title: In Vivo ectopic expression of OVOL2 and HNF1A in the context of PDA

first: Lorenzo

last: Tomassoni

summary: We tested whether ectopic expression of OVOL2 and HNF1 either individually or in combination—the gene pairs inducing the most efficient MOS → GLS transdifferentiation in vitro—could recapitulate MOS → GLS cell state transdifferentiation in vivo

overall_design: Plasmids containing ORF constructs for OVOL2, HNF1A, and the OVOL2-HNF1A as well mCherry (identical to those used for the in vitro assays) were lentivirally transduced into morphogenic KP4 cells. Puromycin-selected cells were then pooled and injected into athymic mice (n = 3). Animals were sacrificed at 14 days following injection and mCherry positive cells were scRNA-seq profiled

first: Pasquale

last: Laise

first: Mikko

last: Turunen

first: Alvaro

last: Garcia

first: H

last: Maurer

first: Ela

last: Elyada

first: Bernhard

last: Schmierer

first: Jeremy

last: Worley

first: Jordan

last: Kesner

first: Xiangtian

last: Tan

first: Ester

last: Fernandez

first: Xue

last: Yuanqing

first: Andrea


title: Human PDA Cell Lines: single cell RNA-Sequencing in nomral versus serum-free media

status: Public on Jan 16 2026

city: New York

address: 1130 St Nicholas Avenue

country: USA

institute: Columbia University

type: Expression profiling by high throughput sequencing

summary: Six Human PDA Cell Lines (KP4, Miapaca2, Panc1, Patu8902, PK45H, SW1990) were cultured in both normal and serum-free media conditions and then were subjected to Single-Cell RNA Sequqncing.

overall_design: Six human PDA cell lines were seeded in a 6-well plate (Falcon, ref. 353046) using recommended media. 4h later, media was changed or replaced for serum-free media and culture them for 48h. Cells were then processed for single cell RNAseq using Total Seq B antibodies and the recommended protocol from Biolegend

middle: C

last: Worely

first: Chen

last: Yining

first: Kelly

last: Wong

first: Urszula

middle: N

last: Wasko

first: Somnath

last: Tagore

first: Alexander

middle: L

first: Sabrina

last: Ge

first: Alina

last: Iuga

first: Aaron

last: Griffin

first: Winston

first: Gulam

middle: A

last: Manji

first: Mariano

middle: J

last: Alvarez

first: Faiyaz

last: Notta

first: David

last: Tuveson

first: Kenneth

middle: P

last: Olive

last: Califano


title: Propofol Differentially Modulates Neurodevelopmental Processes in Region-Specific Human Brain Organoids

city: Zunyi

first: Hong-Qing

last: She

address: 6 Xuefu West Road, Honghuagang District

country: China

institute: Zunyi Medical University

summary: Background: Propofol, a commonly used anesthetic, has controversial effects on fetal neurodevelopment. This study explores the hypothesis that propofol exerts region- and time-specific effects on distinct brain regions, with implications for clinical safety during pregnancy. Methods: Human dorsal forebrain (hCS) and ventral forebrain (hSS) organoids were exposed to 20 μM propofol for 6 hours on day 11 (D11). Morphological changes, including diameter and surface area, were analyzed at D11, D18, and D25. Immunofluorescence and PCR analyses were conducted at early (D18/25), middle (D40), and late stages (D80) post-intervention. Electrophysiological analyses were performed from D40 to D70, and bulk RNA-seq analyses were conducted on organoids at D18 to assess transcriptional changes. Results: Propofol exhibited region-specific effects on hCS and hSS organoids. In hCS organoids, excitatory neuron markers were consistently reduced at all stages, while in hSS organoids, inhibitory neuron markers were transiently affected at early stages but recovered over time. Electrophysiological analysis revealed increased action potential incidence in hCS organoids, indicating enhanced neuronal excitability despite decreased excitatory neuron markers. Transcriptomic analysis demonstrated metabolic reprogramming, including improved oxygen supply and a shift toward efficient aerobic pathways, alongside modulation of developmental signaling pathways. Conclusions: Propofol exerts region- and time-specific effects on neural development, involving both neuronal maturation and metabolic adaptation. These findings emphasize the need for careful evaluation of anesthetic safety in pregnant women and its potential long-term neurodevelopmental consequences.

overall_design: Human induced pluripotent stem cells (hiPSCs) were differentiated into ventral forebrain organoids and dorsal forebrain organoids, and subjected to propofol exposure on day 11 (D11).  Organoids without drug treatment were used as controls.  RNA-seq was performed to analyze gene expression profiles, with all samples collected on day 18 (D18) in triplicate.

first: Chang-Le

last: Fang

first: Qi-Jun

last: Li

first: Ruo-Lan

last: Du

first: Ke-Qian

last: Liu

first: Qiu-Xia

last: Xiao

first: Xiao-He

last: Tian

first: Wen-Yuan

last: Wang

first: Liu-Lin

last: Xiong


title: Stem cell-like CD8+ T cells lacking PD-1 adapt to chronic stimulation by reducing TCR signaling and self-renewal capacity [scRNA-seq]

city: Lausanne

first: Tania

last: Wyss

address: Rue du Bugnon 25A

country: Switzerland

institute: SIB Swiss Institute of Bioinformatics

summary: CD8+ T cells responding to chronic viral infection adapt to continuous antigenic stimulation by the upregulation of inhibitory receptors. Here we addressed how memory-like CD8+ T cells (TML), which sustain the immune response in chronic infection due to their stem cell-like properties, adapt to chronic stimulation when they cannot express the co-inhibitory receptor PD-1. We found that PD-1 deficient TML cells did not show evidence of a compensatory increases in the expression of other inhibitory receptors or exhaustion-related genes. Rather, these cells displayed a reduced ability to activate multiple signaling pathways downstream of the TCR. By-passing proximal TCR signaling events restored the activation of PD-1-deficient TML cells. Consistent with reduced TCR signaling, PD-1-deficient TML cells had a reduced ability to expand and self-renew in response to recall stimulation. Transient blockade of PD-1/PD-L1 interaction similarly reduced the stemness of TML cells. Thus, in the absence of PD-1, stem-like CD8+ T cells adapt to chronic stimulation by reducing the capacity of the TCR to transmit activating signals, which limits the stemness of these cells. Thus, PD-1 preserves the stemness of TML cells ensuring the long-term maintenance of the immune response to chronic infection.

overall_design: Mouse P14 cells were flow sorted from the spleen of Vbeta5 hosts (CD45.1) transferred with WT or PD-1 KO Tcf7GFP P14 cells at d28 post LCMV infection. WT P14 cells were sorted from hosts infected with either Armstrong (Arm) or clone13 (cl13), while PD-1 KO Tcf7GFP P14 cells were exposed to cl13 only.  The sorted cells were restimulated in vitro with plate bound anti-CD3/28 antibodies for 4 h (stimulated) or were cultured without activation (rested) and then subjected to scRNAseq analysis. Rested and stimulated cells derived from the same donor mouse.

first: Werner

last: Held

first: Julia

middle: M

last: Maier

first: Melanie

last: Charmoy


title: Single-Cell Barcoded Lineage Tracing in KP4 PDA Cell Line

first: Lorenzo

last: Tomassoni

summary: The KP4 PDA cell line was used to experimentally assess cross-lineage plasticiyty between different states of Pancreatic Malignant Cells

overall_design: Cells were transduced with an average of ~3 million random 27-nucleotide barcodes and were subjcted to single-cell RNA-Seq at two different time points, named as T0 and T1 (10 population doubligins).

first: Pasquale

last: Laise

first: Mikko

last: Turunen

first: Alvaro

last: Garcia

first: H

last: Maurer

first: Ela

last: Elyada

first: Bernhard

last: Schmierer

first: Jeremy

last: Worley

first: Jordan

last: Kesner

first: Xiangtian

last: Tan

first: Ester

last: Fernandez

first: Xue

last: Yuanqing

first: Chen

last: Yining

first: Kelly

last: Wong

first: Urszula

last: Wasko

first: Somnath

last: Tagore

first: Alexander

first: Sabrina

last: Ge

first: Alina

last: Iuga

first: Aaron

last: Griffin

first: Winston

first: Gulam

last: Manji

first: Mariano

last: Alvarez

last: Notta

first: Andrea


title: In vitro arrayed OVOL2 overexpression in PDA Cell Lines (bulk RNA-Seq)

first: Lorenzo

last: Tomassoni

summary: In this assay OVOL2 was overexpressed in 3 Morphogenic PDA cell lines to investigate its ability to repgram cells toward the Lineage subtype.

overall_design: Unperturbed samples of HPAFII, KP4, PATU8988S, PK45H and PANC1 cell lines were sequenced. Other samples of KP4, PANC1 and PK45H cell lines were sequenced as well following OVOL2 overexpression

first: Pasquale

last: Laise

first: Mikko

last: Turunen

first: Alvaro

last: Garcia

first: H

last: Maurer

first: Ela

last: Elyada

first: Bernhard

first: Jeremy

last: Wang

first: Faiyaz

first: David

last: Tuveson

first: Kenneth

last: Olive

last: Califano


title: Single-Cell Barcoded Lineage Tracing in HS766T PDA Cell Line

summary: The HS766T PDA cell line was used to experimentally assess cross-lineage plasticiyty between different states of Pancreatic Malignant Cells

last: Schmierer

last: Worely

first: Jordan

last: Kesner

first: Xiangtian

last: Tan

first: Ester

last: Fernandez

first: Xue

last: Yuanqing

first: Chen

last: Yining

first: Kelly

last: Wong

first: Urszula

last: Wasko

first: Somnath

last: Tagore

first: Alexander

first: Sabrina

first: Alina


title: Dnmt3a-dependent de novo DNA methylation enforces lineage commitment and preserves functionality of memory Th1 and Tfh cells [RNA-seq]

city: Salt Lake City

first: Jeffrey

middle: Scott

last: Hale

state: Utah

address: 15 North Medical Drive

department: Pathology

institute: University of Utah

summary: Following acute viral infection, naïve CD4+ T cells differentiate into T follicular helper (Tfh) and T helper 1 (Th1) cells that generate long-lived memory cells. However, it is unclear how memory Tfh and Th1 cells maintain their lineage commitment. We demonstrate that Tfh and Th1 lineages acquire distinct Dnmt3a-dependent de novo DNA methylation programs that are preserved into memory.  Dnmt3a deletion impairs lineage commitment and functionality of memory Th1 and Tfh cells, resulting in aberrant Runx1 upregulation that represses germinal center Tfh cell differentiation. In contrast, transient pharmacological DNA methyltransferase inhibition during priming impairs repression of Tfh-associated genes while properly silencing Runx1, and results in enhanced Tfh cell functionality in primary and secondary responses to viral infections. Together, these findings demonstrate that Dnmt3a-mediated epigenetic programing is required to enforce T helper lineage commitment and preserve Tfh and Th1-specific functions during the recall response to infection, and reveal novel strategies to improve long-lived adaptive immunity against infectious diseases.

overall_design: Mouse antigen specific WT and Dnmt3a KO CD4+ T cells (SMARTA TCR Transgenic) were sorted from spleens of  LCMV Armstrong infected mice at 7 days post infection (effector Th1 and effector Tfh samples) for RNA sequencing

first: Scott

first: Bryant

last: Perkins


title: In Vivo pooled CRISPRko screens in PDA Cell Lines

first: Lorenzo

last: Tomassoni

summary: We tested the essentiality of PDA state-specific Master Regulators in vivo by transducing a sgRNA library targeting 93 regulatory proteins into Cas9 expressing KP4 and HPAFII human cell lines. Cells were then injected into immunocompromised mice for genomic sequencing.

overall_design: To confirm the essentiality of state-specific MRs in vivo, we transduced a sgRNA library targeting 32 GLS/Lineage and 61 MOS/Morphogenic MRs into Cas9 expressing morphogenic KP4-and lineage HPAFII-cell lines, as representatives of the MOS and GLS state, respectively. All these genes were targeted with 5 sgRNAs / gene. For assay controls, we added 6 core-essential genes as strong positive controls, 100 negative intergenic controls, and 5 non-targeting controls. After puromycin selection, cells were injected into seven immunocompromised mice for each cell line and tumor formation was monitored for 29 days, followed by genomic sequencing

first: Pasquale

last: Laise

first: Mikko

last: Turunen

first: Alvaro

last: Garcia

first: H

last: Maurer

first: Ela

last: Elyada

first: Bernhard

first: Jeremy

last: Worley

last: Ge

last: Iuga

first: Aaron

last: Griffin

first: Winston

first: Gulam

last: Manji

first: Mariano

last: Alvarez

first: Faiyaz

last: Notta

first: Andrea


title: Single-Cell Barcoded Lineage Tracing using KP4 PDA Cell Line - Pre and Post Drug Treatment with Trametinib

summary: The KP4 PDA cell line (Untreated, DMSO-Treated and Trametinib-Treated) was used to experimentally assess plasticity between MAPK-Positive and MAPK-Negative states of Pancreatic Malignant Cells

overall_design: Cells were transduced with an average of ~3 million random 27-nucleotide barcodes and were subjected to single-cell RNA-Seq at two different time points, named as T1 and T2 (10 population doublings). Cells were either Untreated, treated with 0.15% DMSO for 24h or treated with Trametinib in 0.15% DMSO for 24h.

last: Schmierer

first: Jordan

last: Kesner

first: Xiangtian

last: Tan

first: Ester

last: Fernandez

first: Xue

last: Yuanqing

first: Chen

last: Yining

first: Kelly

last: Wong

first: Urszula

last: Wasko

first: Somnath

first: Alexander

last: Wang

first: Sabrina

first: Alina

first: David

last: Tuveson

first: Kenneth

last: Olive

last: Califano


title: TF overexpression assay in PDA cell lines (Bulk RNA-Seq)

first: Lorenzo

last: Tomassoni

summary: In this study the top Lineage Master Regulators of PDA were overexpressed in the KP4 Morphogenic cell line to study their reprogramming effect

overall_design: Unperturbed samples of KP4, PATU and HPAFII cell lines were sequenced. Other samples of KP4 cell line were sequenced as well, perturbing the Lineage Master Regulators one at a time.

first: Pasquale

last: Laise

first: Mikko

last: Turunen

first: Alvaro

last: Garcia

first: H

last: Maurer

first: Ela

last: Elyada

first: Bernhard

first: Jeremy

last: Worley

last: Tagore

last: Ge

first: Aline

last: Iuga

first: Aaron

first: Winston

first: Gulam

last: Manju


title: CRISPR screening in PDAC cell lines

summary: The aim of this study is to investigate the role of all human TFs and coTFs in the status of six PDAC cell lines

overall_design: A gene list including all human TF coTF genes (PMID: 27322546), 50 added core-essential genes (PMID: 28655737) as strong positive controls, and altogether 643 negative control genes comprising known non-essential genes (PMID: 24987113) and genes used as negative control in previous studies (PMID: 30202008). All these genes were targeted with 4 sgRNAs / gene.

last: Schmierer

first: Jordan

last: Kesner

first: Xiantian

last: Tan

first: Ester

last: Fernandez

first: Xue

last: Yuanqing

first: Chen

last: Yining

first: Kelly

last: Wong

first: Urszula

first: Somnath

first: Alexander

last: Griffin

last: Manji

first: Mariano

last: Alvarez

first: Faiyaz

last: Notta

first: Andrea


title: The RNA-binding protein NOVA-1 regulates circRNA expression, alternative splicing, and aging in C. elegans

status: Public on Jan 15 2026

city: Reno

first: Emmanuel

last: Adeyemi

state: NV

address: 1664 N. Virginia Street,

department: Biology

institute: University of Nevada Reno

type: Non-coding RNA profiling by high throughput sequencing

summary: Circular RNA (circRNA) biogenesis is regulated by RNA-binding proteins (RBPs) that alter back-splicing of exons in protein coding genes. However, few in vivo roles for RBPs in the regulation of circRNA biogenesis have been characterized. In this study, we performed total RNA-seq on wildtype and RBP mutant, nova-1. Our findings indicate that the RBP, nova-1, regulates circRNA expression,  alternative splicing, and aging in C. elegans

overall_design: In this project, total RNA-Sequencing was performed to profile circular RNA expression and linear mRNA splicing differences between C. elegans wild-type and NOVA-1 mutant animals. Total RNA was extracted from Day-3 adult worms using the Purelink RNA mini-kit, followed by a DNase I treatment following the manufacturer's protocol. RNA integrity number ranged from 9.6 to 10.0. Samples were stored at -80C prior to RNA-sequencing.

first: Hussam

last: Alshareef

first: Jaffar

last: Bhat

first: Pedro

last: Miura

last: van der Linden


title: Tumor Cell Clustering Enhances Metastatic Competence by Regulating the H3K36 Histone Demethylase KDM2A

city: New Haven

first: Don

last: Nguyen

state: Connecticut

address: 310 Cedar Street

institute: Yale University

summary: Disseminated tumor cells can form clusters via cell-cell adhesion, which increases their capacity to initiate metastasis. Metastatic clusters are characterized by distinct changes in transcription, suggesting that epigenetic mechanisms underlie their unique phenotypic state. By performingfunctional epigenomic studies in models of non-small-cell lung cancer, we identified the histone H3 lysine 36 (H3K36) demethylase KDM2A as being differentially required for the fitness of metastatic cell clusters. This contextual dependency on KDM2A is predicated by tumor cell-cell aggregation, which specifically induces KDM2A binding to CpG island enriched promoters. At these defined genomic loci, KDM2A maintains H3K36 monomethylation, which preferentially correlates with transcriptional activation. KDM2A directly targets oxidative phosphorylation genes and KDM2A activity is required for optimal mitochondrial respiration and apical cell junction integrity in cell clusters. Consequently, suppressing KDM2A reduces metastatic seeding and colonization in multiple organs, including in the brain. These findings reveal a chromatin regulatory mechanism by which homotypic cell communication instructs the epigenome of disseminated tumor cells to potentiate their metastatic competence.

overall_design: RNA-sequencing data of shControl and shKDM2A cells grown under control and cluster forming conditions

first: Carolyn

last: Kravitz

first: Kiran

middle: D

last: Patel

first: Emily

last: Wingrove

first: Dejian

last: Zhao

first: Tang

last: Tang

first: Sampada

last: Chande

first: Yuchen

last: Huo

first: Thomas

middle: F

last: Westbrook

first: Qin

last: Yan

middle: X


title: Oxidative Stress-Induced Immunogenic Cell Death Enhances Whole-Cell Vaccine Efficacy in a Syngeneic Pancreatic Cancer Model

city: St. Catharines

first: SoFong

middle: Cam

last: Ngan

state: ON

address: 1812 Sir Isaac Brock Way

department: Health Sciences

country: Canada

institute: Brock University

summary: RNA sequencing was performed to characterize transcriptional changes induced by sub-lethal oxidative stress in murine pancreatic cancer cells. Panc02 cells were exposed to low-dose hydrogen peroxide and compared with untreated controls. Differential gene expression analysis revealed extensive transcriptomic remodeling, including induction of stress-response programs, inflammatory and antiviral signaling pathways, chemokines involved in immune cell recruitment, and components of the MHC class I antigen presentation machinery. These data define the RNA-level response of pancreatic cancer cells to oxidative stress and provide insight into mechanisms by which stress conditions enhance tumor cell immunogenicity.

overall_design: Murine Panc02 pancreatic cancer cells were cultured in vitro and divided into two conditions: untreated control and oxidative stress treatment with hydrogen peroxide. Total RNA was extracted from biological replicates in each group and used to generate poly(A)+ RNA-seq libraries. Libraries were sequenced using paired-end Illumina sequencing. Reads were quality-controlled, aligned to the mouse reference genome, and quantified at the gene level. Differential expression analysis was conducted to identify genes and pathways altered by oxidative stress.

first: Maryam

last: Katoueezadeh

first: SoFong Cam

first: Siu Kwan

last: Sze


title: A longitudinal study of female and male mice to identify DNA methylation signatures of frailty beyond age

city: Renton

first: Dantong

last: Zhu

address: 401 Terry Ave N

institute: Institute for Systems Biology

type: Methylation profiling by genome tiling array

summary: Frailty is an age-related geriatric syndrome with largely unknown mechanisms. We conducted a longitudinal study of aging C57BL/6JNIA mice, measured frailty index and derived DNA methylation data from PBMCs. We selected frailty-related differentially methylated CpGs and determined differentially methylated regions (DMRs), focusing on both age-independent and -dependent frailty, and using both mixed-sex and sex-stratified subgroups. We propose a joint set of 925 frailty-related DMRs, perform an association study with frailty outcomes, build epigenetic frailty clocks and validate in mice with interventions. Notably, age-independent frailty DMRs are enriched in nervous and endocrine pathways, distinct from signaling and lipid metabolism pathways identified from age-dependent DMRs. We observe hypermethylation in signaling pathways and hypomethylation in lipid metabolism and cytochrome P450 pathways with frailty progression. 36 DMRs show consistent associations in validation. These findings highlight distinct epigenetic signatures underlying frailty and aging, with potential sex-specific mechanisms.

overall_design: We performed a longitudinal study of female (n = 40) and male (n = 49) C57BL/6JNIA mice at up to 5 time points, and derived a total of 315 samples that have DNAm array data and FI measurement. In order to investigate DNAm and mechanisms in naturally aging mice, we used untreated mice (female, n = 20; male, n = 25), including 153 samples, as the discovery cohort.

first: Alice

last: Kane


title: Efficient transgene-free multiplexed genome editing via viral delivery of an engineered TnpB

city: Berkeley

first: Marena

middle: Isabelle

last: Trinidad

state: CA

address: 2151 Berkeley Way

institute: Innovative Genomics Institute

summary: Virus-induced genome editing (VIGE) using compact RNA-guided endonucleases is a transformational new approach in plant biotechnology, enabling tissue-culture-independent and transgene-free genome editing. We recently established a transgene-free VIGE approach for heritable editing at single loci in Arabidopsis by delivering ISYmu1 TnpB (Ymu1) and its guide RNA (gRNA) via Tobacco Rattle Virus (TRV). Here, we greatly improved this system by devising a multiple gRNA expression system and by utilizing an engineered high-activity Ymu1 variant (Ymu1-WFR) to develop an efficient multiplexed genome editing approach.

overall_design: Bulk bacterial RNA sequencing was performed to characterize heterologous RNA expression from a defined plasmid construct in Escherichia coli. Cells were grown under identical conditions and induced with arabinose prior to RNA extraction. Each sample represents total RNA isolated from an induced bacterial culture processed through rRNA depletion and small-RNA-seq library preparation. Sequencing reads were aligned to the Ymu1 locus to assess RNA expression profiles.

first: Honglue

last: Shi

first: Kamakshi

last: Vohra

middle: I


title: The Sh3Pxd2bnee-/- mouse: an attractive model unveiling developmental features in Frank-ter-Haar Syndrome

city: Stanford

first: Julika

last: Huber

address: 257 Campus Drive

institute: Stanford University

summary:
Frank-ter Haar syndrome (FTHS) is an inherited disease associated
with variants of the SH3PXD2B gene, encoding for the podosomal
adaptor protein known as TKS4. FTHS is characterized by multiple
skeletal abnormalities, developmental delay and severe craniofacial
dysmorphology. This study provides an in-depth characterization of
the calvarial phenotype of a mouse model of FTHS and investigates
the potential underlying molecular and transcriptomic mechanisms.
The Sh3Pxd2bnee−/− mouse presents with craniofacial malformations,
AQ4 disrupted suture patterning, decreased osteoregeneration, cell
proliferation and migration both in vitro and in vivo, and impaired
podosome formation. Transcriptomic analysis revealed
downregulation of genes involved in ribosome biogenesis. Moreover,
ribosomal RNA accumulates in cell protrusions of migrating cells. We
established that the craniofacial phenotype of the Sh3Pxd2bnee−/−
mouse is governed by impaired cell migration and proliferation due
to dysfunctional podosome formation, particularly in neural crest-derived
tissues. Transcriptomic and molecular data suggest altered
ribosome-related processes, although further investigation is needed
to clarify the underlying mechanisms.

overall_design: RNA-seq profiling of RNA extracted from skulls of WT and Sh3Pxd2bnee-/- mice at postnatal day 15

first: Siddharth

last: Menon

first: Michael

last: Lopez-Torres

first: Jason

last: Guo

last: Longaker


title: Single cell data from patient-derived xenograft model of PDAC tumor.

first: Lorenzo

last: Tomassoni

summary: The aim of this study is to prove that PDX models of PDAC tumor are able to recapitulate different sub-populations of cells previously identified in human samples

overall_design: Untreated single-cells from one patient-derived PDX model were sequenced

first: Pasquale

last: Laise

first: Mikko

last: Turunen

first: Alvaro

last: Garcia

first: H

last: Maurer

first: Ela

last: Elyada

first: Bernhard

last: Schmierer

first: Jeremy

last: Worley

first: Jordan

last: Kesner

first: Xiangtian

last: Tan

first: Ester

last: Fernandez

first: Xue

last: Yuanqing

first: Chen

last: Yining

first: Kelly

last: Wong

first: Urszula

last: Wasko

first: Somnath

last: Tagore

first: Alexander

last: Wang

first: Sabrina

last: Ge

first: Alina

last: Iuga

first: Aaron

last: Griffin

first: Winston

first: Gulam

last: Manji

first: Mariano

last: Alvarez

first: Faiyaz

last: Notta

first: David

last: Tuveson

first: Kenneth

last: Olive

first: Andrea

last: Califano


title: Disruption of basal cell identity by Rank signaling drives mammary tumorigenesis

city: Madrid

first: Jaime

last: Redondo-Pedraza

state: Madrid

address: Melchor Fernandez Almagro 3, CNIO

department: Molecular Oncology

country: Spain

institute: CNIO

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Rank signaling regulates mammary gland development and epithelial cell differentiation. Rank receptor is expressed by mammary basal and luminal populations, but, unlike luminal Rank, the contribution of basal Rank signaling to MG homeostasis remains poorly studied. We have combined timely regulated basal specific Rank expression with lineage tracing models and unveiled that basal Rank signaling controls basal cell identity in postnatal mammary glands. Ectopic basal Rank disrupts basal but also luminal cell identity, resulting in aberrant luminal-like differentiation of basal cells and impaired lactogenesis. Mechanistically, overactivation of basal Rank signaling leads to basal cell lineage infidelity, illustrated by the appearance of premalignant lesions composed by a basal-derived hybrid population with alveolar features which ultimately generates basal and luminal breast adenocarcinomas. Proteomic, transcriptomic and chromatin analyses support that the loss of tumor suppressive epigenetic regulators driven by basal Rank contributes to epithelial cell dedifferentiation and tumorigenesis. The basal Rank signature generated associates to poor prognosis particularly in human adenocarcinomas of the luminal subtype stressing the clinical relevance of our findings. Interestingly, our results reinforce the idea that luminal breast tumors might originate from basal cells that have suffered a luminal-like aberrant dedifferentiation triggered by Rank signaling

overall_design: Assay for Transposase-Accessible Chromatin DNA sequencing (ATAC-seq) for mammary gland luminal and basal epithelial cells from K14-Ctrl and K14-Rank females; and a basal-luminal hybrid population found in K14-Rank mammary glands  Mice treatment pior ATAC-seq: 4 week old female mice were treated  with 1 mg/ml doxycycline + 20mg/ml sucrose, ad libitum in the drinking water for 4 weeks and treated with 50mg/ml MPA pellet   (Innovative Research of America #NP-161) from week 6 of age for 21 days.

first: Jose

last: Terrón-Bautista

first: Patricia

middle: G

last: Santamaria

first: Felipe

last: Costés-Ledesma

first: Eva

last: González-Suárez


title: Stem cell-like CD8+ T cells lacking PD-1 adapt to chronic stimulation by reducing TCR signaling and self-renewal capacity [RNA-seq]

first: Tania

last: Wyss

summary: CD8+ T cells responding to chronic viral infection adapt to continuous antigenic stimulation by the upregulation of inhibitory receptors. Here we addressed how memory-like CD8+ T cells (TML), which sustain the immune response in chronic infection due to their stem cell-like properties, adapt to chronic stimulation when they cannot express the co-inhibitory receptor PD-1. We found that PD-1 deficient TML cells did not show evidence of a compensatory increases in the expression of other inhibitory receptors or exhaustion-related genes. Rather, these cells displayed a reduced ability to activate multiple signaling pathways downstream of the TCR. By-passing proximal TCR signaling events restored the activation of PD-1-deficient TML cells. Consistent with reduced TCR signaling, PD-1-deficient TML cells had a reduced ability to expand and self-renew in response to recall stimulation. Transient blockade of PD-1/PD-L1 interaction similarly reduced the stemness of TML cells. Thus, in the absence of PD-1, stem-like CD8+ T cells adapt to chronic stimulation by reducing the capacity of the TCR to transmit activating signals, which limits the stemness of these cells. Thus, PD-1 preserves the stemness of TML cells ensuring the long-term maintenance of the immune response to chronic infection.

overall_design: We transferred wild type (WT) or Pdcd1-/- (PD-1 KO) CD8+ T cells expressing a TCR specific for the LCMV epitope gp33-41 (P14 cells) (CD45.2) into Vbeta5 mice (CD45.1) prior to infection with LCMV cl13.  Naive, TML and TEX cells were flow sorted based on their Tcf7-GFP and subjected to bulk RNAseq. The bulk RNAseq experiment also included the corresponding P14 subsets fromWT mice subjected to PD-L1 blockade. Two biological replicates were included for each sorted population.

first: Werner

last: Held

first: Julia

last: Maier

first: Melanie

last: Charmoy


first: Lorenzo

last: Tomassoni

summary: We performed Single-Cell RNA Sequencing of 7 PDA Cell Lines to characterize and quantify PDA subtypes populations

overall_design: Seven PDA cell lines (CAPAN1, HPAFII, HS766T, KP4, MIAPACA2, PANC0403, PATU8988S)  were  cultured and multiplexed together in order to generate baseline Single-Cell RNA-Sequencing from each one of them. To assess whether the confluence state of cell line would affect its protein activity profiles and, consequently, its cell state identification,  the  KP4 cell line  was profiled at high-confluence states  (KP4-HC) and  low-confluence (KP4-LC).

first: Pasquale

last: Laise

first: Mikko

last: Turunen

first: Alvaro

last: Garcia

first: H

last: Maurer

first: Ela

last: Elyada

first: Bernhard

last: Schmierer

first: Jeremy

last: Worley

first: Jordan

last: Kesner

first: Xiangtian

last: Tan

first: Ester

last: Fernandez

first: Xue

last: Yuanqing

first: Chen

last: Yining

first: Kelly

last: Wong

first: Urszula

last: Wasko

first: Somnath

last: Tagore

first: Alexander

last: Wang

first: Sabrina

last: Ge

first: Alina

last: Iuga

first: Aaron


title: Differential chromatin accessibility analysis elucidates mechanisms of coronary artery disease-associated genetic variation

city: Charlottesville

first: Clint

last: Miller

state: VA

department: Center for Public Health Genomics

institute: University of Virginia

summary: Genome-wide association studies (GWAS) have identified hundreds of coronary artery disease (CAD) associated variants; however, the regulatory mechanisms for candidate variants remain mostly unknown. In this study, we evaluated the epigenomic landscape in coronary artery segments using the Assay for Transposase Accessible Chromatin followed by sequencing (ATAC-seq), addressed how chromatin accessibility in human coronary arteries differs from healthy vs ischemic heart derived-coronary arteries, and defined how chromatin accessibility can be used to understand mechanisms of CAD GWAS results. We report 12,226 control-specific (Con) and 1,023 Ischemic-specific (Isc) differentially accessible regions (DARs) in human coronary arteries. Isc tissue samples were enriched for immune cell activation and ~300 CAD-associated variants were mapped in DARs. We also identified CAD-associated variants predicted to alter transcripion factor binding sites related to CAD. Additionally, we identified  endothelial/smooth-muscle-associated genes proximal to Con-DARs and CAD-associated genes proximal to Isc-DARs. In summary, we provide an initial analysis of chromatin accessibility in heathy and atherosclerotic human coronary artery tissue, prioritizing CAD GWAS variants for downstream investigation by altering atherosclerosis-related cis-regulatory activity, and present hypotheses for regulatory mechanisms that could impact CAD progression.

overall_design: We generated and analyzed ATAC-seq chromatin accessibility profiles in human coronary artery tissues from 119 patients stratified into categories based on atherosclerosis disease stages.

first: John

middle: T

last: Stubbs

first: Adam

middle: W

last: Turner

first: Nathan

last: Sheffield


title: Comparative multi-omics analysis reveals shared and derived mechanisms of fin and limb regeneration [ATAC-seq]

city: Baton Rouge

first: Igor

last: Schneider

state: Louisiana (LA)

address: 386 Life Sciences Building, LSU

department: Biological Sciences

institute: Louisiana State University

summary: Comparative studies of vertebrate appendages offer a powerful framework for uncovering shared components of an ancestral regeneration toolkit. Here, we employed a multi-omics comparative approach leveraging the regenerative capacity of the axolotl, zebrafish, and Polypterus senegalus, a fish capable of full fin regeneration. We identified conserved markers of proximal and distal blastema territories, shared activation of DNA damage repair, hif1a-mediated hypoxia response, and sequential activation of pro- and anti-inflammatory program. Apical epithelial ridge markers were expressed in both the wound epidermis and distal mesenchyme during limb and fin regeneration. Notably, hif4a-expressing erythrocytes were uniquely associated with proximal limb and fin amputations but not fin rays, while epidermal myoglobin expression was upregulated only in Polypterus and zebrafish fins. Genome-wide chromatin profiling identified candidate regeneration-responsive elements and a conserved enrichment for AP-1 transcription factor binding. Together, these finding identify shared and derived mechanisms of limb and fin regeneration.

overall_design: ATAC-seq profiling of Polypterus senegalus pectoral fins at homeostasis (uninjured) and at 3 dpa (days post-amputation)

first: Josane

last: F. Sousa

first: Gabriela

last: Lima

first: Louise

last: Perez

first: Hannah

last: Schof


title: Pooled TFs overexpression assay (single-cell RNAseq)

first: Lorenzo

last: Tomassoni

summary: The aim of this study is to investigate the effect of the combinatorial overexpression of the top GLS MRs in a MOS cell line (at single-cell level).

overall_design: Single-cell data from 3 PDA cell lines.

first: Pasquale

last: Laise

first: Mikko

last: Turunen

first: Alvaro

middle: C

last: Garcia

first: H

last: Maurer

first: Ela

last: Elyada

first: Bernhard

last: Schmierer

first: Jeremy

last: Worley

first: Jordan

last: Kesner

first: Xiangtian

last: Tan

first: Ester

last: Fernandez

first: Sue

last: Yuanqing

first: Chen

last: Griffin

first: Winston

first: Gulam

last: Manji

first: Mariano

last: Alvarez

first: Faiyaz

last: Notta

first: David

last: Tuveson

first: Kenneth

last: Olive

first: Andrea

last: Califano


title: scRNA-sequence of NeuroD1-astrocyte therapy post-TBI mouse

city: 广州

first: shuo

last: zhang

address: 黄埔大道西601号

institute: 暨南大学

summary: Astrocytes, the most abundant glial cells in the central nervous system (CNS), play a critical role in brain injury responses. While NeuroD1 overexpression in astrocytes has been shown to reverse glial scar formation and promote tissue repair in mouse models of traumatic brain injury (TBI), the underlying molecular mechanisms remain incompletely understood.

overall_design: To address this, we performed single-cell transcriptomic analysis on a controlled cortical impact (CCI) mouse model using recombinant adeno-associated virus (rAAV)-mediated astrocyte-specific NeuroD1 overexpression.Our findings demonstrate that NeuroD1 overexpression suppresses microglial activation and mitigates brain damage post-TBI. Single-cell RNA sequencing revealed significant shifts in astrocyte subtypes, with upregulated neuroinflammatory pathways and downregulated oxygen metabolism in TBI mice. Conversely, NeuroD1 overexpression reshaped astrocyte subpopulation dynamics, enhanced oxygen metabolism-related gene expression, and attenuated neuroinflammatory signaling. This study provides a comprehensive transcriptomic dataset detailing cellular and astrocyte subtype-specific changes following NeuroD1-mediated brain repair. These insights advance our understanding of cell fate modulation in TBI and support the development of NeuroD1-based gene therapies, offering a valuable resource for future TBI research and therapeutic strategies.

first: Shuo

last: Zhang


title: Transcriptomics Analysis of bulk RNA from liver tissues of male mice treated with high-fat diet and formoterol

city: Charleston

first: Jessica

middle: H

last: Hartman

state: South Carolina

address: 70 President St., Drug Discovery Bldg/Rm DD220

department: Biochemistry and Molecular Biology

institute: Medical University of South Carolina

summary: To establish the consequences of formoterol treatment on high fat diet-induced liver steatosis and inflammation, we isolated RNA from bulk liver tissue of male C57BL/6J mice that were treated with a high-fat diet for 20 weeks, with treatment with vehicle (DMSO) or formoterol (1 mg/kg) daily for the last 4 weeks of the diet. Mice were euthanized using CO2 and cervical dislocation prior to liver dissection.

overall_design: Samples include male mice treated with vehicle (n=4) or formoterol (n=5)

first: Brennan

middle: S

last: Winkler

first: Kristina

last: Stayer

first: Abhinav

middle: K

last: Rao

first: Ehtesham

last: Arif

first: Tsultrim

last: Mendenhall

first: Kristy

last: Thomas

first: Kylie

middle: R

last: Driggers

first: Xiaofeng

last: Zuo

first: Wayne

last: Fitzgibbon

first: Peifeng

last: Deng

first: Yanhui

last: Su

first: Yujing

last: Dang

first: Marie

last: Gerges

first: Daniel

last: Kagan

first: Vishwajeeth

last: Pasham

first: Bethany

last: Wolf

first: Don

last: Rockey

first: Joshua

last: Lipschutz


title: Transcriptomics Analysis of bulk RNA from HepaRG cells exposed to free fatty acids (oleic and palmitic acid), formoterol, or both

first: Jessica

last: Hartman

summary: To establish the consequences of formoterol treatment, fatty acid overload, and the combination in HepaRG cells, we isolated RNA from cell lysates after three days of treatment. Cells were differentiated for four weeks before treatment: for two weeks they were allowed to reach confluence in normal growth media and then they were further differentiated for the following two weeks by adding 1% DMSO to the media for one media change, and then 2% DMSO for two weeks. Cells were subsequently treated with 30 nM formoterol and/or 0.25 mM palmitic + 0.5 mM oleic acids for three days with fresh treatment each day.

overall_design: Samples include control cells treated with 1% BSA as a vehicle and cells treated with formoterol alone, free fatty acids alone, or a combination of the two. N=4 independent replicates are included for each group.

first: Brennan

last: Winkler

first: Kristina

last: Stayer

middle: L


title: CRISPRi screening in PDAC cell lines

first: Lorenzo

last: Tomassoni

summary: The aim of this study is to investigate the role of all human TFs and coTFs in the status of six PDAC cell lines

overall_design: A gene list including all human TF coTF genes (PMID: 27322546), 50 added core-essential genes (PMID: 28655737) as strong positive controls, and altogether 558 negative control genes as assay controls. All these genes were targeted with 4 sgRNAs / gene.

first: Pasquale

last: Laise

first: Mikko

last: Turunen

first: Alvaro

last: Garcia

first: H

last: Maurer

first: Ela

last: Elyada

first: Bernhard

last: Schmierer

first: Jeremy

last: Worley

first: Jordan

last: Kesner

first: Xiangtian

last: Tan

first: Ester

last: Fernandez

first: Xue

last: Yuanqing

first: Chen

last: Yining

first: Kelly

last: Wong

first: Urszula

middle: N

last: Wasko

first: Somanth

last: Tagore

first: Alexander

last: Wang

first: Sabrina

last: Ge

first: Alina

last: Iuga

first: Aaron

last: Griffin

first: Winston

first: Gulam

middle: A

last: Manji

first: Mariano

middle: J

last: Alvarez

first: Faiyaz

last: Notta

first: David

last: Tuveson

first: Kenneth

middle: P

last: Olive

first: Andrea

last: Califano


title: Blockade of T-type Ca²⁺ channels disrupts mitochondrial function and follicle development by inhibiting the PI3K/AKT pathway

city: Nantong

first: Xiuling

last: Zhao

address: NO.16 WenFeng Road,  Chongchuan District

institute: Nantong University

summary: Calcium (Ca²⁺) function as critical secondary messengers in cellular signaling, essential for maintaining ovarian function and promoting oocyte maturation. However, the contribution of T-type voltage-gated calcium channels (VGCCs) to ovarian follicle development remains unclear. To investigate this, we administered flunarizine (FNZ), a potent T-type VGCCs antagonist, to 4-6-week-old female mice via intraperitoneal injection at doses of 0, 3, or 30 mg/kg/day for 7 days. FNZ exposure significantly impaired ovarian morphology, disrupted folliculogenesis, and induced estrous cycle irregularities, concomitant with a marked reduction in serum 17β-estradiol (E2) levels. FNZ led to diminished intracellular Ca²⁺ ([Ca2+]i) levels and mitochondrial dysfunction, resulting in attenuated ATP production. Notably, ovarian tissues exhibited elevated DNA damage, as indicated by increased γH2AX expression. Transcriptomic analysis revealed pronounced alterations in fibroblast growth factor 1 (FGF1) signaling-related genes. Furthermore, FNZ inhibit activation of protein kinase B (p-AKT), an effect that was rescued by recilisib, a specific activator of phosphatidylinositol 3-kinase (PI3K)/AKT. In conclusion, our study establishes that pharmacological inhibition of T-type VGCCs disrupts hormonal secretion and follicular development in adolescent mice, likely through modulation of the PI3K/AKT signaling axis.

overall_design: FNZ was dissolved in a vehicle solution containing 50% polyethylene glycol 400 and 50% physiological saline. FNZ was administered through daily intraperitoneal injections at doses of 30 mg/kg for 7 days. Three ovaries from ICR mice were dissected for group.

first: Shiqin

last: Huang

first: Jin

last: Li

first: Ting

last: Jiang

first: Xuhui

last: Zeng

first: Hao

last: Chen

first: Liping

last: Pu

first: Junyu

last: Nie


title: 5′ UTR length regulates the production of alternative N-terminal protein isoforms in health and disease

city: Cambridge

first: Jimmy

last: Ly

state: Massachusetts

address: 455 main street

institute: Whitehead Institute

summary: The 5′ untranslated region (5′ UTR) of an mRNA is classically viewed as a regulatory region that controls the amount of protein production, but not the resulting protein sequence. Here, we demonstrate that 5′ UTR length also plays a direct role in alternative N-terminal protein isoform production by controlling start codon selection. We find that very short 5′ UTRs enhance leaky scanning, thereby promoting the production of truncated alternative N-terminal protein isoforms. Changes in 5′ UTR length due to alternative transcription initiation can tune the relative abundance of alternative N-terminal isoforms from the same gene. In addition, we identify mutations in rare genetic diseases that alter 5′ UTR length, including a deletion in the VHL 5′ UTR in von Hippel–Lindau disease that shifts translation toward the shorter VHLp19 isoform. Together, our results implicate 5′ UTR length as a determinant of alternative N-terminal isoform production and reveal an underappreciated mechanism by which noncoding mutations can reshape the proteome.

overall_design: Whole cell RNA was extracted from asynchronously growing HeLa cells and CAGE-Seq was performed on the resulting mRNA.

first: Iain

last: Cheeseman


title: Comparative multi-omic analysis reveals conserved and derived mechanisms of fin and limb regeneration [Bulk RNA-seq]

first: Igor

last: Schneider

overall_design: Bulk RNA-seq profiling of Polypterus senegalus intact and 3 dpa (days post-amputation) regenerating fins

first: Josane

middle: F

last: Sousa

first: Gabriela

last: Lima

first: Louise

last: Perez

first: Hannah

last: Schof


title: Comparative multi-omic analysis reveals conserved and derived mechanisms of fin and limb regeneration [spatial transcriptomics II]

overall_design: Cryosections of axolotl  limbs at homeostasis (intact) and at different time points of regeneration (3, 7 and 14 dpa) were used for spatial transcriptomics analysis.


title: Sequencing data of six PDA cell lines.

city: New York

first: Lorenzo

last: Tomassoni

institute: Columbia University

summary: The aim of this study is the classification of PDA cell lines according to the activity of the GLS and MOS MRs

overall_design: Untreated bulk data from six cell lines

first: Pasquale

last: Laise

first: Mikko

last: Turunen

first: Alvaro

last: Garcia

first: H

last: Maurer

first: Ela

last: Elyada

first: Bernhard

last: Schmierer

first: Jeremy

last: Worley

first: Jordan

last: Kesner

first: Xiangtian

last: Tan

first: Ester

last: Fernandez

first: Xue

last: Yuanqing

first: Chen

last: Yining

first: Kelly

last: Wong

first: Urszula

last: Wasko

first: Somnath

last: Tagore

first: Alexander

last: Wang

first: Sabrina

last: Ge

first: Alina

last: Iuga

first: Aaron

last: Griffin

first: Winston

first: Gulam

last: Manji

first: Mariano

last: Alvarez

first: Faiyaz

last: Notta

first: David

last: Tuveson

first: Kenneth

last: Olive


title: Comparative multi-omic analysis reveals conserved and derived mechanisms of fin and limb regeneration

first: Igor

last: Schneider

overall_design: snRNA-seq profiling of Polypterus senegalus regenerating fins. Single nucleus suspensions were prepared from uninjured and regenerating pectoral fins at 1, 3 and 7 dpa (days post-amputation). Regenerating fins were either from fish treated with DMSO (control) or 2.5 uM rapamycin (mTOR inhibition). Nuclei were fixed and frozen using the Evercode Nuclei Fixation Kit v3 (#NF100, #NF200) from Parse Biosciences following the manufacturer’s protocol. Fixed and frozen nuclei were then processed to generate snRNA-seq libraries following the combinatorial barcoding protocol from Parse Biosciences, using the Evercode WT v3 Kit (#WT100A-D, #WT200) as per manufacturer’s instructions.

first: Josane

last: Sousa

first: Gabriela

last: Lima

first: Louise

last: Perez

first: Hannah

last: Schof


title: CTCF-mediated cis-regulatory chromatin insulation enforces a  central B-cell tolerance checkpoint [Micro-C-TALE]

city: Jerusalem

first: Yehudit

last: Bergman

address: Ein Karem

department: Department of Developmental Biology and Cancer Research

country: Israel

institute: The Hebrew University Of Jerusalem

summary:
The generation of a diverse and self-tolerant B cell repertoire is essential for adaptive immunity and is achieved through V(D)J recombination. In mice, Igκ is the dominant light chain, whereas Igλ rearrangement typically occurs in response to nonproductive or autoreactive Igκ recombination, a process termed receptor editing.
Recombination at the RS element deletes the Igκ constant exon, silencing the locus and enabling Igλ expression. However, the epigenetic regulatory framework that orchestrates and governs receptor editing remains poorly defined. Here, we identify a CTCF-binding insulator element (CBE) within the 3′ Igκ super-enhancer (3′-SEκ) that regulates receptor editing and directs the κ-to-λ switch required for Igλ⁺ B-cell development. Mechanistically, loss of this CBE activates an insulated enhancer within the 3′-SEκ, causing aberrant Vκ rearrangements and altered chromatin interactions through disrupted loop extrusion dynamics. Notably, loss of this CBE in mice leads to increased autoantibody production by ten weeks of age, demonstrating that CBE-mediated chromatin architecture shapes B cell fate by constraining autoreactive potential. Collectively, our findings define a novel CTCF-dependent cis- regulatory insulation checkpoint that connects chromatin loop extrusion to antigen- driven receptor editing, thereby enforcing B-cell tolerance.

overall_design: Micro-C-TALE sequencing was performed on ex vivo CD19⁺ pre-B cells isolated from 6–12-week-old WT-Rag1⁻/⁻.hIgM and ΔrsCBE-Rag1⁻/⁻.hIgM mice.

last: Gomes

first: Anna

last: Golberg

first: Eliraz

last: Gitelman

first: Noam

last: Kaplan


title: CTCF-mediated cis-regulatory chromatin insulation enforces a  central B-cell tolerance checkpoint [ChIP-Seq_CTCF]

overall_design: CTCF ChIP-seq was carried out on IL-7–cultured pre-B cells isolated from 6–12-week-old WT and ΔrsCBE mice

first: Merav

last: Hechst


title: Transcriptomic profiling reveals distinct molecular signatures among lesion types in hidradenitis suppurativa

city: Aarhus

first: Hakim

last: Ben Abdallah

state: Denmark

address: Palle Juul-Jensens Boulevard 45

department: Dermatology and Venerology

country: Denmark

institute: Aarhus University Hospital

summary: This study investigated the transcriptome of the various hidradenitis suppurativa lesion types using bulk RNA sequencing.

overall_design: Three-millimetre punch biopsies were collected from draining tunnels (purulent discharge at rest or to palpation) and non-draining tunnels (no purulent discharge) from 20 hidradenitis suppurativa patients and normal skin from eight healthy donors.


title: CTCF-mediated cis-regulatory chromatin insulation enforces a  central B-cell tolerance checkpoint [ChIP-Seq_H3K27ac]

overall_design: H3K27Ac ChIP-seq was performed on ex vivo CD19⁺ pre-B cells isolated from 6–12-week-old WT-Rag1⁻/⁻.hIgM and mutant-Rag1⁻/⁻.hIgM mice.


title: Navigating high-order protein mutational landscapes via deep learning on directed evolution trajectories

city: Peking

first: George

last: Song

address: 5 Yiheyuan Road, Haidian District

institute: Peking University

summary: Accurately predicting the fitness effects of high-order mutations is a grand challenge in understanding and engineering proteins. Existing models, including pre-trained protein language models, struggle to capture the multi-residue interactions that govern these effects. Here, we introduce DENet, a deep learning framework that harnesses the rich co-mutation information within directed evolution (DE) trajectories to reconstruct high-resolution fitness landscapes for deciphering and engineering of complex protein variants. Applied to the cancer target KRAS, DENet-guided screening systematically identified high-order mutants with potent activities and uncovered hidden allosteric mechanisms. For MEK1, DENet discovered complex variants with >1,000-fold increased drug resistance, revealed synergistic tail mutations, and retrospectively identified over 75% of known clinical mutations, largely outperforming existing models. To broaden the framework’s applicability, we developed an in silico strategy that simulates directed evolution to generate crucial co-mutation information from widely available single-mutant datasets. DENet provides a quantitative framework for navigating complex fitness landscapes, uniting the rational engineering of multi-mutation proteins with the elucidation of their allosteric and clinical implications.

overall_design: High-throughput sequencing data were collected from the most highly mutated regions during the directed evolution campaigns of KRAS and MEK1 to identify and quantify mutant variants generated and selected at multiple time points.

first: Yihan

last: Lin

first: Chengzhi

first: Liang

last: Ma

first: Yingfan

last: Xu


title: Single-cell RNA-sequencing of CD45+ immune cells from a kidney transplant biopsy and matched peripheral blood

city: Houston

first: Dawei

last: Zou

state: TX

address: 6670 Bertner Ave

department: Surgery

institute: Houston Methodist Research Institute

summary: T cells are central mediators of transplant rejection, yet the differentiation trajectories that sustain this process remain incompletely defined. To delineate the differentiation trajectory of CD8+ T cells, we performed single-cell transcriptomic profiling of matched allograft and peripheral blood mononuclear cell (PBMC) samples obtained from the same patient.

overall_design: A renal allograft biopsy was obtained percutaneously under ultrasound guidance using a 14-gauge core biopsy needle. The biopsy tissue was immediately placed on ice and rapidly processed for downstream analysis. Peripheral blood mononuclear cells (PBMCs) were collected from the same subject to enable matched sequencing. CD45+ immune cells from the biopsy and PBMC sample were isolated and profiled by single-cell RNA sequencing.

first: Stephanie

last: Yi

first: Yulin

last: Dai

first: Wenhao

last: Chen


title: Toll signalling controls intestinal regeneration in Drosophila

city: Oxford

first: Petros

last: Ligoxygakis

address: South Parks Rd

department: Biochemistry

country: United Kingdom

institute: University of Oxford

summary: This study is part of a manuscript approved for publication in Development (Cambridge UK) showing that the Toll pathway in Drosophila is necessary and sufficient for intestinal stem cell mitosis following infection and under homeostatic conditions. Chronic activation of Toll using a gain of function mutation (Toll10b) induces intestinal stem cell proliferation while blocking differentiation of enteroblasts to enterocytes. Thus, the intestinal tissue becomes replete with progenitor cells. The RNA-seq data deposited here are on whole guts of 20-day old flies where Toll has been activated in intestinal progenitor cells post-developmentally (in adults) from day 1 using the GAL4/GAL80ts system.

overall_design: Whole guts were dissected from flies with the following genotypes: esgts/w1118 (control genetic background), esgts/UAS-Toll10b (overactivation of the Toll pathway), esgts/UAS-Toll10b/UAS-BskDN (blocked overactivity of the Toll pathway by a dominant negative form of JNK). Libraries were generated using the MGIEasy RNA-seq Library Preparation Universal Kit in BGI HongKong. RNA-seq was performed on the DNBSEQ-G400 using the 150 paired-end sequencing protocol. 3 samples (esgts/w1118, esgts/UAS-Toll10b, esgts/UAS-Toll10b/UAS-BskDN) with 3 independent biological repeats were used.

first: Aiswarya

last: Udayakumar

first: Qi

last: Xiao


title: A Polyamine-mediated Posttranslational Modification Required for Macrophage Tissue Residency [scRNA-seq 2]

city: Baltimore

first: Immunometabolism

last: Department

state: Maryland

address: 1650 Orleans Street

department: Immunometabolism

institute: Johns Hopkins University

summary: Tissue-resident macrophages (RTMs) form during embryogenesis, self-renew locally, and regulate tissue homeostasis by clearing dead cells and debris. During tissue damage, however, bone marrow-derived monocytes enter tissue sites and differentiate into RTM, repairing the tissue and replenishing macrophages in the niche. Universal cell-intrinsic mechanisms that control the monocyte to RTM transition remain elusive. We investigated mice with myeloid cell deletions in deoxyhypusine synthase (DHPS), an enzyme that mediates the polyamine-dependent hypusine modification of the translation factor eIF5A, and found that DHPS is required for cell adhesion and signaling programs critical for RTMs. Without DHPS expression, immature tissue macrophages form, but RTM and associated homeostatic functions are lost. Thus, the polyamine-hypusine pathway is a global, tissue-agnostic program driving the differentiation trajectory of monocyte-derived macrophages into RTM.

overall_design: Single cells were isolated from the lung of indicated animals with 3 biological replicates per genotype (KO and DHPS) and prepared for single cell RNA sequencing using the GEM-X 3' Gene Expression kit (v4) from 10X Genomics.

first: Erika

last: Pearce

first: David

middle: E

last: Sanin

first: Gustavo

last: Carrizo


title: Differential downstream signaling in microglia lacking Alzheimer’s-related TREM2 or its adaptor TYROBP/DAP12

city: Hartsdale

first: michelle

last: ehrlich

state: New York

address: 88 Mercer Ave.

institute: Mount Sinai Hospital

summary: Microglia, the primary immune cell in the brain, have multiple activation phenotypes involved in broad functions within the brain, playing roles in neurotoxicity/neuroprotection, release of inflammatory and anti-inflammatory cytokines, and in cell survival, proliferation, and phagocytosis. TREM2 and TYROBP form a transmembrane complex in microglia that modulates intracellular signaling networks, and these proteins are essential regulators of the transition from homeostatic to activated microglia. Recent findings support a TREM2-independent molecular signature that is involved in the early transition of homeostatic to disease-associated microglia (DAM), with the next sequential step of DAM activation from stage 1 to stage 2 being TREM2-dependent. However, the underlying mechanisms determining how TREM2 or TYROBP regulate these downstream phenotypes are largely unknown. We isolated primary microglia from C57BL/6 wild-type (WT) controls, Trem2 knock-out (KO), and Tyrobp KO mice at post-natal day 0-3. Cells were treated with Alzheimer’s disease (AD)-relevant stimuli, such as amyloid beta (Aβ) oligomers or fibrils, or ‘neuroinflammatory-like’ stimuli, such as lipopolysaccharide (LPS). We explored protein and gene expression in the presence or absence of inhibitors of the TREM2/TYROBP downstream signaling pathway. We also performed a high-throughput Olink proteomic analysis of conditioned media from WT, Trem2 KO, and Tyrobp KO stimulated with either LPS or Aβ oligomers or fibrils. Our results show that the absence of either TREM2 or TYROBP is associated with increased basal levels of phosphorylated ERK in primary microglia compared to WT controls. In addition, Trem2 KO and Tyrobp KO cells show a less ramified cell morphology at baseline, as compared to WT microglia. Moreover, stimulating primary microglia with either Aβ oligomers or LPS leads to differential protein and gene expression in cells lacking TREM2 or TYROBP. The dysregulated downstream signal transduction and morphology in the absence of TREM2 or TYROBP suggest their essential roles not only in microglial homeostasis but also in their activation in response to different stimuli.

overall_design: RNA-seq profiling of cortical and hippocampal primary microglia isolated from WT, Trem2 KO, and Tyrobp KO mice at baseline, or after a 24hr-treatment with LPS, A11/OC mix, or OC-only preparations.

first: Gabriela

last: Farias Quipildor

first: Ramona

last: Belfiore

first: Khaled

last: Althobaiti

first: Zahra

last: Najarzadeh

first: Charles

last: Glabe

first: Benjamin

last: Readhead

first: Sam

last: Gandy

first: Stephen

last: Salton

first: Michelle

last: Ehrlich


title: High efficiency CRISPR knock-in demonstrates that TCF1 is insufficient to reverse T cell exhaustion

city: Melbourne

first: Ian

last: Parish

state: VIC

address: 305 Grattan Street

country: Australia

institute: Peter MacCallum Cancer Centre

summary: In this study we investigated the role of TCF1 in reversing T cell exhaustion by CRISPR-HDR technology.

overall_design: C57BL/6 mice received CRISPR-edited CD45.1+ P14 cells and were infected with LCMV Clone 13. 20 days later, the CD45.1 P14 cells were isolated from the spleens of the recipient mice, sorted and analysed.

first: Maria

last: Nogueira de Menezes

first: Nicole

last: Saw


title: A Polyamine-mediated Posttranslational Modification Required for Macrophage Tissue Residency

first: Immunometabolism

last: Department

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

overall_design: Refer to individual Series


title: Effect of depletion of USP16 on gene expression of HGC-27 cells

city: Lanzhou

first: Siying

last: Ren

address: Peili Street, Anning District

department: College of Life Sciences

institute: Northwest Normal University

summary: Gastric cancer has emerged as one of the malignant tumors with increasing incidence in recent years, posing significant challenges to global health. However, effective targeted therapies for gastric cancer remain limited. Despite strict Hippo pathway regulation, the oncoprotein YAP is frequently overexpressed in gastric cancer, indicating that mechanisms beyond phosphorylation must exist to control YAP stability. In this study, we focus on the impact of YAP deubiquitination on the progression of gastric cancer. Our study identifies USP16 as a novel deubiquitinase of YAP and demonstrates it as a direct therapeutic target of ISO in gastric cancer.

overall_design: RNA-seq profling of wildtype HGC-27 cells and their knockdown derivatives(shUSP16)

first: Yehua

last: Li

first: Huanhuan

last: Chen


title: A Cellular and Transcriptomic Atlas of the Aged Mouse Hematopoietic System

city: Tarrytown

first: Kun

last: Xiong

state: NY

address: 777 Old Saw Mill River Road

department: Molecular Profiling and Data Science

institute: Regeneron Pharmaceuticals

summary: Aging is a dominant risk factor for chronic diseases characterized by the functional decline of tissues and organs. During aging, the hematopoietic system declines in regenerative capacity – seemingly attributable to increases in DNA damage, replicative stress, and autophagic flux – resulting in skewing towards a myeloid lineage and away from a lymphoid lineage. Here, we characterized the transcriptomic and cellular landscape of the aged C57Bl6/J mouse hematopoietic system using a combination of bulk RNAseq and single cell RNAseq (scRNAseq). We show that aging leads to global transcriptional alterations in bulk peripheral blood mononuclear cells (PBMCs), lineage marker-depleted bone marrow cells (lin- BM), and in hematopoietic stem cells (HSCs) marked by lineage- (lin-) Sca1+cKit+ (LSK+). These changes indicate widespread activation of inflammatory processes, namely in PBMCs and lin-BM cells. Interestingly, there is also a downregulation of cell cycle genes in HSCs during aging. scRNAseq across 39 hematopoietic cell types revealed age-related skewing in cell composition. Aged PBMCs showed significant decreases in CD4 and CD8 naïve cells concomitant with increases in CD4/8 memory and CD8 exhausted T cell populations. Lin- BM cells showed significant myeloid skewing in common myeloid progenitor (CMP) cells as well as in the HSC population. We also identified a unique HSC population marked by increased Vwf, Wwtr1, and Clca3a1 expression that does not exist in young HSCs, thus likely marking true aged HSCs. Collectively, this works encompasses a significant resource for uses in therapeutically targeting the aged hematopoietic system.

overall_design: Bulk RNAseq and scRNA seq of PBMC and lineage negative bone marrow from young and old mice. Bulk RNAseq of HSC from young and old mice

first: Ryan

last: White

first: Matthew

last: Wakai

first: Allison

last: Surian

first: Christina

last: Adler

last: Negron

first: Min

last: Ni

first: Tea

last: Shavlakadze

first: Yu

last: Bai

first: David

last: Glass


title: CTCF-mediated cis-regulatory chromatin insulation enforces a  central B-cell tolerance checkpoint [4C-Seq]

first: Yehudit

last: Bergman

overall_design: 4C sequencing was performed on v-Abl–transformed 445.3 Rag1-deficient pro-B cells from WT, rsCBE-571 bp–deleted, ΔSIS 445.3, and rsCBE-18 bp–deleted lines.

last: Gomes

first: Eliraz

last: Gitelman


title: Human microglia in brain assembloids display region-specific diversity and respond to hyperexcitable neurons carrying SCN2A mutation

city: West Lafayette

first: Harish

last: Kothandaraman

state: IN

address: 201S South State St

department: Purdue Institute for Cancer Research

institute: Purdue University

summary: Microglia critically shape neuronal circuit development and function, yet their region-specific properties and roles in distinct circuits of the human brain remain poorly understood. In this study, we generated region-specific brain organoids (cortical, striatal, and midbrain), each integrated with human microglia, to fill this critical gap.

overall_design: Single-cell RNA sequencing was performed on human iPSC-derived cortical (hCO+Mg), striatal (hStrO+Mg), and midbrain (hMO+Mg) organoids containing microglia. For each condition, 4–5 organoids derived from three independent hiPSC lines (KOLF2.1J, B07 WT, B11 WT) were pooled into a single sample and processed using the PIPseq-T20 3′ single-cell RNA-seq workflow. Libraries were sequenced on an Illumina NovaSeq S4 platform (2 × 150 bp), and the resulting data were used to compare microglial transcriptional states across distinct brain region–like environments.

first: Jiaxiang

last: Wu

first: Xiaoling

first: Weihao

first: Yang

last: Yang


title: CTCF-mediated cis-regulatory chromatin insulation enforces a  central B-cell tolerance checkpoint [RNA-seq_IGK repertoire]

overall_design: Genome-wide RNA profiling of exvivo small pre-B cells derived from C57Bl6 WT, ΔrsCBE, and ΔrpiaCBE mice of 6-12 weeks of age

first: Avidov

last: Bar


title: Single-cell RNA Sequencing of Wildtype and/or Dazl Mutant Mouse Ovarian Cells Across Embryonic Developmental Stages (E10.5 (Wild type), E11.5 (Wild type and Dazl-/-), E12.5 (Dazl-/-), E15.5 (Wild type)

first: Allan

last: Spradling

address: 3520 San Martin Dr

institute: Howard Hughes Medical Institute Research Laboratories, Department of Embryology, Carnegie Institution for Science

summary: We have identified a Golgi- and microtubule-rich vesicular structure, termed “Visham”, which resembles the Drosophila fusome and accumulates asymmetrically during germ cell cyst divisions starting at E11.5. Genes associated with Visham become enriched in presumptive oocytes during cyst divisions and early meiosis, suggesting its role as a potential oocyte rejuvenation center. To investigate this further, we performed single-cell RNA sequencing (scRNA-seq) of mouse fetal germ cells at E10.5, E11.5, and E15.5, including developmental stages not previously covered in earlier datasets (GSE136441) and thus generated a composit dataset of E10.5-P5 germ cells. Additionally, we analyzed Dazl homozygous and heterozygous mutant ovarian germ cells at E11.5 and E12.5 to study the impact of Dazl loss on cyst formation and Visham-associated gene expression dynamics. This dataset provides a comprehensive resource to explore transcriptional changes across key stages of fetal ovarian development and in the context of Dazl function.

overall_design: Embryonic ovaries were dissected, and fetal sex was identified using Quinacrine and DAPI staining of tail samples to exclude males in E10.5/E11.5 gonad samples. Female gonads were pooled and dissociated into single cells using 0.25% Trypsin at 37°C for 5–7 minutes with gentle trituration. Trypsin was neutralized with 10% FBS, and cells were filtered through a 30 μm strainer. After centrifugation and resuspension in 0.04% BSA, cell viability was assessed with trypan blue, and high-quality samples (>90% viability) were loaded (~10,000 cells) onto the 10X  (Genomics Chromium platform (v3/v4 chemistry) per manufacturer’s instructions. For mutant samples, Dazl+/- males and females were mated, and embryos collected at E11.5 and E12.5. Tail DNA was extracted using the KAPA Express Extract Kit and genotyped using standard JAX protocols (strain 035880). Identified homozygous and heterozygous fetal gonads were dissociated as above and subjected to scRNA-seq on the same 10X chromium platform.

first: Madhulika

last: Pathak


title: Comparative multi-omic analysis reveals conserved and derived mechanisms of fin and limb regeneration [spatial transcriptomics]

city: Baton Rouge

first: Igor

last: Schneider

address: 386 Life Sciences Building, LSU

institute: Louisiana State University

summary: Comparative studies of vertebrate appendages offer a powerful framework for uncovering shared components of an ancestral regeneration toolkit. Here, we employed a multi-omics comparative approach leveraging the regenerative capacity of the axolotl, zebrafish, and Polypterus senegalus, a fish capable of full fin regeneration. We identified conserved markers of proximal and distal blastema territories, shared activation of DNA damage repair, hif1a-mediated hypoxia response, and sequential activation of pro- and anti-inflammatory program. Apical epithelial ridge markers were expressed in both the wound epidermis and distal mesenchyme during limb and fin regeneration. Notably, hif4a-expressing erythrocytes were uniquely associated with proximal limb and fin amputations but not fin rays, while epidermal myoglobin expression was upregulated only in Polypterus and zebrafish fins. Genome-wide chromatin profiling identified candidate regeneration-responsive elements and a conserved enrichment for AP-1 transcription factor binding. Together, these finding identify shared and derived mechanisms of limb and fin regeneration.

overall_design: Cryosections of Polypterus pectoral fins at homeostasis (uninjured) and at different time points of regeneration (1, 3 and 7 dpa) were used for spatial transcriptomics analysis.

first: Josane

last: Sousa

first: Gabriela

last: Lima

first: Louise

last: Perez

first: Hannah

last: Schof


title: Single-cell multiomics of neuronal activation reveals context dependent genetic controls of brain disorders

city: Evanston

first: Christina

last: Thapa

state: Illinois

address: 1001 University Place

department: Center for Psychiatric Genetics (CPG)

institute: Endeavor Health

summary: Despite hundreds of genetic risk loci identified for neuropsychiatric disorders (NPD), most causal variants/genes remain unknown. A major hurdle is that disease risk variants may act in specific biological contexts, e.g., during neuronal activation, which is difficult to study in vivo at the population level. Here, we conducted a single-cell multiomics study of neuronal activation (stimulation) in human iPSC-induced excitatory and inhibitory neurons from 100 donors, and uncovered abundant neuronal stimulation-specific causal variants/genes for NPD. We surveyed NPD-relevant transcriptomic and epigenomic landscape of neuronal activation and identified thousands of genetic variants associated with activity-dependent gene expression (i.e., eQTL) and chromatin accessibility (i.e., caQTL). These caQTL explained considerably larger proportions of NPD heritability than the eQTL. Integrating the multiomic data with GWAS further revealed NPD risk variants/genes whose effects were only detected upon stimulation. Interestingly, multiple lines of evidence support a role of activity-dependent cholesterol metabolism in NPD. Our work highlights the power of cell stimulation to reveal context-dependent “hidden” genetic effects. This dataset aims to validate the cell type-specific transcriptomic effects of the key transcription factors (TFs) of the predicted autism-relevant gne regulatory networks (GRNs).

overall_design: We introduce premature stop codons in TCF4, MEF2C, and RORB via cytosine base editing system with reporter editing enrichment, causing a permanent gene knockout (KO) in iPSCs. The iPSC-derived glutmatergic and GABAergic neurons were co-cultured with primary rat astrocytes and treated with KCl to model neuronal activity. Transcriptional effects of each TF KO were assayed by scRNA-seq.

first: Duan

last: Jubao

first: He

last: Xin

first: Liang

last: Lifan

first: Zhang

last: Hanwen


title: The Ess1 prolyl isomerase represses TERRA transcription and promotes telomere replication in Saccharomyces cerevisiae

city: Shanghai

first: JUXIU

last: CHEN

state: Shanghai

address: 115 Jinzun Road, Pudong New Distract

department: Shanghai Institute of Precision Medicine

institute: Ninth People’s Hospital of Shanghai Jiao Tong University School of Medicine

summary: The conserved Ess1 prolyl isomerase (PIN1 in human) binds the carboxy-terminal domain (CTD) of RNA Pol II, and plays multiple roles in transcription regulation. Consistent with an essential role of the human PIN1 in telomere maintenance, previous screenings have identified the yeast Ess1 as a telomere length maintenance gene. Here, we provide evidence that Ess1 is involved in regulating both telomere transcription and replication. We find that depletion of Ess1 leads to a failure in transcription termination, explaining the essential role of Ess1 in maintaining a low level of telomere repeat containing RNA (TERRA). Furthermore, we show that Ess1 depletion promotes telomere shortening and accelerates senescence in telomerase-deficient cells. Notably, the depletion of Ess1 causes synthetic growth defects and telomere shortening in mre11Δ cells, and compromises rif2Δ-induced telomere elongation. Additionally, Ess1 depletion also accelerates senescence and eliminates type II telomere recombination in rad50Δ tlc1Δ cells. Lastly, Ess1 depletion decreases the accumulation of single-stranded DNA at telomere ends. These results support the model that Ess1 positively regulates both telomerase- and recombination-dependent telomere replication by promoting telomere-end resection. Taken together, this study reveals the yeast Ess1 as a new regulator of telomere transcription and replication via a distinct mechanism from the human PIN1.

overall_design: RNA-seq profiling of ESS1-AID cells treated with either DMSO(control) or 2 mM IAA(experimental) for 3 hours. Each group includes two biological replicates.

first: Juxiu

last: Chen

first: Lin

first: Xiaohan

last: Zhu

first: Jian

first: Zhenfang

first: Ming

last: Lei


title: FRMD5 as a Molecular Hub Altering GR Functions in PTSD

city: Beijing

first: Xi-Yin

last: Zhang

address: College Road

department: Neuroscience Research Institute

summary: Post-traumatic stress disorder (PTSD) is a maladaptive and debilitating psychiatric disorder with limited intervention targets. FRMD5, a scaffold protein, serves as molecular hub involved in diverse biological processes. This study elucidates a tissue-specific mechanism via shorter isoform of FRMD5, characterized by condensate formation and special interactive features, on modulation of glucocorticoid receptor (GR) transcriptional functions. Using a comprehensive set of techniques, we observed a reduction in stress-induced behaviors, highlighting the crucial role of shorter isoform of FRMD5. Further investigations revealed that shorter isoform of FRMD5 forms condensates in nuclear and interacts with a set of proteins involved in GR protein complex and transcriptional cofactors. Besides, FRMD5 accelerates the nuclear translocation and modulates transcriptional functions of GR. Systemic administration of peptides that disrupt FRMD5 functions led to transcriptional functions changes of GR and decreased stress-induced behaviors in mice. Overall, this study uncovers an intrinsic mechanism modulating transcriptional functions of GR via specific FRMD5 isoform, identifying a potential intervention target for PTSD.

overall_design: RNA-seq profiling of L929 cell lines with truncated FRMD5 (Isoform 2) after 4 hours DEX treated, and primary neurons of mice with interfering peptide after DEX treated.

first: Qi-Fan

last: Yang

first: Ting

last: Wu

first: Yun

last: Wang


title: Pancreatic Cancer Datasets

first: Lorenzo

last: Tomassoni

address: 1130 St Nicholas Avenue


title: CTCF-mediated cis-regulatory chromatin insulation enforces a  central B-cell tolerance checkpoint [RNA-seq]

first: Yehudit

last: Bergman

address: Ein Karem

summary:
The generation of a diverse and self-tolerant B cell repertoire is essential for adaptive immunity and is achieved through V(D)J recombination. In mice, Igκ is the dominant light chain, whereas Igλ rearrangement typically occurs in response to nonproductive or autoreactive Igκ recombination, a process termed receptor editing.
Recombination at the RS element deletes the Igκ constant exon, silencing the locus and enabling Igλ expression. However, the epigenetic regulatory framework that orchestrates and governs receptor editing remains poorly defined. Here, we identify a CTCF-binding insulator element (CBE) within the 3′ Igκ super-enhancer (3′-SEκ) that regulates receptor editing and directs the κ-to-λ switch required for Igλ⁺ B-cell development. Mechanistically, loss of this CBE activates an insulated enhancer within the 3′-SEκ, causing aberrant Vκ rearrangements and altered chromatin interactions through disrupted loop extrusion dynamics. Notably, loss of this CBE in mice leads to increased autoantibody production by ten weeks of age, demonstrating that CBE-mediated chromatin architecture shapes B cell fate by constraining autoreactive potential. Collectively, our findings define a novel CTCF-dependent cis- regulatory insulation checkpoint that connects chromatin loop extrusion to antigen- driven receptor editing, thereby enforcing B-cell tolerance.

overall_design: Genome-wide RNA profiling was performed on ex vivo small pre-B cells isolated from 6–12-week-old C57BL/6 WT and ΔrsCBE mice.

first: David

last: Gomes

first: Eliraz

last: Gitelman


title: Integration protocol of microglia and cerebral organoid

city: Boston

first: Yukyeong

last: Lee

state: MA

address: 185 Cambridge Street

institute: MGH

summary: sample analysis by an integrated protocol to induce microglia and neurons

first: Rakesh

last: Karmacharya


title: mRNA transcriptome data of granulosa cells cultured either with BSA (bovine serum albumin) or NEFA (non estrified fatty acid) mix

city: Dummerstorf

first: Vijay simha

last: Baddela

address: Wilhelm-Stahl-Allee 2

department: Reproductive biology

country: Germany

institute: Leibniz institute for farm animal biology

summary: Postpartum negative energy balance could contribute to the reduced fertility rates in females, partly due to elevated levels of non-esterified fatty acids (NEFA) in blood and in ovarian follicular fluid. NEFA has been shown to influence ovarian cell metabolism and function, yet the underlying molecular mechanisms remain incompletely understood. During the present investigation, transcriptome differences induced by a mixture of oleate (200 µM), palmitate (100 µM) and stearate (100 µM) were were  analyzed to understand the specific effects of NEFA in ovarian follicular granulosa cells.

overall_design: RNA from BSA (4n) and NEFA (4n) treated GC were processed for microarray analysis

first: Vijay


title: Metabolic control of innate immune activation in TET2-mutant clonal hematopoiesis [ChIP-seq]

first: Peter

last: Kim

address: 360 Longwood Ave, LC8202

department: Medical Oncology

institute: Dana Farber Cancer Institute

summary: TET2 mutations drive inflammation in clonal hematopoiesis of indeterminate potential (CHIP), but the mechanism by which TET2 inactivation leads to inflammation is not understood. Here, we used in vivo genome-wide genetic perturbations in primary wildtype (WT) or Tet2 knockout (KO) Cas9+ hematopoietic stem-progenitor cells (HSPCs) in a model of zymosan peritonitis to elucidate the basis of Tet2 KO inflammation. We found that Tet2 restrains O-linked N-acetylglucosamine (O-GlcNAc) glycosyltransferase (Ogt), a Tet2 binding partner and metabolic sensor that integrates nutrient availability. Tet2 loss disrupts this Tet2-Ogt interaction, and dysregulated Ogt facilitates widespread H3K4 trimethylation including lipid-related gene loci and inflammatory lipid droplet formation. We identified ATP citrate lyase (Acly) as a critical node for lipid accumulation, inflammation, and myeloid expansion in Tet2 deficiency. In summary, we reveal that Tet2 serves as a negative regulator of the nutrient sensor Ogt, and that Tet2 inactivation leads to aberrant lipid droplet formation and inflammation.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Chipseq profiling from various sources including bone marrow derived macrophages and monocytic human U937 cell line with or without TET2 KO, exposed to various conditions.

first: Christopher

last: Hergott

first: Aidan

last: Miller

first: Amy

last: Deik

first: Meaghan

last: Boileau

first: Kevin

last: Bullock

first: Kerry

last: Pierce

first: Angeline

last: Choy

first: Wesley

last: Shin

first: Marie

last: McConkey

first: Justin

last: Loke

first: Birgitta

last: Ryback

first: Michael

last: Trinh

first: Hong

last: Yue

first: Hojong

last: Yoon

first: Paul

last: Park

first: Shourya

last: Burman

first: Matthew

last: Vander Heiden

first: Eric

last: Fisher

first: Scott

last: Armstrong

first: Clary

last: Clish

first: Benjamin

last: Ebert


title: Engineered miR-122 inhibitors enhance endothelial mitochondrial efficiency and improve vascular function

city: Iowa City

first: Ajit

last: Vikram

state: IA

address: 285 Newton Road

department: Internal Medicine

institute: University of Iowa

summary: MicroRNA-122-5p (miR-122) is primarily expressed by the liver and is increasingly released into the bloodstream during obesity. It impacts the function of non-liver tissues, such as vascular endothelial cells, and increases the risk of diabetic vasculopathy. Using a novel gamma-peptide-nucleic acid-based miR-122 inhibitor (γP-122-I), we show that miR-122 regulates blood glucose level and endothelial function in high-fat diet-fed mice. Targeting γP-122-I to endothelial cells retains its ability to improve vascular function but reduces metabolic benefits compared to the non-targeted version. Our results show that the endothelial cells take up miR-122 through a neuropilin-1-dependent mechanism. The aortic transcriptomics analysis implicates the role of miR-122 in mitochondrial function. The aortas of high-fat diet-fed mice receiving an inhibitor of miR-122 were more efficient in oxygen consumption despite a decline in the expression of mitochondrial electron transport chain complexes, especially complex I. Our in vitro data show that miR-122 overexpression decreases the abundance of mitochondria with high membrane potential and spare respiratory capacity under hyperglycemic conditions. These findings support the therapeutic potential of the γPNA-based miR-122 inhibition in diabetic vasculopathy.

overall_design: 8-week-old mice (male) were fed a high-fat diet (HFD) for eight weeks. Two weeks after dietary intervention, these mice were intraperitoneally injected with either γPSC, γP-122-I, e-γP-SC, or e-γP-122-I (0.25 μmol.kg-1) for six weeks. The aorta was carefully isolated, cleaned, and RNA was isolated.

first: Ravinder-Reddy

last: Gaddam


title: Inhibition of sphingosine metabolism suppresses ferroptosis and rejuvenates aged hematopoietic stem cells to promote cognition and longevity [Low input RNA-seq]

city: Guangzhou

first: Meng

last: Zhao

address: Zhongshan 2nd Road

institute: Sun Yat-sen University

summary: Deletion of sphingosine kinase 2 (Sphk2) suppresses ferroptosis and restores functional capacities in old HSCs

overall_design:
Low input RNA-seq on HSCs
ABC294640 is the specific SphK2 inhibitor used in this study.

first: Changzheng

last: Li


title: Cyclophilin A Stabilizes the Capsid Protein p72 to Facilitate African Swine Fever Virus Replication

city: Harbin

first: Liqin

last: Yang

address: haping road 678

department: Harbin Veterinary Research Institute

institute: Chinese Academy of Agricultural Sciences

summary: African swine fever virus (ASFV) is a large DNA virus that poses a major threat to the global swine industry. Its virion is encapsulated by an icosahedral capsid predominantly composed of the structural protein p72, which constitutes approximately one-third of the total virion mass. Despite its abundance, the mechanisms regulating p72 stability remain poorly understood. Here, we demonstrate that host-mediated stabilization of p72 is essential for efficient ASFV replication. Mass spectrometry of p72 co-precipitates identified host cyclophilin A (CypA, also known as PPIA) as a key binding partner of p72. CypA interacts with p72 both in vitro and in vivo, specifically engaging exposed regions of p72 via its hydrophobic cavity. CypA interaction stabilizes p72 by reducing K63-linked ubiquitination and preventing proteasomal degradation, whereas cyclic CypA inhibitors destabilize p72 by disrupting this interaction and promoting its ubiquitination. Importantly, genetic or pharmacological inhibition of CypA markedly impairs ASFV replication. Mechanistically, CypA inhibition disrupts viral factory formation and virion assembly by decreasing p72 protein accumulation without affecting its transcription. Together, our findings uncover a previously unrecognized host-dependent mechanism regulating capsid protein stability and highlight host CypA as a promising target for antiviral strategies against ASFV.

overall_design: RNA-seq profiling of porcine alveolar macrophages to compare transcriptomic profiles under ASFV infection and NIM811 treatment conditions, including four experimental groups: (1) uninfected with DMSO control, (2) uninfected with NIM811, (3) ASFV-infected with DMSO, and (4) ASFV-infected with NIM811.

first: Hui

last: Kong

first: Xiangpeng

last: Sheng

first: Dongming


title: Inhibition of sphingosine metabolism suppresses ferroptosis and rejuvenates aged hematopoietic stem cells to promote cognition and longevity [scRNA-seq]

overall_design: scRNA seq on ckit positive cell


title: A Polyamine-mediated Posttranslational Modification Required for Macrophage Tissue Residency [Ribo-seq]

first: Immunometabolism

last: Department

address: 1650 Orleans Street

summary: Tissue-resident macrophages (RTMs) form during embryogenesis, self-renew locally, and regulate tissue homeostasis by clearing dead cells and debris. During tissue damage, however, bone marrow-derived monocytes enter tissue sites and differentiate into RTM, repairing the tissue and replenishing macrophages in the niche. Universal cell-intrinsic mechanisms that control the monocyte to RTM transition remain elusive. We investigated mice with myeloid cell deletions in deoxyhypusine synthase (DHPS), an enzyme that mediates the polyamine-dependent hypusine modification of the translation factor eIF5A, and found that DHPS is required for cell adhesion and signaling programs critical for RTMs. Without DHPS expression, immature tissue macrophages form, but RTM and associated homeostatic functions are lost. Thus, the polyamine-hypusine pathway is a global, tissue-agnostic program driving the differentiation trajectory of monocyte-derived macrophages into RTM.

overall_design: Freshly isolated cells from total peritoneal lavage from 3 WT or 3 DHPS KO mice were used for ribosomal analysis.

first: Erika

last: Pearce

first: David

last: Sanin

first: Gustavo

last: Carrizo


title: Dynamics of phage-host interactions in Bacteroides fragilis resolved by single-cell transcriptomics

city: Seattle

first: Dmitry

last: Sutormin

state: WA

address: 401 Terry Ave N

institute: Institute for Systems Biology

summary: The interactions between lytic phages and their hosts are typically studied in bulk culture, which obscures cell-cell differences in infection susceptibility or expression of protective factors. Here, we use bacterial single-cell RNA sequencing to profile the transcriptomes of ~50,000 cells from cultures of a human pathobiont, Bacteroides fragilis, infected with a lytic bacteriophage. From a single sampling, we quantified the asynchronous progression of phage infection in individual bacterial cells and reconstructed the infection timeline, characterizing both host and phage transcriptomic changes as infection unfolded. Further, we discovered phenotypic subpopulations of bacteria that remained uninfected. Each cell's vulnerability to phage infection was influenced by expression of multiple genetic loci, most prominently phase-variable capsular polysaccharide (CPS) biosynthesis pathways and an operon predicted to encode fimbrial genes. These findings uncovered genome-wide phase variation and stochasticity that enable bacterial survival and re-growth without acquiring additional mutations. Overall, we establish bacterial single-cell RNA sequencing as a powerful platform for investigating the dynamics of host-phage interactions and revealing the roles of phase variation and stochasticity in bacterial defenses.

overall_design: Exponentialy growing Bacteroides fragilis NCTC 9343 culture was a) cultivated in chopped meat carbohydrate (CMC) media; b) infected with Bf12P1 phage in CMC media; c) cultivated in CMC media supplemented with 100µM bile acid DCA; d) infected with Bf12P1 phage in CMC media supplemented with 100µM bile acid DCA. Cells were fixed in 4% formaldehyde and processed via microSPLiT protocol.

first: Anika

last: Gupta

first: Norma

last: Morella

first: Anna

last: Kuchina


title: Identification of recurrent tumor-specific markers in patient-matched ovarian cancer samples

city: Yeonsu-gu, Incheon

first: Johyeon

last: Nam

address: 265, Harmony-ro

department: Division of Life Sciences

country: South Korea

institute: Incheon National University

summary: In this study, we performed a transcriptomic comparative analysis on formalin-fixed paraffin-embedded (FFPE) specimens, collecting both primary ovarian cancer tissue and recurrent ovarian cancer tissue in a series from the same patients in real-world settings. Our aim was to identify clinically significant Differentially Expressed Genes (DEGs) associated with the recurrence of HGSOC .

overall_design: We performed RNA-seq on matched primary and recurrent ovarian cancer tissues from 24 patients. This study aims to identify the genetic marker of ovarian cancer recurrence.

first: Min-A

last: Kim

first: Mi-Ryung

last: Han

first: Jaehoon


title: Inhibition of sphingosine metabolism suppresses ferroptosis and rejuvenates aged hematopoietic stem cells to promote cognition and longevity  [CUT&TAG]

first: Meng

overall_design: CUT&Tag seq on HSCs

first: Changzheng


title: Sequencing DNA methylation and hydroxymethylation at co-occurring chromatin features

city: Cambridge

first: Somdutta

last: Dhir

address: Robinson Way

department: CRUK-CI

institute: University of Cambridge

summary: Epigenetic modifications govern chromatin dynamics and cell state. However, current methods cannot simultaneously resolve the presence of multiple DNA modifications at co-occurring chromatin-associated features. It is thus not clear how these features are physically coupled and how their combinations regulate genome function. To address this key question, we report 6-base-CUT&Tag, a method for simultaneous 6-base sequencing at target chromatin features. Using 6-base-CUT&Tag to profile 5mC and 5hmC at co-occurring histone modifications in mouse embryonic stem cells, we identify histone mark-specific signatures of methylation and hydroxymethylation to improve discrimination of different functional enhancer states.

overall_design: NGS sequencing DNA libraries prepared from genomic DNA from mouse ESC cells. Cells were probed using the CUT&Tag methodology using different primary antibodies against a panel of histone marks and 6-base-seq DNA libraries were prepared from the extracted gDNA (6-base-CUT&Tag). 2-3 biological replicates are included per experiment, originated from independent E14Tg2a mESC cultures. Libraries prepared from total tagmented genomic DNA from E14 cells are also included as whole-genome (WG) controls.

first: Rafael

middle: D

last: C Tavares

first: Xuan

last: He

first: Jack

last: Monahan

first: Minna

last: Taipale

first: Paula

last: Golder

first: Aldo

last: Ciau-Uitz

first: Walraj

last: Gosal

last: Tannahill

first: Shankar

last: Balasubramanian


title: A Polyamine-mediated Posttranslational Modification Required for Macrophage Tissue Residency [RNA-Seq]

city: Baltimore

first: Immunometabolism

last: Department

institute: Johns Hopkins University

overall_design: Macrophages from several tissues were obtained from 4 separate WT or DHPS KO mice via FACS.

first: Erika

last: Pearce

first: David

last: Sanin

first: Gustavo

last: Carrizo


title: CTCF-mediated cis-regulatory chromatin insulation enforces a central B-cell tolerance checkpoint [ATAC-seq]

city: Jerusalem

first: Yehudit

last: Bergman

department: Department of Developmental Biology and Cancer Research

institute: The Hebrew University Of Jerusalem

overall_design: ATAC-seq was performed on ex vivo small pre-B cells isolated from 6–12-week-old C57BL/6 WT, ΔrsCBE, and ΔrpiaCBE mice.

last: Gomes

first: Eliraz

last: Gitelman


title: Inhibition of sphingosine metabolism suppresses ferroptosis and rejuvenates aged hematopoietic stem cells to promote cognition and longevity [RNA-seq]

first: Meng

last: Zhao

overall_design: Low input RNA-seq on HSCs

last: Li


title: Cellular Quiescence Displays a Transcriptional Inflammatory Phenotype Similar to the One in Cellular Senescence [scRNA-seq]

city: Belmont

first: Azucena

last: Rocha

address: 42 Alma Avenue

department: MCB

institute: Brown University

summary: Cellular senescence, traditionally viewed as the irreversible arrest of proliferating cells in response to stress and DNA damage, is a key mechanism underlying cellular aging. Several studies have identified a process called ”quiescence-origin senescence,” where quiescent cells bypass proliferation and transition directly into senescence. In this context, we introduce the concept of the transcriptional Quiescence-Associated Secretory Phenotype (tQASP), a putative inflammatory secretory profile that arises in human quiescent cells and may serve as a precursor to the full senescence-associated secretory phenotype (SASP). We report that quiescent cells express tQASP genes in a cell-type-, induction-dependent manner and that this response is influenced by environmental oxygen levels. tQASP expression is also increased in tissue affected by Idiopathic Pulmonary Fibrosis (IPF) and shows a positive correlation with myofibroblast activation. Together, these data support the concept of a continuum between quiescence and senescence and suggest that the tQASP may be an early marker of cellular aging, with significant implications for tissue homeostasis, aging, cancer, and disease progression.

overall_design: single cell RNA Sequencing of quiescent human Lung fibroblasts (LF1) induced by a combination of contact inhibition and serum withdrawal

first: Radha

last: Khalekar

first: John

last: Sedivy

first: Nicola

last: Neretti


title: Profiling the aging transcriptome of the gut wall (longitudinal muscle and myenteric plexus) in mice

first: Loyal

last: Goff

state: MD

address: 733 N Broadway

summary: These data represent the changing transcriptal profiles of the longitudinal muscle and myenteric plexus (LMMP) of the murine ileum with age. Tissues from 1-month, 6-month, and 17-month old mice were characterized.

overall_design: Mice of three ages (1-month, 6-month, and 17-month) and both sexes (n=3 per unique condition; n=2 for 17mo F) were anesthetized with isoflurane and sacrificed by cervical dislocation. The ileum of the small intestine was isolated and flushed followed by peeling of the longitudinal muscle containing myenteric plexus (LMMP) from the underlying tissue. RNA-seq of this tissue was performed to identify factors that define the juvenile or aging myenteric plexus and its local environment.

first: Jared

last: Slosberg

first: Subhash

last: Kulkarni