic SNRPN expression is irreversible upon re-priming. We utilized this live-cell reporter to evaluate chemical and genetic strategies to minimize imprint erasure. Decreasing the level of MEK/ERK inhibition or overexpressing the KRAB zinc finger protein ZFP57 protected a subset of imprints during naïve resetting. Combining these two strategies protected imprint levels to a further extent than either strategy alone. This study offers an experimental tool to track and enhance imprint stability during transitions between human pluripotent states in vitro.

overall_design: WGBS analysis of H9 and H9-SRG primed and naïve hESCs.

last: Fischer

middle: W

first: Sabine

last: Dietmann


overall_design: SLAMseq of MDA-MB-453 cells treated with JQ1, JQ1-VHL, IBET151 or IBET151-VHL

middle: M


title: Treg fitness signatures as a biomarker for disease activity in Juvenile Idiopathic Arthritis

city: London

first: Anne

last: Pesenacker

state: England

address: Pears Building, UCL Institute of Immunity and Transplantation, Rowland Hill Street

department: Div Infection and Immunity

institute: UCL

summary: biomarker discovery pipeline for active and subclinical JIA through 48 gene Treg signature+ nanoString normalised counts from peripheral blood and synovial fluid mononuclear cells and Tregs

overall_design:
a cohort of healthy adult control PBMC (n=20) and Tregs (n=26); pediatric healthy control PBMC (n=5) and Tregs (n=5); JIA SFMC (n=29) and SF Tregs (n=26); active (AJC>=1) JIA PBMC (n=30) and Tregs (n=30); and inactive (AJC=0) PBMC (n=17) and Tregs (n=17), assessed for 48-gene nanoString Treg signature+ for biomarker discovery to identify active JIA
CHARMS,study
JIAP,study

first: Meryl

middle: H

last: Attrill

first: Diana

last: Shinko

first: Telma

last: Martins Viveiros

first: Martina

last: Milighetti

first: Nina

last: de Gruijter

first: Bethany

last: Jebson

first: Melissa

last: Kartawinata

first: Elizabeth

middle: C

last: Rosser

first: Lucy

last: Wedderburn


title: Evidence for a lipofibroblast-to-Cthrc1pos myofibroblast reversible switch during the development and resolution of lung fibrosis

city: bad nauheim

first: Stefan

last: Günther

address: ludwigtr. 43

institute: MPI for heart and lung research

summary: Question addressed in the study: Genetic lineage tracing of alpha smooth muscle actin-positive (ACTA2+) cells in the context of bleomycin-induced pulmonary fibrosis in mice has shown that activated myofibroblasts (aMYFs) differentiate into lipofibroblasts (LIFs) during fibrosis resolution. However, the heterogeneity of the different mesenchymal cell subclusters during fibrosis formation and their contribution to the aMYFs as well as the fate of aMYFs during fibrosis resolution are still unclear.    Materials and methods:  Two-month-old Tg(Acta2-CreERT2); tdTomatoflox mice were used to label Acta2pos cells before (Tam-Bleo condition) or after (Bleo-Tam condition) bleomycin administration. Using scRNA-seq, the origin and fate of Acta2pos cells converging on the aMYF/Cthrc1pos myofibroblast lineage, were analyzed at the peak of fibrosis formation (day 14) as well as during resolution (day 60). Mining of human IPF scRNA-seq data was also carried out. The function of resident mesenchymal cells derived from saline or bleomycin-injured lungs during fibrosis formation or resolution in supporting AT2 progenitor cell renewal was assessed using alveolar organoid assays.    Results:  By employing the dimensionality reduction and UMAP visualization MYF/Cthrc1pos cells were identified and characterized in three scRNA-seq conditions: Saline, Tam-Bleo and Bleo-Tam. Acta2pos cells isolated from saline-treated mice revealed the presence of Acta2pos cells in the expected airway and vascular smooth muscle cells, but also in alveolar fibroblasts, peribronchial and adventitial fibroblasts as well as in a restricted number of Cthrc1pos cells indicating a primed pro-fibrotic status of the non-injured lung. In Tam-Bleo lungs, minimal contribution of these cells to the Cthrc1pos pool was observed. By contrast, labeling of Acta2pos cells following bleo administration (Bleo-Tam) led to massive labeling of Cthrc1pos cells, which following fine clustering were grouped into 4 subclusters named F1-F4. Interestingly, cells in cluster F1 were identified as Cthrc1low and displayed a strong LIF signature (LIFhigh) while cells in cluster F2 were LIFlow and Cthrc1high. Examination of F1 and F2 subclusters during fibrosis resolution suggested that F2 transition back to an F1 status and eventually differentiate into Cthrc1low LIFhigh alveolar fibroblasts. Analysis of the alveolar fibroblast cluster revealed the presence of three subclusters (A, B and C). Cluster C appeared only in Bleo-Tam and displayed a fibrotic signature (Sfrp1, Eln, Ltbp2, Spp1high). These cells, which originate from Acta2neg alveolar fibroblasts are suggested to differentiate into the Cthrc1low LIFhigh F1 subcluster. Data mining of human scRNA-seq from fibrotic lungs supported the conservation of the heterogeneity of the CTHRC1pos population. Alveolosphere assays indicated that resident mesenchymal cells did not reacquire their stem cell niche function at day 60, during fibrosis resolution.    Answer to the question:  Our results support a LIF-to-aMYF reversible switch during fibrosis formation and resolution.

first: Arun

last: Lingampally

first: Saverio

last: Bellusci


title: A transcriptionally active lipid vesicle encloses the injected Chimalliviridae genome in early infection

status: Public on Feb 11 2025

first: Phoolwanti

last: Rani

address: 5401 TATA hall, 9600 Gilman Dr

department: Molecular Biology

institute: University of California, San Diego

summary: Many eukaryotic viruses require membrane-bound compartments for replication, but no such organelles are known to be formed by prokaryotic viruses1–3. Bacteriophages of the Chimalliviridae family sequester their genomes within a phage-generated organelle, the phage nucleus, which is enclosed by a lattice of the viral protein ChmA4–10. Previously, we observed lipid membrane-bound vesicles in cells infected by Chimalliviridae, but due to the paucity of genetics tools for these viruses it was unknown if these vesicles represented unproductive, abortive infections or a bona fide stage in the phage life cycle. Using the recently-developed dRfxCas13d-based knockdown system CRISPRi-ART11 in combination with fluorescence microscopy and cryo-electron tomography, we show that inhibiting phage nucleus formation arrests infections at an early stage in which the injected phage genome is enclosed within a membrane-bound early phage infection (EPI) vesicle. We demonstrate that early phage genes are transcribed by the virion-associated RNA polymerase from the genome within the compartment, making the EPI vesicle the first known example of a lipid membrane-bound organelle that separates transcription from translation in prokaryotes. Further, we show that the phage nucleus is essential for the phage life cycle, with genome replication only beginning after the injected DNA is transferred from the EPI vesicle to the newly assembled phage nucleus. Our results show that Chimalliviridae require two sophisticated subcellular compartments of distinct compositions and functions that facilitate successive stages of the viral life cycle.

overall_design:
Post Knockdown using CRISPR-iART system, the RNA-seq was performed  from biological triplicate samples (ChmAKD_AG1A, ChmAKD_AG1B and ChmAKD_AG1C) and compared with biological triplicate of the non-targating control (NTA, NTB and NTC). These data were also compared with different time point post infection - WT5mpi (5mpiA, 5mpiB and 5mpiC), WT15mpi (15mpiA, 15mpiB), WT20mpi (20mpiA, 20mpiB, 20mpiC), WT30mpi (30mpiA, 30mpiB), and WT60mpi (60mpiA, 60mpiB).
Each GEO sample represents biological replicates that have been merged.

last: Villa


title: Humanized mouse tumor models for evaluation of ILT2 and ILT4 antagonist antibodies

city: South San Francisco

first: Geoff

last: Stone

address: 333 Oyster Point Blvd

institute: NGM Biopharmaceuticals Inc.

summary: Solid tumors are dense three-dimensional (3D) multi-cellular structures that enable efficient receptor–ligand trans interactions via close cell–cell contact.  Immunoglobulin-like transcript (ILT)2 and ILT4 are related immune suppressive receptors that play a role in the inhibition of myeloid cells within the tumor microenvironment.  The relative contributions of ILT2 and ILT4 to immune inhibition in the context of solid tumor tissue has not been fully explored. We present evidence that both ILT2 and ILT4 contribute to myeloid inhibition. We found that while ILT2 inhibits myeloid cell activation in the context of trans-engagement by MHC-I, ILT4 efficiently inhibits myeloid cells in the presence of either cis- or trans-engagement.  In a 3D spheroid tumor model, dual ILT2/ILT4 blockade was required for optimal activation of myeloid cells, including the secretion of CXCL9 and CCL5, upregulation of CD86 on dendritic cells, and downregulation of CD163 on macrophages.  Humanized mouse tumor models showed increased immune activation and cytolytic T cell activity with combined ILT2 and ILT4 blockade, including evidence of the generation of immune niches, which have been shown to correlate with clinical response to immune checkpoint blockade. In a human tumor explant histoculture system, dual ILT2/ILT4 blockade increased CXCL9 secretion, downregulated CD163 expression, and increased the expression of M1 macrophage, IFN-γ, and cytolytic T cell gene signatures. Thus, we have revealed distinct contributions of ILT2 and ILT4 to myeloid cell biology and provide proof-of-concept data supporting the combined blockade of ILT2 and ILT4 to therapeutically induce optimal myeloid cell reprogramming in the tumor microenvironment.

overall_design: Tumors were dissected and dissociated by mincing with a razor blade, followed by grinding through a 70 μm filter cup (Corning) to generate a single cell suspension. Dissociated tumor cells were then washed, counted and human CD45+ cells isolated using the Human CD45 Positive Selection Kit for humanized mice (Stemcell Technologies), according to the manufacturer’s instructions. Following four rounds of washing and magnetic bead isolation, the non-dissociated bead/cell mixture was resuspended in 100 μl of RNAlater (Invitrogen) and stored at -80 °C. To isolate RNA, wells were thawed, cells were pelleted and pellets lysed in 100 μl of RLT buffer, followed by centrifugation to removecell debris and magnetic beads. Supernatant was processed to isolate RNA with the RNEasy mini kit following manufacturer's instructions (Qiagen). Total RNA was prepared for sequencing using the Illumina stranded mRNA library prep kit and sequenced on an Illumina NovaSeq S2 (100bp single-end reads and an average of 40 million reads were generated for each sample). For sequencing alignment, RNA-seq data raw reads were filtered using Trim Galore! (http://www.bioinformatics.babraham.ac.uk/projects/trim_galore/) to remove low quality and adaptor bases. Reads shorter than 20nt were discarded. Filtered reads were mapped to UCSC hg19 and mm9 genome sequences using STAR (v2.7.6a). In the next step, XenofilteR (v1.6) was used on the bam files to exclude mouse RNA sequence reads that originated from host mice. At last, counts of all samples aligned to the human genome were generated using featureCounts (v2.0.1), and edgeR (v3.40.1) was used to get normalized counts and perform differential gene expression analysis.

first: Jane

last: Tian

first: Amir

last: Ashique

first: Sabrina

last: Weeks

first: Tian

last: Lan

first: Yang

last: Hong

first: Hung-I

last: Chen

first: Christina

last: Song

first: Kikuye

last: Koyano

first: Kalyani

last: Mondal

first: Daniel

last: Tsai

first: Isla

last: Cheung

first: Mehrdad

last: Moshrefi

first: Avantika

last: Kikatpure

first: Bin

last: Fan

first: Betty

last: Li

first: Samir

last: Qurashi

first: Lauren

last: Rocha

first: Jonathan

last: Aguayo

first: Col

last: Rodgers

first: Marchelle

last: Meza

first: Darren

last: Heeke

first: Sara

last: Medfisch

first: Chun

last: Chu

first: Shelley

last: Starck

first: Nandini

last: Pal Basak

first: Satish

last: Sankaran

first: Mohit

last: Malhotra

first: Suzanne

last: Crawley

first: Thomas-Toan

last: Tran

first: Dana

last: Duey

first: Carmence

last: Ho

first: Igor

last: Mikaelian

first: Wenhui

last: Liu

first: Lee

last: Rivera

first: Jiawei

last: Huang

first: Kevin

last: Paavola

first: Kyle

last: O'Hollaren

first: Lisa

middle: K

last: Blum

first: Vicky

last: Lin

first: Peirong

first: Anjushree

last: Iyer

first: Sisi

last: He


title: A Massively Parallel CRISPR-Based Screening Platform for Modifiers of Neuronal Activity

city: Palo Alto

first: Steven

last: Boggess

state: California

address: 3181 Porter Dr

institute: Arc Institute

summary: Understanding the complex interplay between gene expression and neuronal activity is crucial for unraveling the molecular mechanisms underlying cognitive function and neurological disorders. Here, we developed pooled screens for neuronal activity, using CRISPR interference (CRISPRi) and the fluorescent calcium integrator CaMPARI2. Using this screening method, we evaluated 1343 genes for their effect on excitability in human iPSC-derived neurons, revealing potential links to neurodegenerative and neurodevelopmental disorders. These genes include known regulators of neuronal excitability, such as TARPs and ion channels, as well as genes associated with autism spectrum disorder and Alzheimer's disease not previously described to affect neuronal excitability. This CRISPRi-based screening platform offers a versatile tool to uncover molecular mechanisms controlling neuronal activity in health and disease.

overall_design: Human iPSCs were transduced with pooled sgRNA library lentivirus at a low MOI (10% transduction efficiency) to minimize multiple infections in which a cell receives more than one sgRNA. Then, iPSCs were selected with puromycin, differentiated into neurons via induction of NGN2 expression, and grown 21 days. Neurons were then dissociated with papain and 20k cells were loaded into a single well of a 10X Chromium GEM-X 3’ chip for GEM preparation. Single cell library preparation was conducted according to the protocol provided for Chromium GEM-X Single Cell 3’ Reagent Kit v4. Enrichment of sgRNA sequences was performed with three semi-nested PCR reactions.

first: Vaidehi

last: Gandhi

first: Ming-Chi

last: Tsai

first: Emily

last: Marzette

first: Noam

last: Teyssier

first: Yu-Ying

last: Chou

first: Xiaoyu

last: Hu

first: Amber

last: Cramer

first: Lin

last: Yadanar

first: Kunal

last: Shroff

first: Claire

middle: G

last: Jeong

first: Celine

last: Eidenschenk

first: Jesse

middle: E

last: Hanson

first: Ruilin

last: Tian

first: Martin

last: Kampmann


title: Memory CD4+ T cells sequentially restructure their 3D genome during stepwise activation

first: Jose

middle: Ignacio

last: de las Heras

address: Max Born Crescent

department: Wellcome Trust Centre

summary: Obtaining a complete picture of 3D-genome organization in tissues and disease states is often blocked by limiting amounts of tissue samples such that multiple types of genome-wide analyses cannot be performed on the same sample. Here, we found that starting with a pure population of memory CD4+ T-cells (mCD4-cells) we could get reproducible Hi-C data with just 50,000 cells. This population is only ~18.75% of the whole T-cell population, but presumably its uniformity enabled high reproducibility with low cell numbers so that Hi-C, RNA-Seq and potentially other analyses could be performed with repeats from the same donor. Moreover, there was strong correspondence between data from 3 different donors, suggesting that variability between individuals observed in other reports may reflect more the dynamic variability in the status of the immune system between individuals rather than variability in genome organization between individuals for a given cell type. We further observed uniform changes between samples and blood donors occurring during activation of the memory mCD4+ T cell population. While some genome organization changes occurred concomitant with changes in gene expression, there were at least as many changes that occurred without corresponding changes in expression. Counter to the hypothesis that TADs are largely invariant structures providing a scaffold for dynamic looping contacts between enhancers and promotors, we found that there were at least as many dynamic TAD changes as loops in this highly differentiated cell type. Stimulation with IL-2 alone triggered many changes in genome organization and many of these changes were strengthened by additional TCR and CD28 co-receptor stimulation. This suggests a stepwise process whereby mCD4+ T cells undergo sequential genome organization changes induced by distinct or combined stimuli likely to "prime" or “de-prime” them for expression responses to subsequent TCR-antigen ligation or additional cytokine stimulation

overall_design: Primary CD4+ memory T-cells were isolated from three blood donors and purified via FACS. Cells were treated with either IL-2, IL2+TCR or left untreated. HiC experiments were performed on 'donor1' in duplicate using either 50000, 250000 or 1000000 cells in order to compare the reliability of the method using reduced numbers of input cells. For donors 2 and 3, 250000 cells were employed in single experiments.

first: Alex

middle: I

last: Ward

first: Eric

last: Schirmer

first: Ariberto

last: Fassati


title: Baseline Gene Expression in Human Derived Cell Lines

city: RTP

first: Logan

middle: J

last: Everett

state: North Carolina

address: 109 T.W. Alexander Dr

department: Office of Research and Development

institute: U.S. Environmental Protection Agency

summary: Many research organizations are exploring the use of high-throughput profiling (HTP) assays such as transcriptomics and image-based phenotypic profiling for in vitro bioactivity screening and hazard evaluation of chemicals. HTP assays can be used to characterize the response of many human-derived cell types to chemicals. However, no cell-based in vitro model will express all the potential molecular targets and pathways that are present in the human body that could be perturbed, resulting in toxicity. Therefore, an envisioned approach for the use of HTP assays in chemical bioactivity screening involves panels of cell lines that express complementary (partially non-overlapping) sets of molecular targets. In this study, baseline gene expression in 53 different human-derived cell lines was characterized using the TempO-Seq human whole transcriptome assay.  The cell line set included 36 human cancer-derived cell lines and 17 engineered or spontanously immortalized human cell lines. The baseline gene expression data can inform what set(s) of molecular targets are expressed across the cell line collection and inform researchers regarding the choice of cell lines for chemical screening studies.

overall_design: Baseline gene expression profiling of 53 different human-derived cell lines. The cell lines were cultured in growth medium recommended by the commercial suppliers from which the cell lines were obtained.

first: Joshua

last: Harrill

first: Clinton

last: Willis

first: Nisha

last: Sipes

first: Laura

last: Word

first: Joseph

last: Bundy

first: Derik

last: Haggard

first: Russell

last: Thomas

first: R Woodrow

last: Setzer


title: Epigenetic editing at individual age-associated CpGs affects the genome-wide epigenetic aging landscape [MSC_hyperL_D11]

city: Aachen

first: Wolfgang

last: Wagner

address: Pauwelsstrasse 20

department: Helmholtz Institute for Biomedical Engineering

institute: RWTH Aachen University

type: Methylation profiling by genome tiling array

summary: Aging is reflected by genome-wide DNA methylation changes, but it is largely unclear how these epigenetic modifications are regulated. In this study, we explored the possibility to interfere with epigenetic clocks by epigenetic editing at individual CpG sites. CRISPR-guided approaches (dCas9-DNMT3A and CRISPRoff) facilitated targeted methylation at age-associated genomic regions that remained stable for up to three months. Furthermore, epigenetic editing evoked many genome-wide off-target effects, which were highly reproducible and enriched at other age-associated CpGs – thus, they are not random off-target effects but seem to resemble co-regulated epigenetic bystander modifications. 4C chromatin conformation analysis at age-associated sites revealed increased interactions with bystander modifications and other age-associated CpG sites. Subsequently, we multiplexed epigenetic modifications in HEK293T, mesenchymal stromal cells, and T cells at five genomic regions that become either hypermethylated or hypomethylated upon aging. While epigenetic editing at age-hypomethylated CpGs appeared less stable, it also resulted preferentially in bystander modifications at CpGs with highest correlation with chronological age. Notably, modifications were simultaneously observed at CpGs that gain and lose methylation with age. These results demonstrate that targeted epigenetic editing can modulate the epigenetic aging network in its entirety and thereby interfere with epigenetic clocks.

overall_design: We analyzed genome-wide DNA-methylation changes with targeted epigenetic editing. Human mesenchymal stromal cells were electroporated with gRNA and dCAS9-DNMT3A/3L. Controls were transfected with plasmides expressing scramble guide RNAs and dCAS9-DNMT3A/3L. Timepoint of isolation is denoted in "time point".

first: Sven

last: Liesenfelder

first: Mohamed

last: Elsafi Mabrouk

first: Jessica

last: Iliescu

first: Monica

last: Varona Baranda

first: Athanasia

last: Mizi

first: Juan

middle: F

last: Perez-Correa

first: Martina

last: Wessiepe

first: Argyris

last: Papantonis