title: Cell Type-Specific Hormonal Signaling Configures Hypothalamic Circuits for Parenting

city: Cambridge

first: Brandon

last: Logeman

address: 16 Divinity

institute: Harvard University

summary: Parenting behavior emerges from hormonally sensitive circuits, but how distinct circuit components are affected by, and contribute to, sex and state dependent changes in infant caregiving remains unclear. Using cell type-specific approaches, we characterized two circuit nodes that are differentially configured by hormones to modulate infant-evoked behavior. An excitatory neuronal population in the anteroventral periventricular nucleus is only active in lactating mothers, increases virgin female caregiving when artificially stimulated and confers oxytocin sensitivity in mothers through a prolactin--STAT5b pathway. These neurons function upstream of another preoptic area population involved in male and female parenting, thus boosting caregiving by mothers. By contrast, androgen signaling in the latter preoptic population reshapes their intrinsic properties to promote pup-directed aggression, revealing cell type-specific tuning of social behavior circuits.

overall_design: RNA-seq and ATAC-seq of hypothalamic neurons enriched via FANS from multiple physiological states including mothers, fathers, virgin males, and virgin females. Please note that processed data files generated from both *L001 and L002 samples are linked to the corresponding *L001 sample records.

first: Patricia

last: Horvath

first: Mustafa

last: Taly

first: Vikrant

last: Kapoor

first: Harris

last: Kaplan

first: Catherine

last: Dulac


title: Receptor Tyrosine Kinases phosphorylate RNA polymerase II at signal-responsive genes

status: Public on Jan 13 2026

city: Memphis

first: Wojciech

last: Rosikiewicz

state: TN

address: 262 Danny Thomas Place

department: Center for Applied Bioinformatics

country: USA

institute: St. Jude Childrens Research Hospital

type: Genome binding/occupancy profiling by high throughput sequencing

type: Expression profiling by high throughput sequencing

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

overall_design: Refer to individual Series


title: Cohesin-mediated chromatin remodeling controls the differentiation and function of conventional dendritic cells.

city: New York City

first: Eduardo

last: Esteva

state: New York

address: 550 1st Ave

institute: NYU Grossman School of Medicine


title: Uncovering a Key Metabolic Role of the GATA1 N-Terminus in Red Blood Cell Development

status: Public on Jan 12 2026

first: Te

last: Ling

address: 262 Danny Thomas Pl

institute: St. Jude Children's Research Hospital

summary: Mutations in GATA1 that result in skipping of exon 2, which encodes the N-terminus, are associated with the myeloid leukemia of Down syndrome and Diamond-Blackfan anemia (DBA). To elucidate the molecular functions of the N-terminus, we employed single-cell RNA sequencing (scRNA-seq) of fetal liver cells derived from Gata1 mutant embryos that express the GATA1 short (GATA1s) isoform in place of full-length GATA1 (GATA1FL). scRNA-seq highlighted defects in erythropoiesis and revealed that the absence of the N-terminus resulted in elevated expression of genes involved in glycolysis, such as the rate-limiting pyruvate kinase PKM. To elucidate the regulation of the PKM gene, we performed precision nuclear run-on sequencing and cleavage under targets and release using nuclease on erythroid cells following acute deletion of GATA1 and determined that PKM is a direct target of GATA1. Mechanistically, we found that substitution of GATA1FL with GATA1s led to increased glycolysis in erythroid progenitor cells but did not affect oxidative phosphorylation. We further discovered that the expression of PKM is significantly elevated in DBA patients with RPS19 mutations, consistent with a role for GATA1 regulation of glycolysis in erythropoiesis. Together, these findings reveal that GATA1 controls not just heme metabolism, but also glycolysis.

overall_design: HUDEP2 cells expressing FKBP12F36V-GATA1 (two clones 6F8 and 7C4) were cultured under differentiation conditions for 24 hours. PRO-seq was conducted at 0.5, 3, and 6 hours in the presence of the acute degradation inducer dTAG-13 (100 nM) to monitor the nascent transcripts.

first: Rashid

last: Mehmood

first: John

middle: D

last: Crispino


title: The accumulation of methylglyoxal and acrolein leads to arginine depletion causing hyperglycemia and renal abnormalities

city: Mannheim

first: Shu

last: Li

address: Neckarpromenade

country: Germany

institute: ECAS

summary: There is growing evidence to support the idea that the accumulation of reactive carbonyl species plays a significant causal role in the pathogenesis of ageing, cardiovascular diseases, diabetes and its complications. Among reactive carbonyl species, methylglyoxal and acrolein, detoxified by Glyoxalase 1 and Aldo-keto reductase 1A1A, could modify arginine residues to form the advanced glycation end products and acrolein-derived guanidino adducts that alter protein structure and function. However, it remained unclear whether the combined loss of Glyoxalase 1 and AKR1A1A impairs arginine metabolism and exacerbates hyperglycemia and its complications. In this study, we generated Glyoxalase 1 and Aldo-keto reductase 1A1A double knockout zebrafish to explore the interplay between carbonyl detoxification and arginine metabolism. Loss of both enzymes led to accumulation of methylglyoxal and acrolein, suppression of the arginine metabolic pathway, and downregulation of insulin signaling. Double mutants exhibited elevated whole-body glucose in larvae and postprandial hyperglycemia in adults, accompanied by glomerular basement membrane thickening, while retinal vasculature remained unaffected. Remarkably, arginine supplementation restored Akt phosphorylation, improved insulin sensitivity, and alleviated renal pathology. Together, these findings identify Glyoxalase 1 and Aldo-keto reductase 1A1A as cooperative regulators of carbonyl detoxification and metabolic homeostasis. Disruption of this dual detoxification system links carbonyl stress to impaired insulin signaling and organ-specific damage through dysregulated arginine metabolism. These results uncover a carbonyl–arginine metabolic axis through which reactive carbonyl species, shaped by the synergistic activity of GLO1 and AKR1A1A, drive glucose dysregulation and tissue-specific injury, highlighting potential targets for metabolic intervention in diabetes and metabolic diseases.

overall_design: glo1-/-akr1a1a-/-zebrafish were generated using CRISPR/Cas9 technology.  Multiple experiments are performed regarding vasculature and organ alterations, glucose homeostasis, transcriptome, and metabolomics in Tg(fli1:EGFP) zebrafish.

first: Hao

first: Jens

last: Kroll


title: Scalable single-cell total RNA sequencing unifies coding and non-coding transcriptomics

city: Stanford

first: Alina

last: Isakova

state: CA

address: 290 Jane Stanford Way

institute: Stanford University

summary: Current single-cell RNA atlases largely capture polyadenylated transcripts while missing critical regulatory layers from non-coding RNA. To address this, we developed TotalX - a generalizable framework that adapts Smart-seq total RNA profiling for use in droplet-based platforms, and captures a broad complement of coding and non-coding RNAsusing a unified pipeline. Applying this approach to developing human brain, we generate a dataset mapping diverse RNA biotypes across all neuronal and non-neuronal lineages, revealingbiotype-specific expression programs with cell-type and temporal specificity. Tracking miRNA dynamics in Cajal–Retzius neurons, transient and early-born neurons in the cortex, we show the enrichment and target anti-correlation of MIR137, associated with schizophrenia and intellectual disability, suggesting tight regulatory control. We apply TotalX to human peripheral blood mononuclear cells and identify transcriptional modules combining coding and non-coding RNAs and tRNA dynamics. Additionally, we analyze dengue-infected hepatocytes and capture non-adenylated viral transcripts that distinguish infection states. This expanded coverage helps with understanding cellular identity and gene regulation at atlas scale

overall_design: HEK293T cells and DENV2 infected Huh7 cells were processed using TotalX method. We also compared TotalX implementations without as well as with two rounds of DASH in miRNA(+) configurations on HEK293T cells


title: Uncovering a Key Metabolic Role of the GATA1 N-Terminus in Red Blood Cell Development [CUT&RUN]

overall_design: CD71hiTer119hi erythroblasts were isolated from E13.5 embryos. Occupancy profiling was performed using the CUT&RUN method.


title: Lycorine alleviates intrahepatic cholestasis by reducing the accumulation of bile acids via TKT/ATF3 pathway

city: nanjing

first: Ying

last: Zheng

address: 639 Longmian Avenue, Jiangning District

country: China

institute: China pharmaceutical university

summary: Cholestatic liver disease (CLD) is characterized by the dysfunction of bile synthesis and excretion, leading to progressive liver injury or fibrosis, but lacks effective treatment drugs. Lycorine, an alkaloid isolated from Lycoris bulbs, possesses antiviral, anti-inflammatory, analgesic, cardioprotective activities. However, there has been no report concerning its effects on CLD so far. We evaluated the hepatoprotective effect of lycorine on intrahepatic cholestasis mouse models induced by α-naphthylisothiocyanate (ANIT) or after multidrug resistance protein 2 knockout (Mdr2-/-). RNA-Sequencing (RNA-Seq), cellular thermal shift assay (CETSA) and mass spectrometry analysis were used to investigate the regulated molecular mechanism of lycorine. Lycorine alleviated intrahepatic cholestasis by decreasing the expression of activating transcription factor 3 (ATF3), and inhibiting the nuclear accumulation of unphosphorylated ATF3, a transcription repression factor of the bile salt export pump (BSEP) responsible for bile acid (BA) secretion. Additionally, lycorine bound with transketolase (TKT) protein directly and promoted its ubiquitination degradation, disrupting the interaction between TKT and ATF3, resulting in the increased interaction between Rho-associated protein kinase 1(ROCK1) and ATF3 and subsequently BSEP transcription repression. Conclusively, our results shed light on the protective effects and mechanisms of lycorine against CLD, which pave way for the design of novel clinical therapeutic strategies.

overall_design: To investigate how lycorine regulates intrahepatic cholestasis, we conducted RNA sequencing (RNA-seq) on the Mdr2 knockout mouse livers treated with vehicle or lycorine.

first: Jiaman

last: Liu

first: YingRong

last: Leng

first: ShengTao

last: Ye

first: Yang

last: Cheng

first: YanQiu

last: Zhang

first: LingYi

last: Kong


title: Effect of increasing dietary lysine level on transcriptomic profile in pig longissimus dorsi muscle following a period with dietary protein restriction after weaning

city: Frederiksberg C

first: Andrew

last: Williams

address: Dyrlægevej 100

country: Denmark

institute: University of Copenhagen

summary: Thus, in the present study the objective was to determine whether increasing dietary Lys levels after a period of dietary protein restriction would lead to distinct muscle transcriptomic profiles in weaner pigs. It was hypothesized that increased dietary Lys levels would lead to increased expression and enrichment of, respectively, genes and pathways regulating muscle protein synthesis.

overall_design: Sixty-seven Swiss Large White piglets from 20 litters of one farrowing series were used in the experiment, where 33 were females and 34 were male castrates. Of the 67 piglets 60 piglets were randomly pre-allocated on the day of weaning to one of 4 dietary treatments implemented from week five to eight (W5-8) after weaning. Remaining seven piglets were not further used in the present study. From weaning and until four weeks after weaning (W1-4), pigs were divided into two groups, a control (STD, N = 18) receiving a diet with standard CP content and a group receiving a diet with a low protein CP content (LP, N = 42). The LP diet was 81/79% (total/digestible) of the CP in the STD diet to force a reduced protein intake during the first four weeks after weaning, while maintaining digestible AA at requirement level in both diets (Table 1). Pigs had ad libitum access to feed. In the period W5-8 the four dietary treatments were a control (STD-STD, N = 12), standard (LP-STD, N = 12), STD + 10% Lys (LP-STD10+, N = 12) and STD + 20% Lys (LP-STD20+, N = 12), where STD-STD and LP-STD treatments received the standard diet, and Lys10 and Lys20, respectively, STD with 10% and 20% greater digestible Lys content and remaining digestible essential amino acids (EAA) above requirement level (Table 1). Pigs in the STD-STD group only included pigs from the STD group from W1-4, whereas the LP-STD, LP-STD10+ and LP-STD20+ groups only included pigs from the LP group of W1-4. Tissue samples from the longissimus dorsi (LD) muscle were collected, snap frozen in liquid nitrogen and subsequently stored in -80 °C until further analysis.

middle: R


title: Genetic determinants that permit growth without the core septin Cdc12 in Cryptococcus neoformans

city: Clemson

first: Rooksana

middle: Elizabeth

last: Noorai

state: South Carolina

address: 105 Collings St, BRC 157

department: Clemson University Genomics and Bioinformatics Facility

institute: Clemson University

summary: Septins are conserved cytoskeletal GTPases that regulate morphogenesis, cytokinesis, and cell-surface organization in fungi. In most model and pathogenic yeasts, including Saccharomyces cerevisiae and Candida albicans, core septin subunits such as Cdc3 and Cdc12 are essential and their loss results in lethal cytokinetic defects. In Cryptococcus neoformans, by contrast, deletion of CDC12 yields viable cells that grow normally under permissive conditions, raising the question of how this pathogen maintains cell integrity without a functional septin complex. To address this, we compared the transcriptomes of wild-type cells and a cdc12Δ mutant. The cdc12Δ strain showed a focused set of transcriptional changes, with induction of genes involved in cell wall remodeling, stress-dependent morphogenetic programs, and pathways that promote fitness under compromised cytoskeletal conditions. Genetic analysis of the most highly induced genes uncovered factors that contribute to proliferation in the absence of Cdc12, including a previously uncharacterized aspartic-type endopeptidase that becomes essential when septin function is lost. These findings outline the compensatory circuitry that permits septin-independent growth in C. neoformans and help explain why this pathogen can tolerate septin loss. This intrinsic plasticity highlights C. neoformans as a powerful comparative model for studying the evolution and function of septin complex in fungal cell biology.

overall_design: Changes in mRNA abundance were identified by comparing 4 replicates of wild type Cryptococcus neoformans samples at room temperature, 21C, to 4 replicates of Septin 12 mutant strain C. neoformans also at room temperature, 21C.

first: Piotr

last: Stempinski

middle: E

first: Vijay

last: Shankar

first: Lukasz

last: Kozubowski


title: Uncovering a Key Metabolic Role of the GATA1 N-Terminus in Red Blood Cell Development [CUT&RUN Human]

first: Te

last: Ling

overall_design: HUDEP2 cells expressing FKBP12F36V-GATA1 were cultured under differentiation conditions for 24 hours. GATA1 occupancy profiling was conducted at 0.5, 3, and 6 hours in the presence of the acute degradation inducer dTAG-13 (100 nM) using the CUT&RUN method.

first: Rashid

last: Mehmood

first: John


title: CTCF ChIP-seq in AC16 cell: A Conserved Chromatin Structural Program Underlies Injury-Induced Cardiomyocyte Proliferation

city: Birmingham

first: Thanh

middle: Minh

last: Nguyen

state: AL

address: 1670 University Blvd, Volker Hall G094

department: Department of Biomedical Engineering

institute: University of Alabama at Birmingham

summary: Because adult mammalian cardiomyocytes cannot reactivate their proliferative machinery, the heart does not recover effectively after injury. In contrast, during the early neonatal period, cardiomyocytes retain a transient regenerative capacity. Myocardial injury performed before postnatal day 3 triggers robust cardiomyocyte proliferation and complete recovery within weeks, and apical resection during the first postnatal days leads to full regeneration without scar formation. In pigs, apical resection at day 1 followed by myocardial infarction at day 28 induces strong cardiomyocyte proliferation and complete functional recovery by day 56. To investigate the molecular programs that underlie this transient regenerative competence, we reanalyzed mouse and pig snRNA-seq datasets using a cell cycle–specific autoencoder (CSA) and a chromatin structural factor–specific autoencoder (CSF). Both frameworks consistently identified a distinct cardiomyocyte population (CM1) enriched for mitotic markers after injury. Gene Ontology analysis revealed coordinated activation of cell-cycle pathways together with chromatin organization and remodeling processes, suggesting that cell-cycle re-entry requires extensive epigenetic restructuring. Cross-species comparison identified a hallmark of chromatin structural regulators associated to cardiomytocyte proliferation. Among these factors, CTCF emerged as a central regulator. Its expression declined after birth but was robustly reactivated in injured mouse and pig hearts. Functional assays demonstrated that CTCF overexpression promotes cardiomyocyte-like cell proliferation, and induce targeted redistribution of CTCF binding at key cell-cycle regulatory loci. Together, these findings identify conserved modulation of chromatin structural factors as a key mechanism underlying cardiomyocyte cell-cycle reactivation and position CTCF as a prominent regulator within this architectural program during cardiac regeneration.

overall_design: Plamids containing seqences to overexpress CTCF (CTCF-OE) and empty (control) were transfected into AC16 cell lines. The cells were harvested three days after transfection. Three biological replicates of approximately 1×10⁶ AC16 cardiomyocyte-like cells transfected with either empty pcDNA vector or pcDNA-CTCF were crosslinked with disuccinimidyl glutarate (DSG; 30 min at room temperature), followed by fixation with 1% formaldehyde for 10 min at room temperature and quenching with 125 mM glycine for 5 minutes. Cells were lysed, and chromatin was sonicated for 60 min using the Pixul platform (Active Motif), under the following parameters: Pulse (N):50, PRF (kHz): 1.00, Process Time: 60 minutes, Burst Rate (Hz): 20. Chromatin was incubated overnight at 4 °C with Protein G/A Dynabeads (Invitrogen, #10003D) pre-bound to 1.5 µg of anti-CTCF antibody (Active Motifs, #91285). Bead-bound complexes were extensively washed and resuspended in TT buffer (10 mM Tris-HCl, pH 8.0, 0.05% Tween-20). On-bead library preparation was performed using the NEBNext Ultra II DNA Library Prep Kit (NEB, #E7645L). Libraries were barcoded using NEXTFLEX Unique Dual Index Barcodes, PCR-amplified for 14 cycles, with Solexa 1GA/1GB primers, and size-selected (200–500 bp) prior to sequencing on an Illumina NovaSeq SP100 platform.For processing, the CTCF ChIP-seq data were aligned via Bowtie genome with Human / GRCh38 no-alt analysis reference genome (https://benlangmead.github.io/aws-indexes/bowtie). Peak calling was done via Macs2 program (https://pypi.org/project/MACS2/).

first: Geng

last: Xiaoxiao

first: Gámez-Luque

last: Javier

first: Porter

last: Riley

first: Eduardo

last: Useche-Bernal

first: Zhang

last: Jianyi

first: Cobo

last: Isidoro

first: Nguyen

last: Thanh

first: Rosa-Garrido

last: Manuel


title: An α-specific PI3K inhibitor improves chemotherapy efficacy by inhibiting hepatic stellate cell activation in liver cancer

city: Woolloongabba

first: Qi

last: Ruan

state: QLD

address: 37 Kent Street

department: Frazer Institute

country: Australia

institute: The University of Queensland

summary: The poor response to current systemic treatments in liver cancer has been associated with the activation of cancer-associated fibroblasts (CAFs). Herein, we aimed to investigate the effects of chemotherapy on hepatic stellate cells (HSCs), the main origin of CAFs in liver cancer. In vitro experiments demonstrated that cisplatin activated HSCs through a paracrine effect from HCC tumour cells while the mechanism remains unknown. RNA sequencing of human primary HSCs incubated in untreated or cisplatin-pretreated Huh7 conditioned medium revealed a PI3K signaling pathwayof HSC activation. Further analyses showed that PI3K pathway, especially PI3K p110a is crucial for HSC activation and by targeting PI3K p110a, HSC activation induced by cisplatin can be completely inhibited and improves cisplatin therapeutic efficacy.

overall_design: RNA-seq profiling of human primary hepatic stellate cells incubated in untreated or cisplatin-pretreated Huh7 cell conditioned medium for 24 hours.

first: Xiaowen

last: Liang


title: Characterization of Subchronic Lung and Brain Consequences Caused by Mouse-Adapted SARS-CoV-2 and Influenza A Infection of C57BL6 mice

city: New Orleans

first: Xuebin

last: Qin

state: LA

address: 1430 Tulane Avenue

department: Pathology

institute: Tulane University

summary: SARS-CoV-2 and, to a lesser extent, influenza A can lead to long-term complications in the respiratory and nervous systems. However, the mechanisms driving post-viral sequelae remain poorly understood. To address this gap, we longitudinally characterized C57BL/6 mice infected with sublethal doses of mouse-adapted SARS-CoV-2 (MA30) or influenza A (PR8). Lung and brain tissues were analyzed at 14-, 21-, and 28-days post-infection (DPI) using histological analysis and bulk-RNA sequencing. In the lungs, both infections caused prolonged inflammation and fibrosis. MA30-infected lungs showed persistent upregulation of inflammatory, coagulation, complement, as well as fibrotic, and extracellular matrix (ECM) remodeling pathways at 21 DPI, alongside downregulation of epithelial junction and metabolic program pathways. In contrast, PR8-infected lungs exhibited a strong acute interferon response and chronic upregulation of basal epithelial markers (e.g., Krt5, Krt14), consistent with epithelial regeneration. Notably, only PR8-infected mice displayed KRT5+ progenitor cell migration into damaged lung regions, indicating divergence in repair mechanisms. Neither MA30-infected, nor PR8-infected mice had detectable brain infection. However, MA30 mice, but not PR8-infected mice exhibited an elevated frequency of microhemorrhages at early timepoints and marked neuroinflammation at all timepoints. Transcriptomic profiling of MA30-infected brains showed enrichment for up-regulation of ECM remodeling, vascular dysfunction, IL6-signaling pathways along with a virus-specific disruption of the hypothalamic–pituitary axis with MA30 infection not seen in PR8-infected brains. These included genes linked to neuroinflammation, sensory processing disruption, and microvascular injury, mirroring clinical features of Long COVID. Together, these findings establish distinct tissue-specific trajectories of long-term pathology following SARS-CoV-2 and influenza infection and provide a foundation for dissecting the mechanisms of post-viral lung and brain disease.

overall_design: Comparative gene expression profiling analysis of RNA-seq data of lung and brain tissues from noninfected, MA30-infected, and PR8-infected B6 mice at acute (7, and 8-9 DPI for MA30 and PR8 infection, respectively) and subchronic phase (21 DPI for both MA30 and PR8 infection).

first: Joshua

last: Currey

first: Chenxiao

last: Wang

first: Meredith

middle: G

last: Mayer

first: Yilin

last: Chen

first: Ana Karina

last: Nisperuza Vidal

first: Michaela

middle: J

last: Allen

first: Mst Shamima

last: Khatun

first: Calder

last: Ellsworth

first: Mohammad

last: Islamuddin

first: Jefferson

last: Evangelista

first: Skye

middle: M

last: Minor

first: Nadia

middle: A

last: Golden

first: Kevin

last: Zwezdaryk

first: Nicholas

last: Maness

first: Robert

last: Blair

first: Jay

middle: K

last: Kolls

first: Derek

last: Pociask

first: Tracy

last: Fischer


title: Receptor Tyrosine Kinases phosphorylate RNA polymerase II at signal-responsive genes [ChIP-seq]

first: Wojciech

last: Rosikiewicz

summary: Receptor Tyrosine Kinases phosphorylate RNA polymerase II and associate with distinct set of signal-responsive genes at different genomic loci.

overall_design: Examination of the targets of EGFR and phospho-Tyr1 RNA Pol II phosphorylated CTD after EGF stimulation, with or without treatment with Erlotinib.

first: Preeti

last: Dabas

first: Beisi

last: Xu

first: Aseem

middle: Z

last: Ansari


title: RNA-seq profiling of METTL1 knockdown in human intestinal epithelial FHC cells

city: Changsha

first: Lichao

last: Yang

state: Hunan

address: renmin middle road

institute: The Second Xiangya Hospital, Central South University

summary: METTL1 is a key methyltransferase responsible for N7-methylguanosine (m7G) modification of RNA and has been implicated in multiple cellular processes. However, its role in human intestinal epithelial cells remains incompletely understood.               In this study, we performed RNA sequencing (RNA-seq) to characterize transcriptomic changes induced by METTL1 knockdown in human fetal intestinal epithelial (FHC) cells. Stable METTL1 knockdown was achieved using lentiviral short hairpin RNA (shRNA), while control cells were infected with a non-targeting control shRNA lentivirus (sh-Ctrl). Knockdown efficiency was validated prior to sequencing.                                       Total RNA was extracted from METTL1-knockdown and sh-Ctrl FHC cells, with three independent biological replicates per group. This dataset provides a transcriptomic resource for investigating the regulatory role of METTL1 in intestinal epithelial cells.

overall_design: Human fetal intestinal epithelial FHC cells were divided into two groups: METTL1 knockdown and shRNA control. Lentiviral shRNA targeting METTL1 (sh-METTL1) was used to stably suppress METTL1 expression in FHC cells, while control cells were infected with a non-targeting control shRNA lentivirus (sh-Ctrl). After confirmation of knockdown efficiency, total RNA was extracted and subjected to high-throughput RNA sequencing. Each group consisted of three independent biological replicates. Differential gene expression analysis was performed to identify transcriptional changes associated with METTL1 depletion.

first: Lianwen

last: Yuan


title: Gsx2 Regulates Oligodendrocyte Precursor Formation in the Zebrafish Spinal Cord

city: Aurora

first: Kim

last: Arena

state: CO

address: RC1S 12801 E 17th Ave

institute: University of Colorado Anschutz

summary: In the spinal cord, neural progenitor cells (NPCs) of the pMN domain produce neurons in early development followed by oligodendrocytee precursor cells (OPCs), which subsequently differentiate as oligodendrocytes (OLs), the myelinating glia of the central nervous system. The mechanisms that specify neural progenitor cells to the OL lineage are not yet well understood. Using zebrafish, we combined single-cell Multi-omics and in vivo genetic approaches to identify gsx2 as a regulator of OPC specification.

overall_design: Multi-ome seq:  Cells were extracted from the trunks of wildtype zebrafish larvae expressing the olig2:egfp transgene at 36 and 48 hours post fertilization. Cells were FAC sorted for EGFP and nuclei were isolated. Joint scRNA-seq and scATAC-seq libraries were produced using the 10X Genomics platform.

first: Kimberly

first: Christina

last: Kearns

first: Rebecca

last: O'Rourke

first: Bruce

last: Appel


title: Cytomegalovirus as a risk factor for exacerbation of Mycobacterium tuberculosis: development of a Mtb-CMV mouse model infection

city: London

first: Hannah

last: Painter

address: Keppel Street

country: United Kingdom

institute: LSHTM

summary: Cytomegalovirus (CMV) infection has been hypothesised to elevate risk of progression to tuberculosis (TB) disease. However, epidemiological human data show conflicting data. To date, an animal model of CMV and Mycobacterium tuberculosis (M. tb), the causative agent of TB, has not been established, to facilitate study of the relationship and mechanisms associated with the co-infection. In this study, C57Bl/6 mice were infected with 10^5 PFU murine CMV. Twelve weeks following mCMV infection, mice were infected with 100 CFU M. tb H37Rv. At one-, three- and four-weeks following M.tb infection, animals were culled by cervical dislocation and lungs were collected for downstream processing for CFU enumeration and bulk RNA sequencing.

overall_design: RNA-sequencing of lung samples obtained from mice co-infected with purified murine cytomegalovirus (mCMV) and Mycobacterium tuberculosis (M.tb). C57Bl/6 mice (6-8-weeks old) were infected with 10^5 PFU mCMV (Smith strain) by intraperitoneal injection. Twelve weeks following mCMV infection, mice were infected with 100 CFU M.tb H37Rv by intranasal infection. At one-, three- and four-weeks following M.tb infection, animals were culled by cervical dislocation and lungs were collected for downstream processing.

first: Helen

last: Fletcher


title: Spatial Transcriptomic Landscape of Invasion Patterns in Human Papillomavirus-Associated Endocervical Adenocarcinoma

city: St Louis

first: Margaret

middle: L

last: Axelrod

state: MO

address: 660 South Euclid Avenue, Campus Box 8118

department: Pathology and Immunology

institute: Washington University in St. Louis

summary: Human papillomavirus (HPV)-associated endocervical adenocarcinoma is the second-most common cancer of the uterine cervix. HPV-associated endocervical adenocarcinoma can be classified into histologic Silva patterns of invasion, which are associated with clinical outcome. However, the mechanisms underlying these patterns of invasion are incompletely understood. We used whole transcriptome spatial transcriptomics to examine gene expression differences separately in the tumor epithelium and the surrounding stromal immune microenvironment (SIME). Seven cases were evaluated, focusing on cases with two distinct patterns of invasion within the same tumor, to control for inter-patient heterogeneity. The most strongly upregulated pathways in both higher-risk tumor epithelium and SIME were associated with extracellular matrix (ECM) remodeling. Transcriptomic-based inference of immune cell populations showed an increase in macrophage populations in higher-risk tumor areas, confirmed by immunohistochemistry. Finally, we derived a four-gene signature from genes upregulated in higher-risk tumor epithelium (KRT6A, TNC, LAMC2 and FN1), which was associated with worse clinical outcome in an independent dataset (The Cancer Genome Atlas). Overall, this work demonstrates that ECM remodeling and macrophage presence are important in the progression to high-risk patterns of invasion in HPV-associated endocervical adenocarcinoma. In addition, we established a prognostic four-gene signature that is predictive of poor outcome.

overall_design: NanoString GeoMX was performed on human HPV associated endocervical adenocarcinoma with tumor (cytokeratin positive) profiled separately from the stromal immune microenvironment (cytokeratin negative)

first: Lulu

last: Sun


title: Protocol for massively parallel RNA assay coupled with immunoprecipitation (MPRNA-IP)

city: Baltimore

first: Taeyoung

last: Hwang

state: MD

address: 855 N. Wolfe St, 3rd floor, Suite 300

institute: Lieber Institute for Brain Development

summary: RNA mediates many cellular functions through interactions with RNA-binding proteins (RBPs), and defining the molecular principles underlying these interactions is essential for understanding RNA biology. Here, we describe a detailed protocol for MPRNA-IP, a Massively Parallel RNA assay coupled with ImmunoPrecipitation that enables high-throughput dissection of RNA–protein interactions in cells. This protocol provides a complete experimental and computational workflow, including the design of RNA tiles, large-scale oligonucleotide cloning, mammalian cell transfection, immunoprecipitation of RNA–protein complexes, sequencing library preparation, and downstream data analysis. As an example application, we synthesized a pool of oligonucleotides tiling six lncRNAs and studied PUM2-binding motifs. Together, these procedures allow high-throughput interrogation of RNA sequences to detect sequence and structural elements that contribute to protein binding. For further information on the development and applications of MPRNA-IP, see Lee et al., Nucleic Acids Research, 2024 (https://doi.org/10.1093/nar/gkae334).

overall_design: Overexpress a pool of oligonucleotides and perform immunoprecipitation in three biological replicates

first: Yu Hsuan

last: Lee


title: Adipose stromal cell-derived cancer-associated fibroblasts promote pancreatic adenocarcinoma through SFRP4 signaling

city: New York

first: Lingyi

last: Cai

address: 2960 Broadway

institute: Columbia University

summary: Progression of pancreatic ductal adenocarcinoma (PDAC) and other carcinomas relies on cancer-associated fibroblasts (CAFs). A subset of CAFs is derived from adipose stromal cells (ASCs) recruited by tumors and the ASC-CAF conversion has been associated with invasiveness and poor prognosis. To explore the underlying molecular mechanisms, we used a model based on primary ASC derived from human visceral adipose tissue co-cultured with human PDAC cell line Capan-1. To investigate cancer progression in vivo, we also used mice orthotopically grafted with mouse KPC cells. Genomic analysis revealed that Capan-1 co-culture induces Wnt and TGFβ signaling and extracellular matrix (ECM) gene expression in ASC. We investigated the function of two markers of the fibroblastic transition highly induced by cancer cells: a long non-coding RNA LINC01614 and a Wnt signaling modulator SFRP4. By using ASC with either SFRP4 or LINC01614 knocked out (ko), we showed that both genes are required for Wnt / TGFβ signaling and ECM induction in ASCs by Capan1. Analysis of changes in Capan-1 genes that rely on LINC01614 and SFRP4 expression in ASC also identified the Wnt and TGF pathways. SFRP4 ko in ASCs suppressed both migration and invasion of Capan-1 cells. We show that tumors in SFRP4 ko mice have less desmoplasia, less epithelial dedifferentiation, reduced growth rate, and reduced progression to metastasis. We conclude that SFRP4 promotes cancer progression in pancreatic cancer and is a promising therapeutic target.

overall_design: Visceral adipose tissue biopsies from a single patient were digested and the stromal vascular fraction was isolated and given the nomenclature vASC. These cells were then engineered to express red fluorescent protein (RFP) via transduction of a lentiviral contruct that also contained a puromycin resistance gene to allow for targeted selection of RFP+ cells. Capan-1 cells were engineered to express green fluorescent protein (GFP) via transduction of a lentiviral contruct that also contained a puromycin resistance gene to select for GFP+ cells. Seperate cohorts of the vASC_RFP cells were further engineered for knockdown of the LINC01614 and SFRP4 genes using CRISPR/Cas9 targeted mutagenesis via lentiviral transduction. The CRISPR/Cas9 construct targeting these genes also contained a blasticidin resistance gene to allow for selection of positively transduced cells. vASC_RFP, vASC_LINC01614_RFP, and vASC_SFRP4_RFP cells were cultured alone or in coculture with Capan-1_GFP cells in replicates for seven days. Cells were then trypsinized, made into single cell suspensions, and sorted (RFP vs GFP) using FACS. Total RNA-sequencing was performed on sorted cells to analyze changes in gene expressions.

first: Mikhail

last: Kolonin

first: Dimitris

last: Anastassiou

first: Joseph

last: Rupert


title: Uncovering a Key Metabolic Role of the GATA1 N-Terminus in Red Blood Cell Development [Bulk RNA-seq]

first: Te

last: Ling

overall_design: Transcriptional profiling utilizing Illumina® Stranded mRNA sequencing was conducted on HUDEP1 clones expressing either the full-length GATA1 (GATA1FL) or the short isoform (GATA1s_1D5 and GATA1s_1KI9) under expansion or differentiation (day 4) conditions, respectively.

first: Rashid

last: Mehmood

first: John

last: Crispino


title: Differential chromatin accessibility response to retinoic acid in neuroblastoma with ATRX in-frame-deletions versus ATRX loss-of-function

first: Sally

middle: Linda

last: George

address: 26 Oakleaf Avenue

institute: The Institute of Cancer Research

summary: Neuroblastoma is a childhood cancer, arising in the developing sympathetic nervous system. Differentiation therapy with 13-cis-retinoic acid (RA) is given to children with neuroblastoma to prevent relapse, however there is little understanding of which patients benefit. ATRX alterations are identified in 10% of high-risk neuroblastomas and associated with poor outcomes. The commonest type of ATRX alterations in neuroblastoma are in-frame multi-exon deletions, followed by nonsense mutations predicted to result in loss-of-function (ATRX LoF). We treated paired ATRX wild-type and LoF neuroblastoma cell-lines with RA: cells with ATRX LoF fail to upregulate direct RA target genes and show reduced chromatin accessibility differentiation and development related genes following RA treatment. Conversely, neuroblastoma models with in-frame deletions mount an appropriate epigenetic response to RA. Taken together this shows that the mechanism of differentiation in ATRX-altered neuroblastoma depends on the type of ATRX alteration, with implications relating to both oncogenesis and therapeutic response.

overall_design: ATRX loss-of-function (E6) and ATRX wildtype (p53(2)) SK-N-SH isogenic cell lines and ATRX in-frame-fusion (IFF) CHLA-90 and SK-N-MM cells were treated with either vehicle (DMSO) or 10 uM RA for 72 hours, before collection for ATAC sequencing.

first: Francois

last: Guillemot


title: Enrichment of DNA hypomethylated regions for biomarker discovery using 5-methylcytosine glycosylase [human_DMNseq]

city: Chicago

first: Yang

last: Li

institute: The University of Chicago

type: Methylation profiling by high throughput sequencing

summary: Here, we describe the DEMETER-assisted 5-Methylcytosine Nicking sequencing (DMN-seq) utilizing 5mC-specific glycosylase DEMETER to nick DNA at 5mC sites, enabling 5mC detection at the single-base resolution. Leveraging this nicking activity to deplete hypermethylated sites, we adapted DMN-seq to preferentially enrich and investigate hypomethylated regions in colorectal cancer samples. When applied to cell-free DNA as low as 0.1 ng, DMN-seq significantly expands the scope of cancer biomarkers by capturing hypomethylated regions, with high sensitivity and reproducibility even in low-input clinical samples.

overall_design: A total of 10 ng of gDNA from CRC patient cohort tumor tissue and adjacent healthy colon tissue was used as the untreated sample, whereas 40 ng was used as the DME-treated sample, 1 replicate was used for CRC cohort gDNA. All gDNA samples were prepared, fragmented, and end-repaired in the same way as those used for methylated DNA enrichment. 1 ng, 0.5 ng. A total of 0.1 ng of cfDNA from CRC patients and healthy controls was used as the untreated sample, whereas 2 technical replicates of the same input level were used as DME-treated samples.

first: Yiding

last: Wang

first: Chang

last: Ye


title: Pathogenic tau in the mouse locus coeruleus produces noradrenergic hyperactivity and neuropsychiatric phenotypes reminiscent of prodromal Alzheimer’s disease

city: Atlanta

first: Anu

last: Korukonda

state: Georgia

address: 615 Michael St

institute: Emory University

summary: Alzheimer’s disease (AD), though defined as a cognitive disorder, often presents neuropsychiatric symptoms such as anxiety, depression, agitation and sleep disruptions years before the onset of frank memory impairment. A hallmark prodromal feature is the accumulation of hyperphosphorylated “pretangle” tau (pTau) in the locus coeruleus (LC), the brain’s primary source of norepinephrine (NE), prior to any other region. While clinical studies link LC tau burden to behavioral abnormalities, causal mechanisms remain unclear. We developed a translationally-relevant mouse model that recapitulates the ‘LC-first’ phenomenon observed in humans using cell type-specific viral expression of pathogenic P364S mutant human tau in LC neurons. Three months post-infusion, pTau accumulation coincided with anxiety- and compulsive-like behaviors and reduced sleep spindles without altering overall sleep architecture. Consistent with the behavioral phenotypes, electrophysiological recordings revealed significant increases in spontaneous and evoked LC activity, accompanied by astrocytic reactivity with no apparent neuronal death. Transcriptomic analysis identified upregulation of the pacemaker potassium/sodium channel Hcn2 in the LC, which may contribute to neuronal hyperexcitability. To further define molecular mechanisms linking pTau to behavioral and cellular phenotypes, we developed a cell type-specific proteomics approach, which revealed synaptic and metabolic alterations in LC neurons associated with tau pathology. Interestingly, early anxiety-like behaviors observed at 3 months diminished at later timepoints (6-9 months) and were replaced by anxiolytic phenotypes. These findings demonstrate that pTau triggers phenotypes reflective of LC-NE hyperactivity in the early stages of AD pathogenesis, laying the foundation for the development of LC-based disease-modifying targets to address neuropsychiatric manifestations.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for LC-mRNA (IP) for P364S mutant human tau-infused or EYFP-infused controls. The background strain is C57Bl/6 and the exact cross is: Th-cre (B6.Cg-7630403G23RikTg(Th-cre)1Tmd/J, The Jackson Laboratory, #008601) crossed with Slc6a2-eGFP/Rpl10a (B6;FVB-Tg(Slc6a2-eGFP/Rpl10a)JD1538Htz/J, The Jackson Laboratory, #031151).

first: David

last: Weinshenker


title: RBProximity-CLIP enables subcellular mapping of RNA-binding protein interactions at nucleotide resolution [CLIP]

city: Bethesda

first: Markus

last: Hafner

state: Maryland

address: 50 South Drive

institute: NIAMS/NIH

summary: RNA-binding proteins (RBPs) orchestrate post-transcriptional gene regulatory (PTGR) processes through specific interactions with RNA molecules, which are tightly regulated in both space and time, including through the subcellular compartmentalization of RBPs and their target RNAs. To characterize these interactions in specific subcellular compartments, we developed RBProximity-CLIP, an approach that integrates targeted APEX2-based proximity labeling with 4-thiouridine–enhanced RNA–protein crosslinking to enable the isolation and profiling of target sites of individual RBPs in a compartment-specific manner . Using this approach, we profiled three RBPs—AGO2, YBX1, and ELAVL1—within the cytoplasmic, nuclear, and nucleolar compartments. RBProximity-CLIP enabled robust isolation of compartment specific RBP-RNA interactome and new insights of compartmental specific gene regulation of distinct RBPs.

overall_design: We developed RBProximity-CLIP methodology for isolating and profiling  RNA binding proteins  (herein: AGO2, YBX1 and ELAVL1) in cytoplasm, nucleus and nucleolar compartments.

first: Iwona

last: Nowak

first: Ahsan

middle: H

last: Polash

first: Hang

middle: T

last: Huynh

first: Mahekdeep

last: Kaur

first: Vivian

last: Lobo

first: Jérémy

middle: B

last: Scutenaire

first: Michelle

last: Fong

first: Ghaliah

last: Alluhaibi

first: Dimitrios

last: Anastasakis

first: Daniel

last: Benhalevy

first: Aishe

last: Sarshad


title: Enrichment of DNA hypomethylated regions for biomarker discovery using 5-methylcytosine glycosylase

first: Yang

last: Li

overall_design: 200 ng of λ-DNA (NEB) or mESC gDNA supplemented with 0.1% (w:w) λ-DNA were fragmented and added with 200 bp dsDNA oligo and 164 bp dsDNA oligo as spike-in. They are subjected to DMN-seq library preparation protocol.

first: Wang

last: Yiding

first: Li

last: Yang

first: Ye

last: Chang


title: Norepinephrine derepresses the Fur regulon of Neisseria gonorrhoeae to enable growth in iron-limited conditions

city: Charlottesville

first: Camille

middle: Shea

last: Westlake

state: Virginia

address: 1340 Jefferson Park Avenue, Pinn Hall Room 7213

department: Microbiology, Immunology, Cancer Biology

institute: University of Virginia

summary: Neisseria gonorrhoeae (Gc) is the Gram-negative bacterium that causes gonorrhea, a prevalent sexually transmitted infection that can have life-threatening clinical sequelae. Gc requires iron for human infection and uses the iron-responsive, iron-binding transcriptional repressor Fur to maintain iron homeostasis. Gc infects mucosal sites, where the neuroendocrine hormone norepinephrine (NE) is produced by the autonomic nervous system and various epithelial and immune cell types.  By RNA-seq, we determined that NE rewires gonococcal gene expression to increase capacity for iron uptake while enabling increased intracellular iron availability. Of the 30 genes that were differentially expressed in NE-treated compared to untreated bacteria, 27 have Fur box-containing promoters.

overall_design: Neisseria gonorrhoeae of strain background FA1090, constitutively expressing the OpaD protein (PMID: 23625842) was grown to mid-log phase in Chelex-treated chemically defined medium (PMID: 32202529), alone or with 12.5 µM Fe(NO3)3 or with 10 µM norepinephrine for 1 hour. Total RNA was isolated using the RNeasy Plus minikit. Ribosomal RNA was depleted using the NEBNext rRNA depletion kit and the remaining RNA was reverse-transcribed into a cDNA library for Illumina sequencing. Reads were mapped to the FA1090 genome (GCA_009757095.1, ASM975709v1).

first: Camile

middle: S

first: Alison

last: Criss


title: SciPhy: A Bayesian phylogenetic framework using sequential genetic lineage tracing data

city: Seattle

first: Samuel

last: Regalado

state: WA

address: William H. Foege Hall, 3720 15th Ave NE

institute: University of Washington

summary: CRISPR-based lineage tracing offers a promising avenue to decipher single cell lineage trees, especially in organisms that are challenging for microscopy. A recent advancement in this domain is lineage tracing based on sequential genome editing, which not only records genetic edits but also the order in which they occur. To capitalize on this enriched data, we introduce SciPhy, a simulation and inference tool integrated within the BEAST 2 framework. SciPhy utilizes a Bayesian phylogenetic approach to estimate time-scaled phylogenies and cell population parameters. After validating SciPhy using simulations, we apply it to two lineage tracing datasets for which we estimate time-scaled trees together with cell proliferation rates. Using simulated and real lineage tracing data obtained from a monoclonal culture of HEK293T cells , we compare SciPhy to other lineage reconstruction methods, and find that SciPhy consistently constructs distinct, and more accurate lineage trees. In particular, for HEK293T cells, SciPhy trees stand out for their later estimated cell division times. In addition, SciPhy reports uncertainty as well as proliferation rates, neither of which are available within a UPGMA analysis. Our second example applies SciPhy to the study of murine gastruloids, and showcases the use of complex models of time-varying population growth to capture realistic aspects of this in-vitro model of early mammalian development. Together, these examples showcase the application of advanced phylogenetic and phylodynamic tools to explore and quantify cell lineage trees, laying the groundwork for enhanced and confident analyses to decode the complexities of biological development in multicellular organisms. SciPhy’s codebase is publicly available at https://github.com/azwaans/SciPhy.

overall_design: Single-cell RNA sequencing was performed on murine gastruloids derived from monoclonal expansion of single mouse embryonic stem cells. Six independently generated monoclonal gastruloids were pooled in pairs and sequenced across three independent 10x Genomics lanes. Cells harbored stably integrated CRISPR-based DNA recording barcodes (“DNA Typewriter”) that accumulate ordered genetic insertions during proliferation. Lineage tracing barcode sequences recovered from single cells were analyzed using a Bayesian phylogenetic framework to reconstruct time-scaled lineage trees and infer cell population growth dynamics.

first: Sophie

last: Seidel

first: Antoine

last: Zwaans

first: Junhong

last: Choi

first: Jay

last: Shendure

first: Tanja

last: Stadler


title: Transposon sequencing reveals Burkholderia gene fitness in a spaceflight-relevant plant-pathogen interaction

city: Kennedy Space Center

first: Aubrie

middle: E.

last: O'Rourke

state: Florida

address: Kennedy Space Center

institute: National Aeronautics and Space

summary: The spaceflight environment imparts unique selective pressures on the plants and microbes of plant growth chambers on the International Space Station (ISS), which generally manifests through genetic signatures associated with a heightened response to stress. Terrestrially, a baseline understanding of the gene fitness response for any plant growth promoting microbe when in a tripartite relationship with host and pathogen is currently unknown and is important to characterize before closed-environment spaceflight implementation. To that end, this study evaluated the behavior of an ISS plant habitat isolate of Burkholderia contaminans as tomato seeds transition to seedlings and assessed gene fitness during challenge with Fusarium oxysporum f. sp. lycopersici (FOL), the causal agent of Fusarium wilt. To probe mechanisms underlying the bacterial-fungal interaction between B. contaminans and FOL in the tomato root zone, a genome-wide transposon mutant library was developed of the B. contaminans isolate.  Using a seed film delivery method vetted for spaceflight, a Tn-Seq library of B. contaminans was applied to Solanum lycopersicum cv. Red Robin seeds. Transposon sequencing (Tn-Seq) analysis revealed that the type II secretion system (T2SS) was critical for root zone establishment, whereas a Nudix hydrolase was specifically important for responding to FOL infection, and provided further confirmation that antifungal and siderophore-producing gene clusters were not.

overall_design: In each plant growth vessel (magenta jar) two seed films were placed in one corner of each for the following scenarios: a) “NoSeed”-a control where only the B. contaminans Tn-Seq library (108 CFU/mL) is applied, b) “Root” or “Shoot”-a treatment where B. contaminans Tn-Seq library (108 CFU/mL) plus seed is applied and c) “FOLRoot” or “FOLShoot”-a treatment of B. contaminans Tn-Seq library (108 CFU/mL) plus seed for additional FOL challenge. The final 108 CFU/mL count was obtained by adding 10 µL droplets of the bacterial cultures at 1010 CFU/mL to the seed film. The B. contaminans Tn-Seq library was revived by slightly thawing the freezer stock and pipetting 100 mL of Tn-Seq library into 3 mL of TSB containing 50 mg/mL kanamycin and incubated at 35°C with shaking at 200 rpm for 16 hours. Three separate Tn-Seq starter cultures were prepared to serve as timepoint 0 (T0) and to inoculate the seed films as the three biological replicates (TnA, TnB, and TnC) of the experiment. Cells which were not used for the experiment were retained as T0 for Tn-Seq sequencing. Magenta vessels were placed into a controlled-environment chamber set to 50% RH and ambient CO2. At seven days after initiation, vessels were thinned to one plant per vessel and 10 µL of 103 CFU/mL FOL culture was applied to the corner opposite to the established plant in the fungal control and experimental treatment vessels. Plants were grown for 28 days total before they were photographed and harvested. At harvest, shoot and root tissues were separated and placed into 50 mL Falcon tubes with 10 mL of 50 mg/mL kanamycin in TSB media and allowed to grow for 16 hours at 35°C with shaking at 200 rpm to select for the transposed B. contaminans Tn-Seq library. Plant material was removed from the Falcon tube, the bacterial culture was pelleted by centrifugation at 3000 rpm for 5 min, and media decanted. Three cell pellets were obtained for scenarios a, b roots, b shoots, c roots, c shoots, and timepoint 0 for a total of 18 cell pellets for DNA extraction. A total of 19 Tn-Seq libraries were prepared including the input library that made the freezer stock.

first: Anya

last: Volter

first: Jessica

last: Atkin

first: Aaron

last: Curry

first: Anirudha

last: Dixit


title: Epicardium Regulates Ventricular Compaction

city: Houston

first: Mingfu

last: Wu

address: 4349 Martin Luther King Boulevard

department: Department of Pharmacological and Pharmaceutical Sciences

institute: University of Houston

summary: We used single nucleus RNA sequencing (snRNAseq) to show that Numb and Numblike specific deletion in epicardium regulates ventricular compaction via the Numb Family Proteins-Fgf Signaling

overall_design: Whole hearts harvested from control (no Cre) and Tbx18Cre/+ mediated Epicardium specific Numb and Numblike double knockout (Tbx18Cre/+; Numbflox/flox; Numblikeflox/flox) E15.5 mice embryos were snRNA sequenced. We analyzed all the cell types in the heart at E15.5 and compared the different expression profiles between control and knockout for all the clusters, to determine how Numb Family Proteins in epicardium regulate ventricular compaction.

first: Anika

last: Nusrat

first: Shiyanth

last: Thevasagayampillai

first: Aaranyah

last: Kandasamy

first: Preethi

last: Gunaratne


title: Polianthes tuberosa L. Extract Suppresses Melanogenesis through Concurrent Inhibition of cAMP/CREB and MAPK Signaling Pathways

city: WuXi

first: Li

last: qiaozhen

address: Meicun street

institute: Jiangnan University

summary: Ultraviolet (UV) radiation is a primary environmental stimulus for skin hyperpigmentation. Polianthes tuberosa L. (PT), rich in polyphenols and flavonoids, possesses antioxidant and anti-inflammatory properties, yet the anti-melanogenic mechanism of the PT extract (PTE) remains unexplored. Network pharmacology revealed nuclear factor erythroid-derived 2-like 2 (NFE2L2), superoxide dismutase 1 (SOD1), nuclear factor kappa B subunit 1 (NFKB1) and interleukin-6 (IL-6) as core targets. This suggests that the therapeutic mechanism of the PTE may involve the coordinated modulation of oxidative stress (via NFE2L2/SOD1) and inflammation (via NFKB1/IL-6)—two interconnected pathways that critically drive melanogenesis following UV exposure. The PTE dose-dependently inhibited UV-induced release of inflammatory cytokines and paracrine melanogenic factors in the immortalized human keratinocyte cell line. It also mitigated UV-induced collagen reduction in fibroblasts. In the mouse melanoma cell line B16F10, the PTE significantly suppressed melanogenesis and tyrosinase activity (p < 0.001). Integration of transcriptomic and proteomic data provided a complementary view of molecular regulation at both the messenger RNA and protein levels. This convergent evidence indicated that PTE acts as a concurrent inhibitor of the cyclic adenosine monophosphate (cAMP)/protein kinase A/cAMP-response element binding protein and mitogen-activated protein kinase signaling axes, as validated by quantitative polymerase chain reaction and western blot analyses. This dual inhibition led to the downregulation of microphthalmia-associated transcription factor and its downstream melanogenic enzymes. Our findings underscore the potential of PTE as a multifaceted, natural whitening agent for cosmetic applications.

overall_design: RNA-seq of Murine B16F10 melanoma cells in control and Polianthes tuberosa L. extract-treated group


title: Gene expression profiling of cultured alveolar macrophages following induction of immunological cell death

city: Suita

first: Daisuke

last: Motooka

state: Osaka

address: 3-1, Yamadaoka

country: Japan

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

summary: Alveolar macrophages (AMs) play a crucial role in protecting the lungs from pathogens by inducing immunogenic cell death (ICD). However, the type of cell death that effectively induces protective immunity remains to be fully understood. In our investigation of the mechanisms regulating AM activation and lung immune responses, we found that AMs are highly susceptible to necroptosis, a form of ICD. Treatment with pan-caspase inhibitor,  emricasan, directly induced cell death in AMs, resulting in the release of interleukin 1α. To understand the characteristics of inflammatory responses induced by emricasan, we performed a comprehensive bulk RNA-seq analysis of cultured AMs treated with emricasan for 6 hours.

overall_design: In vitro cultured alveolar macrophages (prepared from C57BL/6J mice) were cultured with or without emricasan (40 μM) for 6 hours.

first: Takumi

last: Adachi

first: Etsushi

last: Kuroda


title: Convergent antibody mutation trajectories convert functional self-tolerance to genetic self-tolerance in IGHV4-34 B cells.

city: Darlinghurst

first: Timothy

middle: John

last: Peters

state: NSW

address: 384 Victoria St

department: Immunology

institute: Garvan Institute of Medical Research

summary: Preventing autoantibody secretion by rendering self-reactive B cells functionally silent through clonal anergy has long posed the question of why fill the circulating B cell repertoire with cells that cannot secrete antibody? Here we address this question from the perspective of B cells that comprise 5-10% of the human circulating repertoire, expressing self-reactive surface immunoglobulins employing the IGHV4-34 heavy chain variable element. Using gene targeting to construct mice expressing a representative human IGHV4-34 antibody on the surface of many B cells, we show these cells are prevented from autoantibody secretion by B cell clonal anergy marked by downregulation of surface IgM, induction of tolerance-response mRNAs, and exclusion from the marginal zone and B1 cell subsets. This functionally tolerant state is overridden when the IGHV4-34 B cells cross-react with a virus, which stimulates the self-reactive B cells to hypermutate in germinal centres. Within 16 days of infection, 99% of daughter cells have acquired one of five heavy chain mutations that diminish binding to self but preserve virus binding, and 33% had combined 2 or 3 of these mutations to become genetically self-tolerant and virus specific. These results demonstrate, from the perspective of a pathologically important human autoantibody class and the world’s most successful virus vaccine, how human antibody specificity is sculpted in the progeny of anergic B cells to yield antibodies that bind a virus but not self.

overall_design: We generated knock-in transgenic mice expressing a representative human IGHV4-34 antibody on a large percentage of the nascent B cell repertoire. We focussed on the unmutated ancestor of antibody 589, a highly mutated IGHV4-34 antibody against A14L, a nonglycosylated major protein of the vaccinia virus inner membrane. Antibody 589 was isolated from a healthy adult who had received vaccinia virus during the global vaccination effort that led to smallpox eradication in the 1970’s. Three decades later the donor was boosted with vaccinia virus and after 14 days individual plasma cells were sorted from the blood and antibody heavy and light chain mRNAs sequenced and expressed.

first: Christopher J.

last: Jara

first: Sherin

last: Zachariah

first: Katherine JL

last: Jackson

first: Timothy J

first: Etienne

last: Masle-Farquhar

first: Eric

last: Lam

first: Megan

last: Faulks

first: Amanda

last: Russell

first: David C.

last: Tscharke

first: Robert

last: Brink

first: Christopher C.

last: Goodnow

first: Joanne H.

last: Reed


title: RBProximity-CLIP enables subcellular mapping of RNA-binding protein interactions at nucleotide resolution [RNA-seq]

first: Markus

last: Hafner

overall_design: About 6 x 105 cells were seeded in 60-mm plates and transfected the next day with 50 pmol of siRNA targeting YBX1 (ThermoFisher Scientific, Cat.#4390824, siRNA ID s9731) or a control siRNA (ThermoFisher Scientific, Cat.#4390843) preincubated for 20 min at room temperature with 5 µL of Lipofectamine RNAiMAX transfection reagent (ThermoFisher Scientific, Cat.#13778150) in OPTI-MEM medium. Cells were collected and split in two 72 h after transfection. Total proteins were extracted with RIPA buffer on one half of the cells and used for western blot to assess the silencing efficiency. Total RNAs were extracted on the second half of the cells with TRIzol™ Reagent (ThermoFisher Scientific, Cat.#15596018) and treated with DNAse I using the Direct-zol RNA Miniprep kit (Zymo research, Cat.#R2052) according to the manufacturer’s instructions. Each siRNA transfection was performed in biological triplicate. rRNA-depleted libraries were generated from 1 µg of total RNA using the NEBNext® rRNA Depletion Kit v2 (New England Biolabs, Cat.#E7400X) and NEBNext® Ultra™ II Directional RNA Library Prep Kit for Illumina® (New England Biolabs, Cat.#E7760L) according to the manufacturer’s instructions. The quality of the initial RNA material and of the final cDNA libraries was assessed using a TapeStation system (Agilent Technologies). Final libraries were sequenced on an Illumina NovaSeq X Plus platform (Illumina) to generate paired-end reads (2 × 50 bp).

first: Iwona

last: Nowak

first: Ahsan

last: Polash

first: Hang

last: Huynh

first: Mahekdeep

last: Kaur

first: Vivian

last: Lobo

first: Jérémy

last: Scutenaire

first: Michelle

last: Fong

first: Ghaliah

last: Alluhaibi

first: Dimitrios

last: Anastasakis

first: Daniel

last: Benhalevy

first: Aishe

last: Sarshad


title: Deficiency Disrupts Photoreceptor Viability and Synaptic Integrity in a Choroideremia Mouse Model

city: Wuhan

first: Yin

last: Shen

address: Zhang Road No. 99 Jiefang Road 238

institute: Renmin Hospital of Wuhan University

summary: Choroideremia, an X-linked retinal dystrophy causing progressive vision loss, arises from deficient Rab escort protein-1 (REP1), critical for the prenylation of Rab proteins. The precise mechanisms linking REP1 dysfunction to retinal degeneration remain unclear. Here, we generated conditional REP1 knockout mice to model choroideremia and dissect REP1’ s role in retinal homeostasis. Histological analysis revealed severe photoreceptor (PR) layer thinning (91.2 ± 2.34 μm vs. 188.1 ± 3.46 μm in controls) by postnatal day 30, accompanied by disrupted synaptic architecture in the outer plexiform layer (OPL). Electroretinogram (ERG) demonstrated profound visual dysfunction, with scotopic a-wave and b-wave amplitudes reduced by 69.6% and 86.7%, respectively, indicating PR and perhaps bipolar cell (BC) impairment. RNA-Sequencing and immunofluorescence labeling showed downregulation of PR, synaptic, and phototransduction-related molecules as well as disrupted structural integrity of PRs. Transmission electron microscopy (TEM) revealed ultrastructural synaptic defects, including shortened synaptic ribbons and loss of invaginated triads. Our findings establish an essential role for REP1 in maintaining PR viability and synaptic connectivity. Furthermore, we also provide mechanistic insights into the progression of choroideremia associated with the activation of microglia mediated by the NF-κB pathway, suggesting that neuro-immune is one of the potential therapeutic targets for preserving retinal circuitry in patients with choroideremia.

overall_design: RNA-seq profiling of C57BL/6J wildtype mice and conditional REP1 knockout mice to model choroideremia at P30.

first: Jiangbo

last: Yan

first: Tianlu

last: Zhang

first: Yuxin

last: Du

first: Zeyuan

last: Pu

first: Yazhen

last: Ma


title: Enrichment of DNA hypomethylated regions for biomarker discovery using 5-methylcytosine glycosylase [mouse_DMNseq]

first: Yang

last: Li

overall_design: mESC gDNA supplemented with 0.1% (w:w) λ-DNA were fragmented and added with 200 bp dsDNA oligo and 164 bp dsDNA oligo as spike-in. They are subjected to DMN-seq library preparation protocol.

first: Yiding

last: Wang

first: Chang

last: Ye


title: cmLumiOpto gene therapy in combination with chemotherapy to treat triple negative breast cancer.

city: COLUMBUS

first: Zhuoxin

last: Zhou

state: OH

address: Gateway Lofts Columbus, 2211 Dublin Rd

institute: Ohio State University

summary: Triple-negative breast cancer (TNBC) is aggressive, resistant to chemotherapy, and prone to recurrence. While PARP inhibitors (PARPi) help some patients, most do not respond. We developed a novel therapy combining mitochondria-targeted luminoptogenetics (cmLumiOpto) with PARPi to enhance treatment. Delivered via an anti-CD276 monoclonal antibody-conjugated exosome-AAV (mAb-Exo-AAV), cmLumiOpto disrupts mitochondrial membrane potential, inducing cancer cell death. In vitro, the combination increased cytotoxicity; in vivo, it reduced tumor burden by 95–100%, suppressed xenograft growth, and blocked metastasis in TNBC models. Mechanistic studies showed mitochondrial depolarization, DNA damage, cytokine release, and immune infiltration, highlighting a promising strategy for TNBC therapy. The mechanism of action was further explained through bulk RNA sequencing post-treatment. These findings highlight the therapeutic potential of our cmLumiOpto gene therapy for treatment of triple negative breast cancers.

overall_design: Metastasized 4t1 tumour cells were harvested from the lungs of BALB/cJ mice after three weeks of treatment with cmLumiOpto gene therapy in combination with PARPi.Total RNA was extracted from the harvested tumour cells using the RNeasy Plus Mini Kit (Qiagen, Germantown, MD, USA). Bulk RNA sequencing with cDNA library construction were performed at Novogene America (Sacramento, CA, USA) using the Illumina HiSeq™ X Ten platform.


title: Evaluation of targeted and immune combination therapies in a rat model of hormone receptor-positive breast cancer  [RNA]

city: Boston

first: Kornelia

last: Polyak

address: 450 Brookline Ave. S1070

institute: Dana-Farber Cancer Institute

summary: Breast cancer hormone receptor (HR) estrogen and progesterone (ER/PR) positive tumors were generated in an outbred rat model (Sprague-Dawley) through the injection of the carcinogen N-nitrosomethylurea (NMU). These tumors were treated with immune checkpoint blockade (anti PD-L1) in combination with targeted therapies, including an TNK1/IKBKE inhibitor, a KMT5B/C inhibitor, a TGF-b inhibitor, and a selective estrogen receptor degrader (SERD). This dataset includes single-cell RNA sequencing (scRNA-seq) data from tumors subjected to various treatment setups. Whole-tumor digestion was performed to preserve a comprehensive representation of the tumor microenvironment. The study aims to identify mechanisms that sensitize HR postive tumor subtypes to immune checkpoint blockade therapy.

overall_design: Tumors were induced by intraperitoneal injection of 50 mg/kg NMU on days 46 and 53 of age. Treatment with NIS793 (a TGFBR inhibitor), IKBKEi (a TNK1/IKBKE inhibitor), A196 (a SUV420H1/SUV420H1 inhibitor), Fulvestrant (an estrogen receptor antagonist), ⍺PD-L1, or a combination of targeted therapy with immune checkpoint blockade (ICB) was initiated when tumors reached 8 mm in size, more than one tumor was detected, or a tumor remained unchanged in size for more than three days. Rats were randomly assigned to treatment groups, and tumor size was measured every other day using calipers.

first: Ernesto

last: Rojas Jimenez

first: Triet

last: Bui

first: Pengze

first: Marco

last: Seehawer

first: Jun

last: Nishida


title: Evaluation of targeted and immune combination therapies in a rat model of hormone receptor-positive breast cancer

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

overall_design: Refer to individual Series


title: Cornulin transcriptomic analysis in Cal27 oral cancer cells

city: Chandigarh

first: Arnab

last: Pal

state: Chandigarh UT

address: Sector 12

department: Biochemistry

country: India

institute: Post Graduate Institute of Medical Education and Research

summary: RNA-Seq profiling of Cal27 Oral squamous carcinoma cells overexpressing Cornulin (CRNN), GFP control, and untreated group (UNT) to study differential gene expression. This project investigates the transcriptomic alterations induced by the overexpression of Cornulin (CRNN) in the Cal27 human oral squamous cell carcinoma cell line. The study design included three experimental groups: (1) Cornulin-overexpressing Cal27 cells, (2) GFP-expressing Cal27 cells as vector controls, and (3) untreated Cal27 cells. Total RNA was extracted using Trizol reagent, followed by RNA quality assessment via Bioanalyzer. Strand-specific mRNA libraries were prepared using the TruSeq Stranded mRNA kit, pooled at 6 nM, and sequenced using an Illumina NovaSeq 6000 S4 flow cell in a paired-end (2x100 bp) configuration. Raw FASTQ files were analyzed by aligning to the human genome (GRCh38), and gene-level quantification was performed. Differential expression analysis was conducted using the DESeq2 R package. DEG lists from each comparison group were subjected to pre-ranked gene set enrichment analysis (GSEA) to identify enriched pathways and gene ontologies. The dataset provides insights into Cornulin-mediated gene regulation.

overall_design: Paired-end stranded RNA sequencing of Cornulin-overexpressing Cal27 cells vs. GFP vector control and untreated Cal27 cells. Two biological replicates per group.

first: Arindam

last: Maitra


title: CCL18 as a immunosuppressive biomarker for prognosis in ABC diffuse large B-cell Lymphoma

city: Madrid

first: Juana

last: Serrano López

address: Avda Reyes Católicos 2

department: Hematology

country: Spain

institute: IIS-FJD

summary: Objective: decipher TME of ABC lymphomas to describe new biomarkers and treatment strategies. Methods: We analyzed 43 DLBCL samples (23 GCB and 20 ABC) by Nanosring PanCancer Immune Profiling Panel with additional 30 cell-of-origin genes. ABC lymphomas showed distinct expression profile than GCB with a markedly downregulation of MAPK10, more entropy with eosinophils enrichment. Multivariate analysis identified CCL18 as an independent adverse prognostic factor in ABC. lymphomas. Connectivity mapping identified proteasome inhibitors and CDK4/6 inhibitors as promising therapeutic candidates.

overall_design: Transcriptome analysis of 43 DLBCL samples (23 GCB and 20 ABC) by using Nanostring PanCancer Immune Profiling Panel woth 30 cell-of-origin genes. MAPK10 methylation by Pyrosequencing. Deconvolution analysis by CIBERSORT and GSEA. Data analysis: Cox regression and LASSO regularization. Therapeutic candidates through connectivity mapping.

first: Marta

last: Rodríguez-Moreno

first: Francisco

last: Rojas

first: Jesús

last: Frutos

first: Ignacio

last: Mahillo

first: Cristina

last: Serrano

first: Alberto

last: López

first: Teresa

last: Morales-Ruiz

last: Roldán-Arjona

first: Joaquín

last: Sánchez-García

first: Ana

last: Río-Machín

first: Daniel

last: Morillo

first: Raúl

last: Córdoba

first: Pilar

last: Llamas-Sillero

first: Socorro

last: Rodríguez-Pinilla

last: Serrano-López


title: Fecal microbiota transplantation mitigates cardiac remodeling and functional impairment in mice with chronic colitis

city: Galveston

first: Kevin

middle: M.

last: Lopez

state: Texas

address: 301 University Blvd

department: Internal Medicine

institute: University of Texas Medical Branch at Galveston

summary: Inflammatory bowel disease (IBD) is a chronic inflammatory disorder with significant extraintestinal manifestations, including cardiovascular derangements. However, whether and how chronic colitis impairs heart function remain unclear. We investigated the impact of chronic colitis on heart function and transcriptome using two mouse models: DSS-treated and Il10-/- mice. Echocardiography was employed to assess heart function and molecular characterization was performed using RNA-sequencing, RT-qPCR, and western blot. Both models exhibited significant cardiac impairment, including reduced ejection fraction and fractional shortening, along with increased collagen deposition, inflammation, and myofibril reorganization. Molecular analyses revealed upregulation of fibrosis markers and β-catenin reactivation, indicating a pro-fibrotic cardiac environment. RNA-sequencing unveiled a shared upregulation of eicosanoid-associated and inflammatory genes (Cyp2e1, Map3k6, Pck1, Cfd) across both models, along with model-specific alterations in pathways governing cAMP and cGMP signaling, arachidonic and linoleic acid metabolism, and immune cell responses. DSS colitis caused differential regulation of 232 cardiac genes, while Il10-/- colitis yielded 105 dysregulated genes. Importantly, therapeutic fecal microbiota transplantation (FMT) restored heart function in both models, characterized by reduced fibrosis markers and downregulated pro-inflammatory genes, while also mitigating intestinal inflammation. Notably, Il10-/- mice showed relatively less cardiac recovery following FMT, highlighting IL-10’s cardioprotective and anti-inflammatory contribution. Our findings provide direct evidence that chronic colitis impairs heart function, elucidate novel insights into colitis-induced cardiac remodeling, and suggest that FMT mitigates cardiac dysfunction by attenuating systemic inflammation and correcting gut dysbiosis. Further evaluation of gut-heart interactions and microbiome-based therapies is needed to improve cardiovascular health in IBD patients.

overall_design: To investigate the genetic and transcriptomic changes to the heart in chronic colitis and how therapeutic fecal microbiota transplantation affects such changes. Differential gene expression profile analysis was performed using RNA sequencing of total RNAs isolated from the colons of two independent mouse models using C57BL/6J mice and 129(B6)-Il10tm1Cgn/J mice strains. The comparative groups for C57BL/6J mice include (1) vehicle control, (2) DSS, (3) DSS+FMT. The comparative groups for 129(B6)-Il10tm1Cgn/J mice include (1) wild type, (2) IL-10 KO, and (3) IL-10 KO+FMT. Gene ontology and KEGG pathway encrichment was performed on the gene lists for each model, independently.

first: Xiaoying

last: Zhong

first: Max

last: Liu

first: Ying

last: Xiao

first: Rongliwen

last: Ou

first: Thierry

last: Kochkarian

first: Don

middle: W

last: Powell

first: Ken

last: Fujise

first: Qingjie

last: Li


title: A single-cell transcriptomic landscape of peripheral leukocytes in healthy dogs

city: Jeju-si

first: Myung-Chul

last: Kim

state: Jeju-do

address: 102, Jejudaehak-ro, Jeju-si, Jeju-do

country: South Korea

institute: Jeju National University

summary: Single cell RNA sequencing (scRNA seq) is a powerful approach for dissecting cellular heterogeneity in both human and veterinary medicine. Although prior canine studies have profiled peripheral leukocytes at the single-cell level, the accurate identification, functional characterization, and clinical relevance of specific immune subsets remain underexplored. Here, we aimed to generate a comprehensive single cell atlas of healthy canine peripheral blood leukocytes and to uncover the molecular mechanisms that underpin their homeostasis. We performed scRNA seq on peripheral leukocytes from six clinically healthy, small breed adult dogs using the 10x Genomics Chromium platform. Sequencing reads were aligned and annotated on canFam4, and data were processed in Seurat (v.5.1.0) with standard quality control, normalization, and integration workflows. Downstream bioinformatic and statistical analyses were conducted in RStudio (v.4.2.0). After filtering, we obtained 30,040 high quality single cell transcriptomes, which clustered into 51 distinct immune subsets. Leveraging canFam4 improved recovery of single-cell and key lineage and functional markers, such as CD14⁺CD33+ monocytes, XCR1⁺CD1D⁺ dendritic cells, CEACAM1+CD24⁺ neutrophils, and IL32⁺BATF⁺ regulatory T cells previously underrepresented in canFam3.1 based studies. We also identified interferon enriched CD14⁺ monocyte and CD4⁺ T subsets implicated in canine myxomatous mitral valve disease. Analysis of CD274⁺ myeloid and PDCD1⁺ T cell clusters provided evidence roles for IL 10 signaling, malate metabolism (MDH1/MDH2), and viral response pathways (LAG3, CD47, TBX21) in maintaining homeostasis. Finally, cohort specific differences in T cell exhaustion scores and proliferative (cycling) T cell fractions pointed to variable antigenic exposures. The single cell transcriptomic landscape of circulating canine leukocytes unveiled novel, clinically relevant immune subsets and the molecular pathways that maintain immune homeostasis. Our dataset and analytical framework represent valuable resources for translational research, particularly in cancer immunology.

overall_design: Peripheral leukocytes were isolated from six clinically healthy, small breed adult dogs. Each PBMC sample was subject to scRNA-seq using 10x Genomics Chromium platform.

first: Taeeun

last: Gu

first: Hyeewon

last: Seo

first: Yewon

last: Moon

first: Nicholas

last: Borcherding

first: Ryan

last: Kolb

first: Woo-Jin

last: Song

first: Chung-Young

last: Lee

first: Hyun Je

first: Weizhou

last: Zhang


title: Transcriptome analysis of mural granulosa cells and cumulus cells during ovulation in mouse ovary

city: Ube

first: Amon

last: Shiroshita

state: Yamaguchi

address: Minamikogushi 1-1-1

institute: Yamaguchi university

summary: This study investigates the transcriptomic changes in mural granulosa cells and cumulus cells collected at different time points during the ovulation process in mice following hCG stimulation. RNA-seq was performed to identify dynamic gene expression patterns associated with ovulation, and the data serve as a resource for understanding temporal regulation of ovarian function.

overall_design: Total RNA was extracted from mural granulosa cells and cumulus cells isolated from mouse ovaries at multiple time points (0h, 4h, 8h, 12h) after hCG injection. PolyA-enriched libraries were prepared and sequenced on an Illumina NovaSeq 6000 platform. Read alignment and gene expression quantification were performed using CLC Genomics Workbench with default parameters.


title: Transcriptomic analysis reveals the potential role of TOE1 in hepatocellular carcinoma

city: Shenzhen

first: Yi

last: Cai

address: Shenzhen Campus of Sun Yat-sen University, Guangming New District, Shenzhen

institute: Sun Yat-sen University School of Medicine

summary: This study integrates bioinformatics analysis and in vitro cellular experiments to elucidate the role of TOE1 (Target of EGR1) in Hepatocellular Carcinoma (HCC) progression. TOE1, a deadenylase belonging to the DEDD exonuclease superfamily, is primarily localized in the Cajal bodies of the nucleus. While the biological functions of TOE1 have been partially characterized, its role in cancer remains unclear. We analyzed TOE1 expression, its correlation with clinical pathological features, prognosis, and immune infiltration in HCC using databases and platforms such as The Cancer Genome Atlas (TCGA), COSMIC, cBioPortal, and TIMER. Additionally, we validated TOE1 expression levels in clinical samples. Through the knockdown of TOE1 in HCC cell lines and subsequent RNA-seq analysis, we explored its functional role and potential molecular mechanisms in HCC malignant progression. The research focuse on uncovering the role of TOE1 in HCC development and its underlying mechanisms. Comprehensive bioinformatics interrogation revealed that TOE1 is significantly upregulated in HCC and correlates with poor patient prognosis. It may influence tumor development by modulating the stemness of tumor cells and immune cell infiltration, thereby reshaping the tumor microenvironment. Furthermore, in vitro experiments suggested that TOE1 promotes HCC proliferation and metastatic potential via modulating the Hippo signaling pathway. This research highlights the pivotal role of TOE1 in HCC, indicating its promise as a novel target for early detection, therapeutic strategies, immunological intervention, and prognosis assessment in HCC. These findings provide fresh perspectives for precision medicine in the context of HCC.

overall_design: To investigate the functional role of TOE1 in HepG2 cells, we performed transient siRNA-mediated knockdown of TOE1.

first: Jie Yu

last: Ao

first: Si Ting

last: Xu

first: Bo Yu

last: Yang

first: Xian

last: Sun

first: Yan

last: shi


title: Transcriptomic profiling of a commensal Escherichia coli lysogen in simulated intestinal fluid reveals broad changes in both core and accessory regions of the genome.

city: Edmonton

first: Kat

last: Pick

address: 116 St & 85 Ave NW

country: Canada

institute: University of Alberta

summary: Escherichia coli is a diverse species, ranging from commensal strains that are part of the gut microbiome, to virulent strains that cause a variety of different infections. Much of this genomic diversity is attributable to horizontally-acquired elements, including bacteriophages. RNA sequencing has helped to reveal how pathogenic and laboratory-adapted strains respond to the gut environment, but relatively few studies have been conducted in commensal strains. Further, traditional transcriptomic data analysis techniques are not tailored to capture changes in horizontally-acquired elements, leaving them relatively understudied. Here we performed RNA sequencing on commensal E. coli MP1, which was recently isolated from mice. We analyzed the transcriptomes of MP1 grown in standard laboratory conditions or in simulated intestinal fluid (SIF), in strains that either carry or lack the vB_EcoP_Kapi1 prophage. We find that both lysogeny and growth media cause broad transcriptional changes in MP1, with growth media playing a larger role. Functional enrichment analysis revealed up-regulation of aerobic respiration, purine biosynthesis, and xenobiotic export, and down-regulation of flagellar motility and chemotaxis, acid stress response, fatty acid beta oxidation, and transport processes in SIF. In Kapi1 lysogens, transport processes were also down-regulated. Hypothesizing that horizontally-acquired genes may have been missed by traditional functional enrichment analyses, we next performed an in-depth analysis of these elements. Some genes on all identified elements were expressed, and many were differentially expressed; sugar uptake and contact-dependent inhibition systems were down-regulated in SIF, while the pathogenicity island PAI-X and multiple dynamin family proteins were strongly up-regulated in Kapi1 lysogens. Finally, we analyzed the Kapi1 transcriptome, revealing expression of λ-like regulatory genes and a putative novel accessory gene during lysogeny. Our results demonstrate that growth media and lysogeny have profound impacts on the host transcriptome. Many of the processes differentially regulated in SIF have been previously shown to be differentially regulated in the gut. In Kapi1 lysogens, the only genes consistently up-regulated were encoded by horizontally-acquired elements, suggesting cross-talk. Importantly, these genes were completely missed in traditional functional enrichment analyses, highlighting a significant gap in our knowledge of the transcriptional landscape of horizontally-acquired elements.

overall_design: RNA-seq profiling of Escherichia coli KP7 and MP13 grown to mid-log phase in LB or simulated intestinal fluid (SIF). KP7 and MP13 are isogenic derivatives of the commensal strain MP1; KP7 is mcherry-tagged and lacks the Kapi1 prophage, while MP13 is gfp-tagged and carries the Kapi1 prophage

first: Tracy

last: Raivio


title: A Dual-Function Genomic Region Links SCRT2 to ISLET1 repression in dI3 interneuron precursors

city: São Paulo

first: Irene

last: Yan

state: São Paulo

address: Av Prof Lineu Prestes

country: Brazil

institute: University of Sao Paulo

summary: How transcriptional programs coordinate the transition from neural progenitors to lineage-committed neurons in the spinal cord remains poorly understood. While much is known about transcription factors acting in the proliferative and differentiated zones, the role of intermediate zone (IZ) factors during lineage specification is less clear. Here, we investigate the function of SCRATCH2 (SCRT2), a postmitotic repressor expressed in the IZ, during dorsal interneuron differentiation. Chromatin profiling revealed that SCRT2 primarily binds to intergenic, transcriptionally inactive regions near neurogenic genes. Among these, we identified a conserved regulatory element, ECR4, located between ISLET1 and PARP8. Functional assays showed that ECR4 drives neural transcription and is composed of two subregions: ECR4B, an enhancer activated by ISLET1 and POU4F1, and ECR4A, which contains SCRT2 binding motifs and mediates transcriptional repression. Mutation of the vCES-box, a predicted SCRT2-binding motif within ECR4A, abolished repression, confirming a repressive regulatory interaction. Overexpression of SCRT2 in vivo reduced the number of ISLET1+ dorsal interneurons, supporting a model in which SCRT2 represses ISLET1 through ECR4 to modulate dI3 lineage specification. These findings identify a novel regulatory mechanism linking intermediate zone transcriptional repression to dorsal interneuron development in the spinal cord.

overall_design: CUT&RUN and ATAC-seq of HH25 chick neural tubes.

first: Vitória

last: Botezelli

first: Ee

last: Liau

first: Carolina

middle: P

last: Goes

first: Tatiane

middle: Y

last: Kanno

first: Shirley

middle: C

last: Anticona

first: Ana

last: Azambuja

first: Marcos

last: Simoes-Costa

first: Chao Yun

middle: I


title: Transcriptome analysis of Mus musculus skin from experimental studies on grape seed extract NSPCC1 intervention in skin aging

city: Guangzhou

first: Zhaoyun

last: Xue

address: Second of the Zhongshan Road

institute: Sun Yat-sen University

summary: To investigate the effects of Grape Seed Extract NSPCC1 on D-galactose combined with UVA/B-induced skin photoaging model and related transcription factors in C57BL/6J mice.

overall_design: RNA-seq profiling of control,D-galactose combined with UVA/B-induced skin aging model and NSPCC1 treated the skin of mice

first: Zhen

last: Zhang

first: Yufang

last: Wang

first: Yihan

first: Jianwen

last: Chen

first: Ruikun

last: He


title: In Vivo Modeling of Human γδ T Cell Ontogeny Reveals Terminal Deoxynucleotidyl Transferase as a Key Regulator of Type 3 Vδ2 T Cell Development

city: GOSSELIES

first: Abdulkader

last: AZOUZ

address: Rue Adrienne Bolland, 8

country: Belgium

institute: Université Libre de Bruxelles

summary: Profound differences in T cell receptor (TCR) repertoire and functional profiles between human and murine γδ T cells pose significant challenges for translational γδ T cell research. Therefore, we generated humanized immune system (HIS) NBSGW (NOD,B6.PrkdcscidIl2rγ-/-KitW41/W41) mice reconstituted with human fetal liver CD34+ hematopoietic stem and progenitor cells (HSPCs) enabling human γδ T cells evaluation in vivo. The HIS mice accurately recapitulated the TCR-associated thymic programming of human γδ T cells—alongside αβ T cells—and their peripheral effector functions, including the generation of phosphoantigen-reactive Vγ9Vδ2 T cells uniquely found in humans. Moreover, terminal deoxynucleotidyl transferase (TdT) was identified as a key regulator of type 3 Vδ2 T cell development. These findings demonstrate that HIS mice are a powerful model to screen human γδ T cell-targeting immunotherapies and to obtain mechanistic insights in human γδ T cell biology.

overall_design:
γδ and αβ T cells were sorted from the thymi and peripheral organs of humanized immune system (HIS) mice transplanted with human HSPCs, followed by 5'-RNA single cell transcriptome, ADT and TCR sequening (10x genomics).
In parallel, after knocking-out (KO) DNTT in the input HSPCs of HIS mice by CRISPR, γδ and αβ T cells were sorted from the thymi and spleens of control or KO HIS mice, followed by 5'-RNA single cell transcriptome, ADT and TCR sequening (10x genomics).

first: Céline

last: La

first: Aurélie

last: Detavernier

first: Maria

last: Papadopoulou

first: Guillem

last: Sanchez Sanchez

first: Vincent

last: Martens

first: Séverine

last: Thomas

first: Muriel

last: Nguyen

first: Valérie

last: Acolty

first: Maxime

last: Melchior

first: Desiree

last: Venturoli

first: Bérengère

last: de Toeuf

last: Azouz

first: Moosa

last: Rezwani

first: Yohannes

last: Tafesse

first: Isoline

last: Verdebout

first: Anthony

last: Rongvaux

first: Lena

last: Boehme

first: Tom

last: Taghon

first: Koen

last: Venken

first: Dirk

last: Elewaut

first: Stanislas

last: Goriely

first: David

last: Vermijlen


title: Gonadal and chromosomal sex effects on hepatic transcriptome

city: Los Angeles

first: Karen

last: Reue

state: CA

address: 695 Charles E. Young Drive South

institute: University of California, Los Angeles

summary: The Four Core Genotypes (FCG) mouse model independently segregates gonadal development and sex chromosomes, which allows for the impact of gonadal and chromosomal sex to be evaluated. We evaluated the impact of biological sex on hepatic transcriptome in the FCG mice with intact gonads and gonadectomized after a high-fat diet.

overall_design: High-throughput sequencing of liver total RNA from Four Core Genotypes (FCG) mice with intact gonads and gonadectomized (GDX) fed a HFD for 11 wks and 15 wks, respectively.

first: Carrie

last: Wiese


title: Acute myeloid leukemia microenvironment impairs neutrophil maturation and function through NFκB signaling [ATAC-seq]

city: Birmingham

first: Paran

last: Goel

state: Alabama

address: Department of Hematology and O, 701 20th Street South

institute: University of Alabama, Birming

summary: Acute myeloid leukemia (AML), an aggressive hematological malignancy, is driven by oncogenic mutations in stem and progenitor cells that give rise to AML blasts. While these mutations are well-characterized, their impact on healthy hematopoiesis—those blood cells exposed to AML but not mutated—has not been well-characterized. As the marrow is the major site for granulopoiesis, neutrophils are heavily influenced by AML pathobiology. Indeed, most AML patients report neutropenia, rendering them susceptible to infections. However, since AML studies use peripheral blood mononuclear cells devoid of neutrophils, the characterization of neutrophil dysfunction remains poorly understood. To investigate AML-exposed neutrophils, a pre-clinical AML mouse model was used where primary leukemic cells were transplanted into non-irradiated neutrophil reporter (Ly6G- tdTomato; Catchup) hosts. Neutrophils could not completely mature, suggesting impaired granulopoiesis. Single- cell transcriptomics of AML-exposed neutrophils revealed higher inflammation signatures and expression of CD14, an inflammatory marker. To address the factors contributing to this biology, an ex vivo cytokine screen was performed on marrow neutrophils and identified that NFκB signaling drove CD14 expression. AML-exposed neutrophils displayed widespread chromatin remodeling, and de novo motif discovery predicted increased binding sites for CCAAT-enhancer-binding proteins (C/EBPs) and Interferon regulatory factors (IRFs). Moreover, AML-exposed neutrophils inhibited T-cell proliferation, highlighting their immune-suppressive capability. Finally, similar biology of immature, inflammatory neutrophils was found in AML patients, again indicating dysregulated granulopoiesis. Collectively, these data show that AML-associated inflammation alters neutrophil granulopoiesis, impairs neutrophil function, and drives immunosuppression, thus contributing to patient susceptibility to infection.

overall_design: ATAC seqencing was performed with tdTOM+ neutrophil subpopulations (50000) were sorted from AML (Meis1-HoxA9 overexpression) mice and controls.


title: Acute myeloid leukemia microenvironment impairs neutrophil maturation and function through NFκB signaling [scRNA-seq]

summary: Acute myeloid leukemia (AML), an aggressive hematological malignancy, is driven by oncogenic mutations in stem and progenitor cells that give rise to AML blasts. While these mutations are well-characterized, their impact on healthy hematopoiesis—those blood cells exposed to AML but not mutated—has not been well-characterized. As the marrow is the major site for granulopoiesis, neutrophils are heavily influenced by AML pathobiology. Indeed, most AML patients report neutropenia, rendering them susceptible to infections. However, since AML studies use peripheral blood mononuclear cells devoid of neutrophils, the characterization of neutrophil dysfunction remains poorly understood. To investigate AML-exposed neutrophils, a pre-clinical AML mouse model was used where primary leukemic cells were transplanted into non-irradiated neutrophil reporter (Ly6G-tdTomato; Catchup) hosts. Neutrophils could not completely mature, suggesting impaired granulopoiesis. Single-cell transcriptomics of AML-exposed neutrophils revealed higher inflammation signatures and expression of CD14, an inflammatory marker. To address the factors contributing to this biology, an ex vivo cytokine screen was performed on marrow neutrophils and identified that NFκB signaling drove CD14 expression. AML-exposed neutrophils displayed widespread chromatin remodeling, and de novo motif discovery predicted increased binding sites for CCAAT-enhancer-binding proteins (C/EBPs) and Interferon regulatory factors (IRFs). Moreover, AML-exposed neutrophils inhibited T-cell proliferation, highlighting their immune-suppressive capability. Finally, similar biology of immature, inflammatory neutrophils was found in AML patients, again indicating dysregulated granulopoiesis. Collectively, these data show that AML-associated inflammation alters neutrophil granulopoiesis, impairs neutrophil function, and drives immunosuppression, thus contributing to patient susceptibility to infection.

overall_design: CatchUp mice were transplanted with Meis1-HoxA9 leukemic cells or CD45.2+ control bone marrow cells. Single-cell RNA-sequencing was performed 14 days after, when animals developed acute myeloid leukemia. Bone marrow neutrophils from (4) control and (4) AML mice were isolated by fluorescence-activated cell sorting (FACS) and analyzed by scRNA-seq using 10x genomics platform. In parallel, we sorted bone marrow neutrophils from healthy CatchUp mice and treated them with either GCSF, GCSF + GMCSF, or GCSF + GMCSF + IKK16 for 24 hours. We subsequencly sorted neutrophils from each condition (n=2/condition) and analyzed by scRNA-seq using 10x genomics platform.


title: Stage-specific microRNA profiling of FACS-sorted Math1-positive commissural neurons in developing chick embryos

city: Toledo

first: Bhakti

middle: Ajit

last: Khot

state: Ohio

address: 3050 W Towerview Blvd, Wolfe Hall Room 3238

department: Dept of Molecular, Cellular & Developmental Biology

institute: University of Toledo

type: Non-coding RNA profiling by high throughput sequencing

summary: This study aims to systematically identify miRNAs and characterize microRNA (miRNA) expression in a specific population of Math1-positive commissural neurons in the developing chick spinal cords. To achieve this, chick embryos were electroporated with constructs encoding a Venus-tagged, enhancer-driven Math1, followed by FACS isolation of Venus-Math1-positive neurons from developing spinal cords at pre-crossing (HH18–20) stages. By performing miRNA sequencing on these sorted populations, the study seeks to uncover stage-specific miRNA expression signatures that may play critical roles in the differentiation, identity, and maturation of Math1-positive commissural neurons during spinal cord development.

overall_design: To identify miRNAs and explore expression profiles of miRNAs from Math1-positive neurons harvested at the precrossing HH 18-20 stage of chicken spinal cord

first: Guofa

last: Liu

first: Harindi

last: Suriyaarachchi