title: Feeding with a ketonic diet confers complete protection against malaria

city: Shenyang

first: Yiwei

last: Zhang

address: 120 Dongling Road

institute: Shenyang Agricultural University

summary: Environmental factors restrict malarial parasite development, but the influence of host metabolic variations on the infectivity of blood-stage parasite is not fully understood. Here, we show that the administration of a ketogenic diet (KD) to mice conferred complete protection against Plasmodim berghei infection. The β-hydroxybutyrate (βOHB), a predominant ketone body, was elevated in P. berghei infected mice on a KD, and  inhibited the proliferation of both P. berghei and P. falciparum. A KD and βOHB induced metabolic reprogramming in the parasites resulting in reduced cellular nicotinamide adenine dinucleotide (NAD), which led to the downregulation of genes controlling parasite development, erythrocyte invasion and pathogenicity. This study reveal that A KD-derived βOHB can ultimately cause the developmental arrest of Plasmodium parasites and benefits host resistance to malaria.

overall_design: The underlying mechanism of ketone bodies-mediated parasite inhibition on P. falciparum was explored via single-cell RNA sequencing (scRNA-seq) analysis. A single-cell transcriptome atlas of P. falciparum 3D7 before and after βOHB treatment was first constructed


title: Immunoglobulin A controls intestinal virus colonization to preserve immune homeostasis.

status: Public on Mar 24 2025

city: Chicago

first: Joseph

middle: Joy

last: Sifakis

state: IL

address: 900 East 57th Street

department: Chemistry

country: USA

institute: The University of Chicago

type: Expression profiling by high throughput sequencing

summary: Immunoglobulin A (IgA) is the predominant immunoglobulin isotype in mammals, primarily secreted at type I mucosal surfaces. Despite its abundance, the precise role of secretory IgA in the intestinal lumen, where it coats a diverse array of commensal microbiota, has remained elusive. Our study reveals that germinal center IgA responses are essential for preventing chronic colonization of the gut by specific viruses. In absence of IgA, chronic viral colonization triggers an antigen-driven expansion of CD8αβ+ intraepithelial lymphocytes (IELs). Although, these IELs are unable to clear the virus, they contribute to maintaining homeostasis by regulating its load and type-I interferon responses. Consequently, IgA deficiency increases susceptibility to colitis in genetically susceptible host or following chemical induction, but only in presence of viral pathobionts requiring IgA for their clearance. These findings underscore the potential vulnerability of IgA deficient individuals to immunopathology when exposed to selective viral pathobionts.

overall_design: scRNA-seq profiling of jejunal intraepithelial CD8αβ+ lymphocytes from three Ighasec+/- and Ighasec-/- SPF mice.

first: Wioletta

last: Lisicka

first: Zachary

middle: M

last: Earley

middle: J

first: Bana

last: Jabri

first: Samantha

last: Riesenfeld


title: Spatial single cell RNA profiles from medulloblastoma Group 3/4 tumors with oncogene aberrations

status: Public on Mar 25 2025

city: Heidelberg

first: Konstantin

last: Okonechnikov

state: Germany

address: Im Neuenheimer Feld 280

department: Pediatric Neurooncology

country: Germany

institute: German Cancer Research Center

summary: Medulloblastomas Group 3/4 are cerebellar tumors  with strong inter- and intratumoral heterogeneity.  In our research project we focused on specific Group 3/4 tumor cases with somatic aberrations MYC,MYCN and PRDM6 using various single cell  techniques.

overall_design: Cohort of MB G34 tumors was processed with Resolve Bioscience protocol in order to obtain spatial single cell profiles


title: Epigenetic and transcriptomic profiling of mouse neural stem cells

city: Ikoma

first: Taito

last: Matsuda

country: Japan

institute: Nara Institute of Science and Technology

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Neural stem cells (NSCs) in the hippocampus of mammalian brains decline rapidly and persistently with age, eventually leading to impairment of hippocampal memory function in later life. However, the mutual relationship between epigenetic remodeling and transcriptional regulation that initially compromises hippocampal NSC activity during the early stage of chronological aging has not been well elucidated. We here performed single-cell RNA sequencing (scRNA-seq) and single-cell assay for transposase-accessible chromatin sequencing (scATAC-seq) for NSCs and newly generated neurons in a chronological manner. Integrated analysis using these datasets revealed that the chromatin profile of hippocampal NSCs and their progeny undergoes continuous alterations from neonatal to mature adult stages, accompanied by considerable changes in transcriptional regulation. We further show that a decrease in the expression of Setd8, encoding the only known enzyme that catalyzes histone H4 monomethylation at lysine 20 (H4K20me1), underlies age-related changes in the characteristics of mouse NSCs in the hippocampus: the downregulation of Setd8 expression elicits age-related changes in gene expression and epigenetic regulation, and impairs NSC activity, leading to functional deficiency in hippocampal memory. Thus, our study identifies a mechanism for an unexpectedly early-onset decline of NSC activity and hippocampal neurogenesis that precedes aging.

overall_design:
We used 50,000 cultured NSCs to construct a library for each sample. Samples collected from cultured mouse NSCs in Setd8 knock down after selection with puromycin were subjected to RNA-seq, ChIP-seq (H4K20me1) and ATAC-seq.

For scRNA-seq and scATAC-seq, the cells were sorted on a FACS Aria II (BD Biosciences), with 7-AAD (559925, BD Pharmingen) added to exclude nonviable cells from the analysis. Cells dissociated from the DG of WT mice were used to draw gates for EGFP+ and EGFP- cell populations.
The EGFP+ cells were sorted into N2 medium and centrifuged at 200 x g at 4C for 10 min. The supernatant was carefully removed, leaving 30 uL to resuspend the cell pellet. One microliter of cells was mixed with 10uL Trypan Blue to determine cell viability.
For scRNA-seq and scATAC-seq analyses, 700-1200 cells/uL were used.

first: Shuzo

last: Matsubara


title: The impact of Coptis chinensis Franch root rot on leaves

city: wuhan

first: yu

middle: kai

last: kaidi

address: shizishangjie

country: China

institute: Huazhong Agricultural University

summary: Root rot disease poses a devastating threat to Coptis chinensis Franch, a medicinal plant prized for its bioactive alkaloids. To dissect its defense mechanisms, we conducted integrated transcriptomic and metabolomic analyses on resistant (R), early-stage infected (S-ES), and late-stage infected (S-LS) plants. Our findings reveal a disease severity-dependent escalation in flavonoid metabolism. Key metabolites, such as kaempferol and quercetin derivatives, were significantly increased compared to R, paralleled by progressive upregulation of biosynthetic genes (PAL, CHS, CHI, FLS). Strikingly, salicylic acid (SA)-associated metabolites and pathway genes (NPR1, NPR3/NPR4) showed no differential expression across groups, contrasting with typical SA-mediated defenses in other species. This study uncovers flavonoid biosynthesis as the primary defense strategy in C. chinensis during root rot progression, while SA signaling may not be the main defense mechanism. These results provide actionable targets for enhancing disease resistance in medicinal plants through metabolic engineering

first: kaidi

last: Yu

first: yuying

last: Yang


title: Immunoglobulin A controls intestinal virus colonization to preserve immune homeostasis [ZE309]

overall_design: bulk RNA-seq profiling of the intestinal tissue from duodenum, jejunum, ileum and colon of GF or SPF Ighasec+/- and Ighasec-/- mice


title: Angiotensin Converting Enzyme (ACE) expression in microglia reduces amyloid β deposition and neurodegeneration by increasing SYK signaling and endolysosomal trafficking

city: Los Angeles

first: Warren

last: Tourtellotte

state: California

address: 8700 Beverly Blvd

department: Pathology and Laboratory Medicine

institute: Cedars-Sinai

summary: Angiotensin Converting Enzyme (ACE) has been studied for its role in the regulation of myeloid cell bactericidal and antitumor activity. Here, we generated a mouse model with Tamoxifen-dependent ACE expression and Cx3CR1-Cre control. These mice were crossed to 5xFAD and 9 month forebrains were harvested and FFPE processed to assess the ACE-dependent changes in amyloid-reactive microglia.

overall_design: Paraffin tissue blocks were faced until matching anatomical regions were visible across all blocks and 3 hippocampal donor cores (1mm diameter) were extracted from each block and placed into a recipient tissue microarray block using a GrandMaster TMA (3D Histotechnology, Hungary) instrument

first: Andrew

last: Gomez

first: Hyae Ran

last: Byun

first: Shaogen

last: Wu

first: AKM Ghulam

last: Mhuammad

first: Jasmine

last: Ikbariyeh

first: Jaelin

last: Chen

first: Alek

last: Muro

first: Lin

last: Li

first: Kenneth E

last: Bernstein

first: Richard

last: Ainsworth


title: Ground-state pluripotent stem cells are characterized by Rac1-dependent cadherin-enriched F-actin complexes

city: Singapore

first: Shiying

last: LIU

state: Singapore

address: 61 Biopolis Dr, Proteos, Singapore 138673

department: Institute of Molecular and Cell Biology (IMCB)

country: Singapore

institute: Agency for Science, Technology and Research (A*STAR)

summary: Pluripotent stem cells (PSCs) exhibit extraordinary differentiation potential and are thus highly valuable cellular model systems. However, although different PSC types corresponding to distinct stages of embryogenesis have been in common use, aspects of their cellular architecture and mechanobiology remain insufficiently understood. Here, we investigated how the actin cytoskeleton is regulated in different pluripotency states. We observed a drastic reorganization during the transition from ground-state naïve mouse embryonic stem cells (mESCs) into converted prime epiblast stem cells (EpiSCs). mESCs are characterized by prominent actinenriched cortical structures that contain cadherin-based cell–cell junctional components, despite not locating at cell–cell junctions. We term these structures ‘non-junctional cadherin complexes’ (NJCCs) and show that they are under low mechanical tension, depend on the ectodomain but not the cytoplasmic domain of E-cadherin, and exhibit minimal Ca2+ dependence. Active Rac1 was identified as a negative regulator that promotes β-catenin dissociation and NJCC fragmentation. Our data suggests that NJCCs might arise from the cis-association of E-cadherin ectodomain, with potential roles in ground-state pluripotency, and could serve as structural markers to distinguish heterogeneous population of pluripotent stem cells.

overall_design: RNA-SEQ profiling of ground-state naïve mouse embryonic stem cells (mESCs) in feeder-free 2iLIF cultures and converted prime epiblast stem cells (EpiSCs) in vitro following the protocal indicated in the manuscript.

last: Liu

first: Pakorn

last: Kanchanawong


title: A multimorphic variant in ThPOK causes a novel human disease with T cell abnormalities, immune dysregulation, and severe fibrosis [scRNA-Seq]

status: Public on Mar 23 2025

city: Vancouver

first: Stuart

last: Turvey

state: BC

address: 938 W 28th Ave

country: Canada

institute: BCCHRI

summary: ThPOK is a transcription factor which acts as a master regulator of CD4+ T cell lineage commitment. We report the first human disease caused by genetic alteration in ThPOK, specifically, a damaging heterozygous de novo variant in ThPOK (NM_001256455.2: c.1080A>C, p.K360N). This patient exhibited the unusual constellation of persistent CD4+ T cell deficiency, allergy, interstitial lung disease, corneal vascularization and scarring, developmental delay, and growth failure. The ThPOKK360N variant displayed abnormal multimorphic activity, interfering with ThPOKWT (antimorph), failing to bind wild-type ThPOK consensus sequences (amorph), and showing novel DNA-binding specificity (neomorph). Single-cell RNA sequencing revealed defects in CD4+ and CD8+ T cell maturation and activation (hypomorph). Recapitulated in lentivirally transduced healthy control T cells and fibroblasts, transcriptomic analysis showed ThPOKK360N-transduced T cells had impaired TCR activation, and ThPOKK360N-transduced fibroblasts with increased pro-fibrotic gene expression. This novel human disease confirms ThPOK's role in CD4+ T cell development but also uncovers novel roles in TCR activation and regulation of fibrotic pathways in fibroblasts.

overall_design: Sinlge cell RNA sequencing on patient sample alongside three healthy controls with or without stimulation with CD3/CD28 T cell activator.

first: Maryam

last: Vaseghi-Shanjani

first: Mehul

last: Sharma

first: Simran

last: Samra

first: Pariya

last: Yousefi


title: iPSC-derived NK cells engineered with CD226 effectively control acute myeloid leukemia

city: Beijing

first: Cai

last: Runze

address: Xueyuan Rd.,Haidian District

institute: Peking University Health Science Center

summary: CD226 plays a vital role in NK cell cytotoxicity, interacting with its ligands on tumor targets. Acute myeloid leukemia (AML) cells have developed mechanisms to escape NK cell cytotoxicity, including inducing downregulation of CD226 on NK cells. Induced pluripotent stem cell -derived NK (iPSC-NK) cells offer an important source of standardized off-the-shelf NK cell therapy to treat AML patients. In this study, we engineered iPSC-NK cells with CD226 to assess the ability of killing AML cells. iPSC-NK cells engineered with CD226 have a typical NK cell phenotype and demonstrate improved anti-AML activity and multiple cytokines releasing at low effector-to-target ratios. Transcriptomic analysis revealed upregulation of immune effector function pathways associated with cytotoxicity and immune activation in CD226-overexpression iPSC-NK cells. In an AML xenograft model, mice treated with CD226 overexpression iPSC-NK cells exhibited significantly reduced leukemia burden, prolonged survival, decreased systemic inflammation compared to those treated with Control iPSC-NK cells. Overall, our study provided evidence that iPSC derived-NK cells engineered with CD226 represent a promising candidate for off-the-shelf immunotherapy, particularly in AML and other CD226 ligand-expressing malignancies.


title: TNF signature in advanced melanoma patients treated with immune checkpoint inhibitors: results from the MELANFα clinical study

city: Toulouse

first: Anne

last: Montfort

address: 2 avenue Hubert Curien

country: France

institute: INSERM

summary: Resistance to immune checkpoint inhibitors (ICI) in cancer patients is not fully understood and predictive biomarkers are lacking. MELANFα (NCT03348891) is an open-label, prospective, multicenter cohort of 60 patients with advanced melanoma receiving ICI (bitherapy: ipilimumab + nivolumab; monotherapy: pembrolizumab or nivolumab). In this study we collected blood from patients taken at baseline (week0) and 6 weeks after treatment initiation (week6), isolated peripheral blood monuclear cells and assessed the impact of treatment systemic immune responses to identify potential markers of response/resistance.

overall_design: CITE-seq PBMC from 8 advanced melanoma patients taken at baseline (Week 0, W0) and 6 weeks after the initiation of treatment (Week 6, W6) with nivolumab (1mg/kg) + ipilimumab (3mg/kg). Each sample was stained with 11 CITE-seq antibodies (ADT) before single cell sequencing

first: Mathieu

last: Virazels

first: Amélie

last: Lusque

first: Stéphanie

last: Brayer

first: Matthieu

last: Genais

first: Carine

last: Dufau

first: Jean

last: Milhès

first: Thomas

last: Filleron

first: Cécile

last: Pagès

first: Vincent

last: Sibaud

first: Laurent

last: Mortier

first: Olivier

last: Dereure

first: Maha

last: Ayyoub

first: Amandine

last: Fabre

first: Nathalie

last: Andrieu-Abadie

first: Vera

last: Pancaldi

first: Céline

last: Colacios

first: Nicolas

last: Meyer

first: Bruno

last: Ségui


title: A multimorphic variant in ThPOK causes an inborn error of immunity with T cell defects and fibrosis [fibroblasts]

first: Stuart

last: Turvey

overall_design: Fibroblasts were grown from patient and healthy control lung tissue biopsy before lenti virus transduction and RNA was studied via RNA seq.

first: Maryam

last: Vaseghi-Shanjani

first: Mehul

last: Sharma

first: Simran

last: Samra

first: Pariya

last: Yousefi


title: Single cell resolution analysis of multi-tissue derived human iNKT cells reveals novel transcriptional paradigms

status: Public on Mar 22 2025

city: St. Louis

first: Melissa

last: Mavers

state: MO

address: 660 Euclid Ave

department: Pediatrics

institute: Washington University School of Medicine

summary: Invariant natural killer T (iNKT) cells are evolutionarily conserved innate lymphocytes important for host defense against pathogens. Further, they are increasingly recognized to play a role in tumor immune surveillance and in protection against graft versus host disease, and they are of particular importance as a universal donor for cellular therapies. Therefore, a thorough understanding of the biology of iNKT cells is critical. Murine studies have revealed the existence of transcriptionally and functionally distinct subsets, similar to T helper cell subsets. However, a comprehensive study of human iNKT cell heterogeneity is lacking. Herein, we define the transcriptomic heterogeneity of human iNKT cells derived from multiple immunologically relevant tissues, including peripheral blood, cord blood, bone marrow, and thymus, using single cell RNA-sequencing. We describe human iNKT cells with a naïve/precursor transcriptional pattern, a Th2-like signature, and Th1/17/NK-like gene expression. This combined Th1/17 pattern of gene expression differs from previously described murine iNKT subsets in which Th1- and Th17-like iNKT cells are distinct populations. We also describe transcription factors regulating human iNKT cells with distinct gene expression patterns not previously described in mice. Further, we demonstrate a novel T effector memory RA+ (TEMRA)-like pattern of expression in some human iNKT cells. Additionally, we provide an in-depth transcriptional analysis of human CD8+ iNKT cells, revealing cells with two distinct expression patterns—one consistent with naïve/precursor cells and one consistent with Th1/17/NK-like cells. Collectively, our data provide critical insights into the transcriptional heterogeneity of human iNKT cells, providing a platform to facilitate future functional studies and to inform the development of iNKT-based cellular therapies.

overall_design:
iNKT cells were enriched by magnetic beads and fluorescence activated cell sorted (FACS) using antibodies to the invariant T cell receptor. CD24+ stage 0 iNKT precursor cells were removed via FACS to ensure adequate numbers of differentiating and mature thymic iNKT cells to better facilitate comparison across tissues.

***************************************************************
raw data undergoing submission to relevant respository (dbgap/EGA)
***************************************************************

first: Reyka

last: Jayasinghe


title: Characterizing the Immune Infiltrates in Secondary Syphilis: Implications for Transmission and Pathology

status: Public on Mar 21 2025

city: Ann Arbor

first: Poulami

last: Dey

address: 1301 Catherine St

institute: Michigan Medicine

summary: Provisionally accepted

overall_design: biopsies from secondary syphilis

first: Johann

middle: E

last: Gudjonsson

first: Lam

middle: C

last: Tsoi


title: A multimorphic variant in ThPOK causes an inborn error of immunity with T cell defects and fibrosis [T cells]

overall_design: T cells were isolated from PBMCs of a healthy control and RNA seq was studied with or without transduction with ThPOK variant.


title: Transcriptional profiling of Mycobacterium tuberculosis exposed to antibiotics and drug combinations in broth culture.

city: Seattle

first: Ethan

last: Bustad

state: WA

address: 1900 Ninth Ave.

department: CGIDR

institute: Seattle Children's Research Institute

summary: We profiled  Mtb transcriptional response to a range of different antibiotic drug exposures in 7H9 broth culture by RNA-seq. Compound abbreviations: (EMB = ethambutol, BTZ = BTZ043, TYL = tyloxapol). We also hypothesized that an improved Mtb TRN could be inferred using a large, diverse compendium of RNA-seq data and therefore sought to supplement the existing repository of Mtb expression data available on SRA with additional conditions that could inform on Mtb's underlying regulatory programs. We went on to infer Mtb's TRN using a collection of inference methods in combination with this compendium, and employed it to build an interpretable machine learning model of Mtb fitness under stress.

overall_design: RNA-seq analysis was performed on samples taken from Mtb cultures subjected to various stresses.

first: José Manuel

last: Ezquerra-Aznárez

first: Jessica

last: Assadi

first: Robert

middle: D

last: Morrison

first: Shuyi

last: Ma

first: David

middle: R

last: Sherman

first: Santiago

last: Ramón-García


title: Effect of DDTC(I) on mRNA of SaOS2 cells in osteosarcoma

city: Taiyuan

first: Ruhao

last: Zhou

email: zrhi1998@163.com, zhouruhao@sxmu.edu.cn

address: 382 Wuyi Road,

institute: Shanxi medical university

summary: To investigate the mechanism of DDTC(I) on SaOS2 cells

overall_design: Comparative gene expression profiling of RNA-seq data for SaOS2 cells and their DDTC(I) treatments


title: MEG3 Upregulation in Myotonic Dystrophy and Its Correction by Antisense Conjugates Targeting Disease-Causing Mutations [miRNA-seq]

city: Oxford

last: Seoane Miraz

address: IMS-Tetsuya Nakamura Building, Old Road Campus, Roosevelt Drive, Headington

department: Paediatrics

country: United Kingdom

institute: University of Oxford

type: Non-coding RNA profiling by high throughput sequencing

summary: MEG3 upregulation has been confirmed to independently promote apoptosis in cancer, cardiovascular disease, and Alzheimer’s Disease. In this study, we unveiled that MEG3 is highly upregulated in Myotonic Dystrophy 1 (DM1) human muscle cells when compared with wild type cells. It is also increased in the muscle of DM1 mouse models, 15-fold in HSA-LR, and 6-fold in LC15. We also observed an increase of MEG3 in SMA mice (5-fold) indicating that MEG3 levels could play a role in the neuromuscular pathology leading to cell loss. Additionally in DM1, RNA sequencing revealed transcript retention of miRNAs and LncRNAs in the nucleus versus the cytoplasm, a phenomenon not observed with mRNAs. This nuclear retention could exacerbate pathological mechanisms involving non-coding RNA, such as MEG3-led apoptosis. In DM1 mice (HSA-LR), the correction of DM1 pathology with a peptide antisense oligonucleotide conjugate targeting the repeat expansion, a family of compounds currently being tested in a DM1 clinical trial, was able to also reduce the levels of MEG3. When direct interventions for the root cause of a disease are unavailable, the downregulation of MEG3 and its effect on apoptosis may serve as an alternative therapeutic approach, notably in conditions that arise from complex interactions of genetic and environmental factors.

overall_design: To detect misregulated miRNAs in sub-cellular compartments from healthy and DM1 affected human myotubes, we used PARIS kit to fraction human myotubes into nucleus and cytoplasm fractions. Then RNA-Seq was performed and expression levels of miRNAs were determined.

last: Seoane-Miraz

first: Miguel

middle: A

last: Varela

first: Matthew

last: Wood


title: Drosophila Hamlet mediates epithelial tissue assembly of the reproductive system

city: CANBERRA

first: Jiayu

last: Wen

state: ACT

address: 133 Garran Rd

department: Department of Genome Science

country: Australia

institute: John Curtin School of Medical Research, The Australian National University

summary: Epithelial tissue fusion requires coordinated molecular events at the ends of two epithelial structures. Regulatory mechanisms controlling these events remain largely elusive. In the Drosophila reproductive system (RS), this fusion unites the gonad and the genital disc-derived tissues, into a continuous tube. This study unveils the pivotal role of Hamlet (Ham), a Drosophila PR-domain containing transcription factor, in orchestrating epithelial tissue fusion in the RS. Loss of ham leads to sterility and disconnection between the testes (TE) and seminal vesicles (SV). Systematic analysis of Ham-downstream genes reveals cytoskeletal, metabolic regulators and signaling pathway components. Ham activates genes for epithelial differentiation and remodeling, while repressing genes required for tissue growth and patterning. Using multiplexed in situ hybridization, we demonstrate spatial-temporal gene expression dynamics in contacting epithelia. Key Ham downstream effectors include E-Cadherin (E-Cad), Toll (Tl) and Wnt2 signaling pathways, regulating tissue interaction and fusion. Our findings present a comprehensive gene network crucial for heterotypic epithelial tissue fusion. Mammalian Ham orthologs PRDM3 and PRDM16 are highly expressed in epithelial tissues, suggesting a conserved role across species.

overall_design: Using RNA-seq to identify differentially expressed genes between wt and ham mutant conditions. CUT&Tag was used to map Ham binding peaks. By integrating RNA-seq and CUT&Tag data, direct targets of Ham were identified.

first: Qi

last: Dai

first: Huazhen

last: Wang


title: Transcriptional response to rotenone exposure in C. elegans and the effects on progeny

status: Public on Mar 20 2025

city: Durham

first: Kinsey

last: Fisher

state: NC

address: 124 Science Drive

department: Biology

institute: Duke University

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

overall_design: Refer to individual Series


title: Sustaining Cyclin D1 Expression Renders Kidney Cancer Cells HIF2-Independent

city: Boston

first: Nitin

middle: H.

last: Shirole

state: MA

address: 450 Brookline Avenue, M448, Dana Farber Cancer Institute

institute: Dana Farber Cancer Institute

summary: Inactivation of theVHLtumor suppressor gene stabilizes HIF2a, which drives clear cell renal carcinoma (ccRCC). The HIF2ainhibitor belzutifan is approved for ccRCC treatment, butde novoand acquiredresistance are common. HIF2a, bound to ARNT, transcriptionally activates hundreds of genes. We performed genome-wide CRISPRa screens in HIF2a-dependent ccRCC lines treated with the belzutifan tool compound PT2399 to identify HIF2a-responsive genes that confer belzutifan resistance when not downregulated. Sustaining the expression of the HIF2atarget geneCCND1, encoding Cyclin D1, promoted HIF2a-independence/belzutifan resistance. This activity requires Cdk4/6 activation by Cyclin D1, but is not solely due to inhibitory phosphorylation of the canonical Cyclin D1 target, pRB. Indeed, ccRCC lines lacking all three pRB family members remained at least partially HIF2a-dependent. In this context. However, kinase-defective Cyclin D1 variants at least partially overrode belzutifan’s antiproliferative effects, suggesting that ccRCC promotion by Cyclin D1 requires the phosphorylation of pRB paralogs and one or more kinase-independent Cyclin D1 activities.

overall_design: To identify candidate enhancer RNAs and profile the immediate transcriptional effects of DMSO and PT2399 drug treatment at 6 hour timepoint, PRO-seq (Precision Run-On Sequencing) was performed to investigate single nucleotide resolution of nascent RNA 3' ends.

middle: H

first: Seth

last: Goldman

first: Karen

last: Adelman

first: William

middle: G

last: Kaelin


title: Real-time quantitative PCR analysis of EV-associated miRNAs from Human Treg cells

city: Napoli

first: Paola

last: De Candia

address: via Pansini 5

country: Italy

institute: UNIVERSITA' DEGLI STUDI DI NAPOLI FEDERICO II - Dipartimento di Medicina molecolare e Biotecnologie mediche

type:
Expression profiling by RT-PCR
Non-coding RNA profiling by array

summary: MicroRNAs (miRNAs) are small non-coding molecules targeting messenger RNAs and inhibiting protein translation. Regulated by dynamic micro-environmental cues, miRNAs in turn modulate key biological processes, including cell growth and development, energy utilization and homeostasis. In particular, miRNAs control the differentiation, survival and activation of both CD4+ T conventional (Tconv) and CD4+ T regulatory (Treg) cells, key players of the adaptive immunity; in particular, miRNA-mediated gene expression regulation contributes to the physiological response of those cells to infections and the pathological loss of immune homeostasis in autoimmunity. Upon T cell receptor (TCR) stimulation, Treg cells release a  fingerprint of miRNAs in association with extracellular vesicles (EVs), which is specific and significantly different from that released by Tconv cell counterpart (GSE183713). In this study, we have highlithed how Treg-cell derived EVs, and their miRNA content, are able to block the proliferation and activation of Tconv cells, demonstrating a strong tolerogenic function.

overall_design: Human Treg cell-derived EVs were isolated from conditioned media using size exclusion chromatography (Exo-spin™ columns, Cell Guidance) according to manufacturer’s protocol. Isolated EV eluate was total RNA extracted using RNAqueous-4PCR Kit (Ambion, U.S.A.) and a fixed volume of eluted RNA sample was used as input for reverse transcription reaction by miRCURY-LNA RT Kit according to manufacturer’s instruction (Qiagen, U.S.A.). EV-associated miRNAs (n = 752) were profiled by using the complete human miRCURY LNA miRNA panel I+II (V5, Qiagen, U.S.A.).

last: de Candia

first: Dario

last: Di Silvestre


title: Immunoglobulin A controls intestinal virus colonization to preserve immune homeostasis [ZE303]

city: Chicago

first: Joseph

last: Sifakis

address: 900 East 57th Street

institute: The University of Chicago

summary: Immunoglobulin A (IgA) is the predominant immunoglobulin isotype in mammals, primarily secreted at type I mucosal surfaces. Despite its abundance, the precise role of secretory IgA in the intestinal lumen, where it coats a diverse array of commensal microbiota, has remained elusive. Our study reveals that germinal center IgA responses are essential for preventing chronic colonization of the gut by specific viruses. In absence of IgA, chronic viral colonization triggers an antigen-driven expansion of CD8αβ+ intraepithelial lymphocytes (IELs). Although, these IELs are unable to clear the virus, they contribute to maintaining homeostasis by regulating its load and type-I interferon responses. Consequently, IgA deficiency increases susceptibility to colitis in genetically susceptible host or following chemical induction, but only in presence of viral pathobionts requiring IgA for their clearance. These findings underscore the potential vulnerability of IgA deficient individuals to immunopathology when exposed to selective viral pathobionts.

overall_design: RNA-seq profiling of the intestinal tissue (jejunum, ileum) or sorted epithelial cells (jejunum) from Ighasec+/- and Ighasec-/- mice treated with isotype or anty-CD8b depleting antibody for 3 weeks.

first: Wioletta

last: Lisicka

first: Zachary

last: Earley

first: Bana

last: Jabri

first: Samantha

last: Riesenfeld


overall_design: We performed RNA-Seq to identify HIF2a transcriptional targets after treating OSRC2 and TURH4TKB cells with HIF2a inhibitor PT2399 for 24hrs and 48 hrs.

first: Devishi

last: Kesar


title: Next Generation Sequencing unveils the impact of CD4+ regulatory T cell (Treg)-derived extracellular vesicles (EVs) on the transcriptome of CD4+ T cells.

summary:
Purpose: The goal of this study was the transcriptome high-throughput data analysis of CD4+ T cells isolated from peripheral blood of healthy subjects and T cell receptor (TCR)-stimulated in vitro in the presence of Treg cell-derived EVs, compared with Tconv cell-derived EVs, or with control (mock) EVs (particles isolated from unconditioned medium).
Methods:  total RNA profile of  human CD4+ T cells from healthy subjects. Libraries were prepared for sequencing on NextSeq 500 (llumina technology).
Results: Paired differential expression and pathway enrichment analysis showed that CD4+ T cells treated with Treg cell-derived EVs are characterized by lower expression level of a plethora of transcripts linked to protein catabolism and cell growth.
Conclusions: Several transcripts are differently regulated between CD4+ T cells treated with  Treg cell-derived EVs and the same cells treated with either Tconv cell-derived EVs or mock EV controls.

overall_design: CD4+ T cell mRNA profiles of 3 healthy subjects treated with Treg cell-derived EVs, or Tconv cell-derived EVs or controls were generated by deep sequencing, using Illumina platform.


title: Transcriptional profiling of Mycobacterium tuberculosis strains exposed to antibiotic treatment.

status: Public on Mar 19 2025

first: Ethan

last: Bustad

summary: We profiled the transcriptional response of different, phylogenetically distinct Mycobacterium tuberculosis (Mtb) strains to isoniazid (INH) and rifampicin (RIF) using RNA-seq. We also hypothesized that an improved Mtb TRN could be inferred using a large, diverse compendium of RNA-seq data and therefore sought to supplement the existing repository of Mtb expression data available on SRA with additional conditions that could inform on Mtb's underlying regulatory programs. We went on to infer Mtb's TRN using a collection of inference methods in combination with this compendium, and employed it to build an interpretable machine learning model of Mtb fitness under stress.

first: Tige

last: Rustad

first: Jessica

last: Assadi

first: Robert

last: Morrison

first: Shuyi

last: Ma

first: David

last: Sherman


title: Transcriptional profiling of Mycobacterium tuberculosis exposed to caseum mimics

summary: We profiled the transcriptional response of Mycobacterium tuberculosis exposed to different caseum mimic conditions using RNA-seq. We also hypothesized that an improved Mtb TRN could be inferred using a large, diverse compendium of RNA-seq data and therefore sought to supplement the existing repository of Mtb expression data available on SRA with additional conditions that could inform on Mtb's underlying regulatory programs. We went on to infer Mtb's TRN using a collection of inference methods in combination with this compendium, and employed it to build an interpretable machine learning model of Mtb fitness under stress.

overall_design: RNA-seq analysis was performed on samples taken from Mtb cultures subjected to various caseum mimic stresses.

first: Richard

last: Jones Jr


title: Phase variation in Mycobacterium bovis frd operon disrupts the succinate/HIF-1α/IL-1β axis to drive immunometabolic rewiring and enhance pathogenicity [RNA-seq]

city: BEIJING

first: Yuhui

last: Dong

state: Select

address: HaiDian District, Mingyuanxilu No.2

department: College of Veterinary Medicine

institute: China Agricultural University

summary: Tuberculosis (TB), caused by Mycobacterium tuberculosis complex (MTBC) pathogens, remains a global health threat. While bacterial genetic adaptations during host adaptation are poorly understood, phase variation in genomic homopolymeric tracts (HT) may drive pathogenicity evolution. Here, we demonstrate that M. bovis exploits HT insertion mutations in the fumarate reductase-encoding frd operon to subvert host immunometabolism. This study investigates the immunometabolic consequences of frd operon mutations in M. bovis-infected macrophages. RNA sequencing (RNA-seq) was performed on mouse bone marrow-derived macrophages (BMDMs) infected with wild-type M. bovis or its isogenic Δfrd mutant for 12 hours. Comparative transcriptomic analysis identified 192 upregulated and 344 downregulated genes in Δfrd-infected BMDMs, revealing significant suppression of innate immune pathways (e.g., defense response to Gram-positive bacteria, hypoxia adaptation) and upregulation of neutrophil chemotaxis/IL-17 signaling. Notably, Δfrd infection downregulated Gapdh (a glycolytic gatekeeper) and disrupted glucose metabolism-related pathways.

overall_design: RNA-seq was performed on BMDMs infected with wild-type M. bovis or its isogenic Δfrd mutant for 12 hours.

first: Xiangmei

last: Zhou


title: Transcriptional profiling of Mycobacterium tuberculosis exposed to antibiotic treatment in broth culture.

summary: We profiled  Mtb transcriptional response to a range of different antibiotic drug exposures in 7H9 broth culture by RNA-seq. Compound abbreviations: (AMK = amikacin, BDQ = bedaquiline, CPF = ciprofloxacin, LVF = levofloxacin, MXF = moxifloxacin, CFZ = clofazimine, PA824 = pretomanid, RPE = rifapentine, PZA = pyrazinamide). We also hypothesized that an improved Mycobacterium tuberculosis (Mtb) transcriptional regulatory network (TRN) could be inferred using a large, diverse compendium of RNA-seq data and therefore sought to supplement the existing repository of Mtb expression data available on SRA with additional conditions that could inform on Mtb's underlying regulatory programs. We went on to infer Mtb's TRN using a collection of inference methods in combination with this compendium, and employed it to build an interpretable machine learning model of Mtb fitness under stress.


title: Profiling Mtb expression during hypoxia and reaeration by RNA-seq

summary: We hypothesized that transcription factor activities found to be predictive of overall Mycobacterium tuberculosis (Mtb) fitness in a pooled culture of transcription factor induction strains could also be used to predict growth phenotypes under other growth conditions using interpretable machine learning models. To test, we measured Mtb transcriptional responses grown under hypoxic and reaeration stress without recombinant transcription factor perturbation as input data for our modeling.

overall_design: RNA-seq analysis was performed on samples taken from cultures subjected to a 12-day hypoxia and reaeration timecourse. Strains were cultured in oxygen-limited conditions for 7 days, followed by reaeration on days 7-12. Four biological replicates were maintained and sampled.


title: Transcriptional profiling of Mycobacterium tuberculosis exposed to proton pump inhibitor treatment in broth culture.

summary: We hypothesized that an improved Mtb TRN could be inferred using a large, diverse compendium of RNA-seq data and therefore sought to supplement the existing repository of Mtb expression data available on SRA with additional conditions that could inform on Mtb's underlying regulatory programs. We profiled Mtb transcriptional response to a range of different proton bump inhibitor exposures in 7H9 broth culture by RNA-seq. Compound abbreviations: (PAN = pantoprazole, RAB = rabeprazole, VER = verapamil, LAN = lansoprazole, OMP = omeprazole).  We went on to infer Mtb's TRN using a collection of inference methods in combination with this compendium, and employed it to build an interpretable machine learning model of Mtb fitness under stress.

first: Kristin

middle: N

last: Adams


title: The Hira complex regulates Gli3R-dependent transcription in Hedgehog signaling and medulloblastoma cell growth and migration

status: Public on Mar 18 2025

city: Warsaw

first: Weronika

last: Skarzynska

address: Banacha 2c

country: Poland

institute: Centre of New Technologies

summary: Regulation of the Hedgehog pathway activity may be supported by coactivators and corepresors of its main effectors- Gli transcription factors. While activation processes are well studied, repression mechanisms remain elusive. We identified chromatin remodelling complex Hira to interact with Gli3R protein, showed that its loss-of-function changes Hh pathway activity, and examined possible mechanism behind the observed effect. We also established that Hira influences the viability and migratory abilities of Hh-dependent medulloblastoma Daoy cells. Our study paves the way for a better understanding of processes involved in Hh pathway regulation and Hh-dependent carcinogenesis. This work was supported by grant from the National Science Centre (NCN): PRELUDIUM 2022/45/N/NZ3/02113.

overall_design: CUT&RUN experiment in NIH/3T3 mouse cell line with doxocycline inducible (Dox) expression of HA-Gli3R and Hira protein loss-of-function

first: Brygida

last: Baran

first: Pawek

last: Niewiadomski


title: The Hira complex regulates Gli3R-dependent transcription in Hedgehog signaling and medulloblastoma cell growth and migration [RNA-seq]

overall_design: RNAseq experiment in Daoy human cell line in Hira protein loss-of-function condition

first: Pawel


first: Nitin

last: Shirole

overall_design: Chromatin immunoprecipitation DNA sequencing (ChIP-Seq) to identify HIF2a genomic loci. We performed ChIP-Seq using an anti-FLAG antibody in OSRC2 cells in which CRISPR/Cas9 was used to append a FLAG-HA epitope tag to the N-terminus of HIF2a expressed from the endogenous HIF2a locus.

first: Sudeepa

last: Syamala

first: Shweta

last: Kukreja

first: Rong

last: Li

first: Xintao

last: Qiu

first: Paloma

last: Cejas

first: Henry

middle: W

last: Long

first: William

last: Kaelin


title: Concurrent loss of the Y chromosome in cancer and T cells impacts outcome [Bulk RNA-seq CRIPR]

city: Baltimore

first: XINGYU

last: CHEN

state: MD

address: 3501 St Paul St, Baltimore, MD 21218

department: Department of Urology

institute: Cedars-Sinai Medical Center

summary: Loss of the Y chromosome (LOY) in peripheral blood mononuclear cells (PBMCs) is the most common somatic alteration in normal men and is associated with greater mortality from epithelial cancers. In tumors, epithelial cell LOY is also associated with worse survival. This raises the fundamental question of whether a relationship between LOY in PBMCs, PBMC-derived immune cells, and cancer cells exists, and if so, what the consequences are. We addressed this through a comprehensive pan-cancer analysis of bulk and single-cell RNA sequencing data from 29 distinct human tumor types, and from autochthonous and syngeneic murine models of cancer. In both human and murine tumors, malignant epithelial cells had the highest LOY prevalence, which was associated with significant phenotypic changes. Remarkably, LOY was also found in stromal and immune cells of the tumor microenvironment (TME) of both species, with the presence of LOY in malignant epithelial cells predicting LOY in benign cells. In addition, LOY was correlated between paired tumor and PBMC samples from patients. Among benign cells, LOY drove the strongest phenotypic shift in CD4+ T and CD8+ T cells, with both showing signs of immunosuppression. The magnitudes of LOY in epithelial cells, CD4+ T cells, and CD8+ T cells were independent predictors of overall survival, with tumors exhibiting concurrent epithelial and T cell LOY having the worst outcomes. These findings provide a novel model linking LOY in immune and malignant epithelial cells and explain in part why LOY in PBMCs and tumors are both associated with greater cancer mortality.

overall_design: Bulk RNA-seq: CRISPR Y-KO and CRISPR Y-Scr MB49 Y⁺ cell lines (FASTQ)

first: Xingyu

last: Chen

first: Yiling

last: Shen

first: Suhyeon

last: Choi

first: Mukta

last: Basu

first: Lena

last: Hoelzen

first: Martina

last: Tufano

first: Hany

last: Abdel-Hafiz

first: Saravana

middle: K

last: Kailasam Mani

first: Maryam

last: Ranjpour

first: Jiani

last: Zhu

first: Krishnan

middle: V

last: Ramanujan

first: Ekaterina

last: Koltsova

first: Vinicius

last: Calsavara

first: Simon

last: Knott

first: Dan

last: Theodorescu


title: Concurrent loss of the Y chromosome in cancer and T cells impacts outcome [scRNA-seq MB49]

overall_design: scRNA-seq (MB49): MB49 clone 5 (C5) tumors implanted in C57Bl/6J mice (FASTQ).


title: Concurrent loss of the Y chromosome in cancer and T cells impacts outcome [Bulk RNA-seq FACS]

overall_design: Bulk RNA-seq: FACS-isolated CD45⁻ cells from CRISPR Y-KO vs. CRISPR Y-Scr MB49 tumors (FASTQ).


first: Nitin

last: Shirole


title: DNA microarray analysis using jejunum from wild-type or IAP knockout (Akp3-/-) mice administered with IIAEK (Ile-Ile-Ala-Glu-Lys)

status: Public on Mar 17 2025

city: Gifu

first: Satoshi

last: Nagaoka

address: 1-1 Yanagido

institute: Gifu university

summary: To understand the molecular mechanism by which IIAEK ameliorates hepatic and intestinal cholesterol metabolism, we performed DNA microarray analysis using liver, duodenal, and jejunal samples from the control and IIAEK groups of WT or IAP KO mice. We found that 3503 transcripts were identified in the jejunum of WT mice [WT, Control (WC) vs. WT, IIAEK (WI)] and 1388 in IAP-KO mice [IAP-KO, Control (KOC) vs. IAP-KO, IIAEK (KOI)] ( ≥1.2 fold-change, p < 0.05).

overall_design: Seven week-old wild type (WT) and IAP knockout (IAP-KO) (Akp3-/-) mice were administered orally the pentapeptide IIAEK (Ile-Ile-Ala-Glu-Lys) (600 mg/kg B.W./d) once a day for 14 d. After fasting for 8 h, the jejunum was collected from the mice for DNA microarray analyses (n=3/group).


title: Concurrent loss of the Y chromosome in cancer and T cells impacts outcome [WES]

first: XINGYU

last: CHEN

overall_design: Whole-Exome Sequencing (WES): CRISPR Y-KO and CRISPR Y-Scr (control) MB49 Y⁺ cell lines (FASTQ).

first: Xingyu

last: Chen


title: Humanized Mouse Model Reveals T-Cell ANXA2 as a Potential Therapeutic Target in Ischemic Stroke

city: Guangzhou

first: Tianyuan

last: Zhang

address: The First Affiliated Hospital of Jinan University

institute: Jinan university

summary: human peripheral blood mononuclear cells (PBMCs) were transferred into immunodeficient mice to generate a mouse model with humanized T cells, and this model was applied to a stroke model. We used single-cell RNA sequencing (scRNA-seq) to reveal the heterogeneity of humanized T cells and other mouse immune cells in the humanized mouse model

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: immunce cells (including humanzied T cells and mouse immunce cells) of the humanized mouse were isolated by fluorescence-activated cell sorting (FACS) and analyzed using scRNAseq.

first: Xiaojuan

last: Zheng

first: Aijun

last: Lu

first: Shuyuan

last: Li

first: Keshen

first: Xiaoxiong

last: Huang

first: Yanfang

last: Liu

first: Wei

first: Shengming

first: Jingchong

first: Niu

last: He

first: Chi

last: Tsang

first: Hongcheng

last: Mai

first: Anding

last: Xu


title: DNA microarray analysis using liver from wild-type or IAP knockout (Akp3-/-) mice administered with IIAEK (Ile-Ile-Ala-Glu-Lys)

first: Satoshi

summary: To understand the molecular mechanism by which IIAEK ameliorates hepatic and intestinal cholesterol metabolism, we performed DNA microarray analysis using liver, duodenal, and jejunal samples from the control and IIAEK groups of WT or IAP KO mice. We found that 1995 transcripts were identified in the livers of WT mice [WT, Control (WC) vs. WT, IIAEK (WI)] and 3802 in IAP-KO mice [IAP-KO, Control (KOC) vs. IAP-KO, IIAEK (KOI)] ( ≥1.2 fold-change, p < 0.05). Of these, 1729 transcripts fluctuated only in the WT group (WC vs. WI) and 3536 fluctuated only in the IAP-KO group (KOC vs. KOI).

overall_design: Seven week-old wild type (WT) and IAP knockout (IAP-KO) (Akp3-/-) mice were administered orally the pentapeptide IIAEK (Ile-Ile-Ala-Glu-Lys) (600 mg/kg B.W./d) once a day for 14 d. After fasting for 8 h, the liver was collected from the mice for DNA microarray analyses (n=3/group).


title: BMAL1-HIF2α heterodimers contribute to ccRCC [ChIP-seq]

city: SAN DIEGO

first: Rebecca

last: Mello

state: CA

address: 11093 CLAIREMONT MESA BLVD

institute: The Scripps Research Institute

summary: Circadian disruption enhances cancer risk, and many tumors exhibit disordered circadian gene expression. We show rhythmic gene expression is unexpectedly robust in clear cell renal cell carcinoma (ccRCC). Furthermore, the clock gene BMAL1 is higher in ccRCC than in healthy kidneys, unlike in other tumor types. BMAL1 is closely related to ARNT and we show that BMAL1-HIF2α regulates a subset of HIF2α target genes in ccRCC cells. Depletion of BMAL1 reprograms HIF2α chromatin association and target gene expression and reduces ccRCC growth in culture and in xenografts. Analysis of pre-existing data reveals higher BMAL1 in patient-derived xenografts that are sensitive to growth suppression by HIF2α antagonists. We show that increasing BMAL1 sensitizes ccRCC-derived 786O cells to growth inhibition by PT2399. Together, these findings indicate that an alternate HIF2α heterodimer containing the circadian partner BMAL1 contributes to HIF2α activity, growth, and sensitivity to HIF2α antagonist drugs in ccRCC cells.

overall_design: To characterize the localizations of endogenous BMAL1 and HIF2α in native chromatin, we sequenced genomic DNA associated with BMAL1 or HIF2α in 786O cells. Chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq) were performed using CUT&RUN assay kit (CST #86652) following the manufacturer protocol. 100,000 786O cells infected with lentivirus expressing shRNA targeting “scramble” or BMAL1 were used for each reaction.

first: Katja

last: Lamia


title: ScRNA-seq of Small Intestine Neuroendocrine Tumors

city: Rehovot

first: Einav

last: Somech

address: Hertzel 234

country: Israel

institute: Weizmann

summary: Here we profiled ten low-grade small intestine NET (SiNET) tumor samples as well as one mixed lung tumor by single-cell or single-nuclei RNA-seq. We find that SiNETs are largely separated into two distinct subtypes, in which the neuroendocrine cells upregulate epithelial or neuronal markers, respectively. Surprisingly, in both subtypes the neuroendocrine cells are largely non-proliferative while higher proliferation is observed in multiple non-malignant cell types. Specifically, B and plasma cells are highly proliferative in the epithelial-like SiNET subtype, potentially reflecting the outcome of high Migration Inhibitory Factor (MIF) expression in those tumors, which may constitute a relevant target. Finally, our analysis of a mixed lung neuroendocrine tumor identifies a population of putative progenitor cells that may give rise to both neuroendocrine and non-neuroendocrine (squamous) cells, potentially explaining the origin of the mixed histology. Taken together, our results provide important insights and hypotheses regarding the biology of neuroendocrine neoplasms.

overall_design:
Fresh tumor samples were collected at the time of surgery and processed immediately. Tumors were minced and enzymatically dissociated using a tumor dissociation kit (Miltenyi Biotec) in a GentleMACS Octo-dissociator at low speed. A single-cell suspension was obtained by filtering through a 60-100 µm cell strainer, followed by RBC removal (Roche) and dead cell removal (Miltenyi). Cell viability and density were assessed using Trypan blue staining and a Countess II automated hemocytometer. 10x Genomics Single-Cell Chromium Controller was used to load 8,000-10,000 cells per channel.  For frozen tumors, dissociation was performed with nuclei isolation using ST-based buffers (Slyper et al.) or the EZ-lysis method (Sigma). Tissue was homogenized in ice-cold buffer using a Dounce tissue grinder (Sigma), followed by centrifugation (500g, 4°C), washing, and resuspension in a PBS-based buffer with BSA and RNase inhibitor. Nuclei suspensions were filtered through a 40 µm strainer and counted, with a final concentration of 1,000 nuclei/µl for 10x Chromium loading.  For library preparation, the 10x Genomics Chromium Next GEM Single Cell 3′ GEM, Library & Gel Bead Kit v3.1 was used for fresh tumor/single-cell samples, while the Chromium Next GEM Single 5′ GEM was used for frozen tumor/single-nuclei samples. Libraries were generated following the standard 10x Genomics single-cell RNA-seq protocol, including reverse transcription, cDNA amplification, fragmentation, and adapter/sample index attachment.  Sequencing was performed on an Illumina NovaSeq-6000 using either:      SP-100 kit (2 pooled libraries per lane)     S1-100 kit (4 pooled libraries per two lanes)     Read lengths: Read 1: 26 nt, Read 2: 55 nt, Index 1: 8 nt, Index 2: 0 nt.

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

first: Itay

last: Tirosh

first: Debdatta

last: Hadler

first: Avishai

last: Spitzer

first: Chaya

last: Brabolin

first: Michael

last: Tyler

first: Reut

last: Halperin

first: Biton

last: Moshe

first: Amit


title: BMAL1-HIF2α heterodimers contribute to ccRCC

summary: Circadian disruption enhances cancer risk, and many tumors exhibit disordered circadian gene expression. We show rhythmic gene expression is unexpectedly robust in clear cell renal cell carcinoma (ccRCC). Furthermore, the clock gene BMAL1 is higher in ccRCC than in healthy kidneys, unlike in other tumor types. BMAL1 is closely related to ARNT and we show that BMAL1-HIF2α regulates a subset of HIF2α target genes in ccRCC cells. Depletion of BMAL1 reprograms HIF2α chromatin association and target gene expression and reduces ccRCC growth in culture and in xenografts. Analysis of pre-existing data reveals higher BMAL1 in patient-derived xenografts that are sensitive to growth suppression by HIF2α antagonists. We show that increasing BMAL1 sensitizes ccRCC-derived A498 cells to growth inhibition by PT2399. Together, these findings indicate that an alternate HIF2α heterodimer containing the circadian partner BMAL1 contributes to HIF2α activity, growth, and sensitivity to HIF2α antagonist drugs in ccRCC cells.

overall_design: To characterize the localizations of endogenous BMAL1 and HIF2α in native chromatin, we sequenced genomic DNA associated with BMAL1 or HIF2α in A498 cells. Chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq) were performed using CUT&RUN assay kit (CST #86652) following the manufacturer protocol. 100,000 A498 cells infected with lentivirus expressing shRNA targeting “scramble” or BMAL1 were used for each reaction.


title: Tumor-site directed A1R expression enhances CAR T cell function and improves efficacy against solid tumors. [human RNAseq constitutive]

status: Public on Mar 16 2025

city: Melbourne

first: Kevin

last: sek

state: VIC

address: 305 Grattan Street

department: Cancer Immunology

institute: Peter MacCallum Cancer Centre

summary: The efficacy of Chimeric Antigen Receptor (CAR) T cells against solid tumors is limited by immunosuppressive factors in the tumor microenvironment (TME) including adenosine, which suppresses CAR T cells through activation of the A2A receptor (A2AR). To overcome this, CAR T cells were engineered to express A1 receptor (A1R), a receptor that signals inversely to A2AR. Using murine and human CAR T cells, constitutive A1R overexpression was demonstrated to significantly enhance CAR T cell effector function albeit at the expense of CAR T cell persistence. Through a novel CRISPR/Cas9 “knock-in” approach we demonstrated that CAR T cells engineered to express A1R in a tumor-localized manner, led to enhanced anti-tumor efficacy dependent on the transcription factor IRF8 and was transcriptionally unique when compared to A2AR deletion. This data provides a novel approach for enhancing CAR T cell efficacy in solid tumors and provides proof of principle for site-directed expression of factors that promote effector T cell differentiation.

overall_design: RNA-seq profiling of NGFR control, A1R and A3R constitutively expressing human anti-LewisY CAR T cells were stimulated with OVCAR ovarian cancer cells for 16 hours. CAR-T cells were FACS sorted and RNA extracted using RNAeasy (Qiagen) kit as per manufacturer's instructions.

last: Sek


title: Tumor-site directed A1R expression enhances CAR T cell function and improves efficacy against solid tumors. [Mouse RNAseq constitutive]

overall_design: RNA-seq profiling of NGFR control, A1R and A3R constitutively expressing mouse anti-HER2 CAR T cells were stimulated with E0771-HER2 breast cancer cells for 16 hours. CAR-T cells were FACS sorted and RNA extracted using RNAeasy (Qiagen) kit as per manufacturer's instructions.


title: Tumor-site directed A1R expression enhances CAR T cell function and improves efficacy against solid tumors. [CD8 inducible human RNAseq]

overall_design: RNA-seq profiling of NR4A2 promoter driven inducible A1R CD8+ human anti-LewisY CAR T cells chronically stimualted with up to 3 rounds of OVCAR ovarian tumors over 72 hours. CAR-T cells were FACS sorted and RNA extracted using RNAeasy (Qiagen) kit as per manufacturer's insdtructions.


title: Tumor-site directed A1R expression enhances CAR T cell function and improves efficacy against solid tumors. [human ATACseq]

overall_design: ATAC-seq profiling of Mock control and NR4A2/A1R human anti-LewisY CAR T cells stimulated with OVCAR ovarian cancer cells for 16 hours. CAR-T cells were FACS sorted and DNA extracted.


title: RNA sequencing analysis of mouse kidney fibroblasts isolated from control kidneys or UUO surgery operated kidneys

city: Toronto

first: Andrew

last: Advani

state: Ontario

address: 6-151 61 Queen Street East

department: Medicine

institute: St. Michael's Hospital

summary: RNA sequencing of fibroblasts isolated from mouse kidneys which were either subjected to left ureter obstruction (UUO) or non obstructed (Control) conditions

overall_design: RNA isolated from primary kidney fibroblasts collected from normal kidneys or UUO kidneys 3 or 7 days after the surgery

first: Harmandeep

last: Kaur


title: NEST: Spatially-mapped cell-cell communication patterns using a deep learning-based attention mechanism

first: Faiyaz

last: Notta

address: 101 College Street

department: Research

institute: University Health Network

summary: Dysregulation of communication between cells mediates complex diseases such as cancer and diabetes. However, detecting cell-cell communication (CCC) at scale remains one of the greatest challenges in transcriptomics. While gene expression remains one of the greatest challenges in transcriptomics. While gene expression measured with single-cell RNA sequencing and spatial transcriptomics reinvigorated computational approaches to detecting CCC, most existing methods exhibit high false positive rates, do not integrate spatial proximity of ligand-receptor interactions, and cannot detect CCC between individual  cells. We overcome these challenges by presenting NEST (NEural network on Spatial  Transcriptomics), which uses a graph attention network paired with an unsupervised contrastive learning approach to decipher patterns of communication while retaining the strength of each signal. We introduce new synthetic benchmarking experiments which demonstrate how NEST outperforms existing tools and detects biologically-relevant CCC along with directionality and confidence across spot- and cell-based technologies measuring several different tissues and diseases. In our applications, NEST identifies T-cell homing signals in human lymph nodes, aggressive cancer CCC in lung adenocarcinoma, and discovers new patterns of communication in pancreatic cancer. Beyond two-dimensional data, we also highlight NEST's ability to detect CCC in three-dimensional spatial transcriptomic data.

overall_design: Two patient samples with resectable (Stage IIB) PDAC were included (Male, 76 years old and Female, 83 years old).  Both patient samples were collected from the University Heath Network Biospecimens Program (Toronto, Canada). Ethical approval was obtained through the University Health Network Research Ethics Board (13-6377).

first: Fatema

last: Tuz Zohora

first: Eugenia

last: Flores-Figueroa

first: Joshua

last: Li

first: Deisha

last: Paliwal

first: Gregory

last: Schwartz


title: Enhancer adoption by an LTR retrotransposon generates viral-like particles causing developmental limb phenotypes [cHiC]

status: Public on Mar 15 2025

city: Berlin

first: Juliane

last: Glaser

address: Ihnestrasse 63-73

department: Development and Disease

institute: Max Planck Institute for Molecular Genetics

summary: Mammalian genomes are scattered with transposable elements (TEs). Silencing of TEs prevents harmful effects caused by either global activation leading to genome instability or insertional mutation disturbing gene transcription. However, whether the activation of a TE can be pathological without directly affecting gene expression is largely unknown. Here, we show that a TE insertion can adopt nearby regulatory activity, producing cell-type-specific viral-like particles (VLPs) in the embryo and affecting organ formation. Failure to silence an LTR retrotransposon inserted upstream of the Fgf8 gene results in their co-expression during development. VLPs assembly in the Fgf8-expressing cells of the developing limb triggers apoptotic cell death, resulting in a limb malformation resembling human ectrodactyly. We rescued this phenotype with mutations in the retrotransposon coding sequence, preventing its full endogenous retroviral cycle. Our findings illustrate how TE insertion is incorporated into the local genomic regulatory landscape and show that VLP production in post-implantation embryos can be pathological.

overall_design:
We study how the insertion of an endogenous retrovitrus lead to a developmental malformation. Using the Dactylaplasia mouse model which carry a MusD ERV insertion at the Fgf8 locus, we carried out capture-HiC, CTCF Chip-seq, 4C-seq, single-cell RNA-sequencing and bulk RNA-seq to identify change in the 3D genome and expression of the MusD mRNA. We further confirm that the MusD retroviral proteins and particles are produced and lead to cell death, causing a string limb phenotype.
Please note that processed data was generated from all Rep* and techRep* samples, and is linked to the corresponding *Rep1 sample records.


title: Enhancer adoption by an LTR retrotransposon generates viral-like particles causing developmental limb phenotypesv [RNA-seq]

overall_design: We study how the insertion of an endogenous retrovitrus lead to a developmental malformation. Using the Dactylaplasia mouse model which carry a MusD ERV insertion at the Fgf8 locus, we carried out capture-HiC, CTCF Chip-seq, 4C-seq, single-cell RNA-sequencing and bulk RNA-seq to identify change in the 3D genome and expression of the MusD mRNA. We further confirm that the MusD retroviral proteins and particles are produced and lead to cell death, causing a string limb phenotype.


title: Enhancer adoption by an LTR retrotransposon generates viral-like particles causing developmental limb phenotypes [ChIP-seq]


title: Enhancer adoption by an LTR retrotransposon generates viral-like particles causing developmental limb phenotypes [scRNA-seq]


title: DNA microarray analysis using duodenum from wild-type or IAP knockout (Akp3-/-) mice administered with IIAEK (Ile-Ile-Ala-Glu-Lys)

first: Satoshi

last: Nagaoka

summary: To understand the molecular mechanism by which IIAEK ameliorates hepatic and intestinal cholesterol metabolism, we performed DNA microarray analysis using liver, duodenal, and jejunal samples from the control and IIAEK groups of WT or IAP KO mice. In total, 1406 transcripts were identified in the duodenum of WT mice ( ≥1.2 fold-change, p < 0.05).  In contrast, we identified  777 transcripts in the duodenum of IAP-KO mice ( ≥1.2 fold-change, p < 0.05).

overall_design: Seven week-old wild type (WT) and IAP knockout (IAP-KO) (Akp3-/-) mice were administered orally the pentapeptide IIAEK (Ile-Ile-Ala-Glu-Lys) (600 mg/kg B.W./d) once a day for 14 d. After fasting for 8 h, the duodenum was collected from the mice for DNA microarray analyses (n=3/group).

first: Satosh

last: i Nagaoka


title: Amniotic Fluid Reduces Liver Fibrosis By Attenuating Hepatic Stellate Cell Activation

city: Galveston

first: W. Samuel

last: Fagg

state: TX

address: 301 University Blvd

department: Department of Surgery

institute: University of Texas Medical Branch

summary: Background & Aims: Persistent activation of hepatic stellate cells (HSCs) drives liver fibrosis, marked by myofibroblast activation (MFA) and epithelial-mesenchymal transition (EMT). Strategies to prevent or reverse this will be critical to treat liver fibrosis successfully. We previously showed that full-term, cell-free human amniotic fluid (cfAF) inhibits MFA and EMT in fibroblasts in vitro. We hypothesize that cfAF treatment can attenuate HSC activation and limit liver fibrosis.  Methods: HSC activation was assessed using murine models of acute (DMN) and chronic (CCl4) liver damage, three-dimensional hepatic spheroids, and HSC cultures. EMT and MFA were induced in vitro using ethanol (spheroids) or TGFβ (HSCs) and evaluated via scratch assays, multi-omics approaches, and RNA/protein analyses.  Results: cfAF treatment prevented weight loss in mice with chronic liver damage without adverse events. Histological analysis revealed a modest reduction in fibrosis with preserved liver architecture in both chronic and acute murine liver damage models. Gene expression profiling and immunostaining indicated cfAF administration to CCl4-treated mice led to reduced EMT- and MFA-related biomarkers along with changes in transcripts associated with liver metabolism, immune regulatory pathways, and cell signaling. In hepatic spheroids exposed to ethanol cfAF treatment lowered COL1A1 protein levels and smooth muscle actin (SMA) RNA abundance in a dose-dependent manner. Treating primary or LX2 hepatic stellate cells with cfAF strongly represses EMT, and multi-omics analyses revealed that it also attenuates TGFβ-induced MFA and the inflammatory phenotype. Thus cfAF treatment prevents liver fibrosis by safeguarding hepatic stellate cell activation.  Conclusions: cfAF treatment limits liver fibrosis by repressing HSC activation in liver fibrosis models. These findings suggest cfAF may be a safe and effective therapy for reducing liver fibrosis and preventing the development of cirrhosis and/or hepatocellular carcinoma.

overall_design: RNA-seq to measure transcript abundance changes in human LX2 hepatic stellate cells, either cultured with or without cell-free human amniotic fluid (cfAF) and in the presence of absence of TGF-beta (and/or with TGF-beta blocker A-83-01)

first: Charles

last: Bowen

first: Frederick

middle: S

last: Ditmars

first: Naiyou

last: Liu

first: Jose

last: Abril

first: David

last: Ajasin

first: William

last: Russell

first: Heather

last: Stevenson

first: Eliseo

last: Eugenin

first: Jeffrey

last: Fair


title: Expression data from induced pluripotent stem cell-derived crypt-villus structural small intestine

status: Public on Mar 10 2025

city: Tokyo

first: Kyosuke

last: Kobayashi

address: 1-29-1 Nanakuni, Hachioji

department: Wellness Science Labs

institute: Meiji Holdings Co., Ltd.

summary:
We established the co-culture model of iPS cell-derived crypt-villus structural small intestine and lactic acid bacteria (LABs).
Using this model, the effect of TNF-α, IFN-γ and LABs (Lactobacillus delbrueckii subsp. bulgaricus 2038 and Streptococcus thermophilus 1131) on the cells was evaluated.

overall_design: We examined 4 experimental groups: control, TNF-α + IFN-γ, TNF-α + IFN-γ + L. bulgaricus 2038 and TNF-α + IFN-γ + S. thermophilus 1131 group. In TNF-α + IFN-γ group, cytokines were added and the cells were incubated for 42 hours. In the lactic acid bacteria (LABs)-added group, after 24 hours of cytokine stimulation, LABs were added and the cells were incubated for 18 hours.

first: Yuri

last: Imai

first: Yuka

last: Shimizu

first: Isamu

last: Ogawa

first: Takaaki

last: Nakai

last: Mizuno

first: Yoshika

last: Kaneda

first: Yumiko

last: Yasuoka

first: Fuka

last: Yamazaki

first: Junko

last: Mochizuki

first: Toshihiro

last: Sashihara

first: Tamihide

last: Matsunaga

first: Takahiro

last: Iwao


title: variant Polycomb Repressive complexes 1 (vPRC1s) in synovial sarcoma

city: Edinburgh

first: Nezha

middle: Suzanne

last: Benabdallah

state: Midlothian

address: Crewe Road South

department: Institute of Genetics and Cancer

institute: University of Edinburgh

summary: Several variant Polycomb Repressive complexes 1 (vPRC1s) co-exist to robustly deposit mono ubiquitination on histone H2A (H2AK119ub1). Their interconnected roles and specific tasks across normal and disease cellular states remain unclear. In synovial sarcoma (SS), a soft tissue tumor driven by the oncogenic fusion protein, SS18::SSX, the SSX tail exhibits a high affinity to H2AK119ub1 marked genomic regions. This mechanism is crucial to sequester the mSWI/SNF complex to PRC1 targets and results in their aberrant expression. We previously showed that PRC1.1 is responsible for H2AK119ub1 deposition in SS and is therefore required for SS18::SSX occupancy. Here, we investigated the role of PRC1 variants in synovial sarcoma.

overall_design: Total RNA from two independent experiments was extracted using a RNeasy mini kit (Qiagen). RNA-Seq library preparation and sequencing were performed at the High Throughput Sequencing Unit of the Genomics & Proteomics core facility of DKFZ using TruSeq Stranded Total RNA Library Prep kit (Illumina) and HiSeq 2000 v4 single-read 50bp sequencing system.

first: Vineet

last: Dalal

first: Ana

last: Banito


title: Pooled tagging and hydrophobic targeting of endogenous proteins for unbiased mapping of unfolded protein responses

city: Philadelphia

first: david

last: smith

state: Pennsylvania

address: 3501 Civic Center Blvd

institute: CHOP

summary: To achieve system-level insights into proteome organization, regulation, and function, we developed an approach to generate complex cell pools with endogenously tagged proteins amenable to high-throughput visualization and perturbation. Pooled imaging coupled to in-situ barcode sequencing identified the subcellular localization of each HaloTag tagged protein, and subsequent ligand-induced misfolding of the library followed by single cell RNA sequencing revealed responses to spatially restricted protein misfolding. These unique datasets characterized protein quality control responses in previously uninterrogated cellular compartments, and cross-compartment analyses revealed mutually exclusive rather than collaborative responses, whereby the Heat Shock Response (HSR) is induced in some compartments and repressed in others where autophagy genes are induced. We further assign protein quality control functions of previously uncharacterized genes based on the similarity of the transcriptional responses to protein misfolding across cellular compartments. Altogether, we present a powerful and efficient method for large-scale studies of proteome dynamics, function, and homeostasis.

overall_design: HEK293 cells were transfected with a library sgRNA. This constitutes the "untag" population. Upon introduction of a donor construct, genes will be edited to contain a HaloTAG. In the presence of the HyT36 ligand, the edited, translated gene prodeuct will misfold. A control population of cells received DMSO rather than HyT36.

first: Stephanie

last: Sansbury

first: Yevgeniy

last: Serebrenik

first: Tomer

last: Lapidot

first: David

last: Smith

first: George

middle: M

last: Burslem

first: Ophir

last: Shalem


title: Tumor-site directed A1R expression enhances CAR T cell function and improves efficacy against solid tumors. [Human RNAseq inducible]

first: Kevin

last: sek

overall_design: RNA-seq profiling of NR4A2 promoter driven inducible A1R CD8+ human anti-LewisY CAR T cells chronically stimulated with up to 3 rounds of OVCAR ovarian tumors over 72 hours in the presence or absence of A2AR agonist NECA (10uM). CAR-T cells were FACS sorted and RNA extracted using RNAeasy (Qiagen) kit as per manufacturer's instructions.

last: Sek


title: Enhancer adoption by an LTR retrotransposon generates viral-like particles causing developmental limb phenotypes [4C]

first: Juliane

last: Glaser

overall_design: We study how the insertion of an endogenous retrovirus lead to a developmental malformation. Using the Dactylaplasia mouse model which carries a MusD ERV insertion at the Fgf8 locus, we carried out capture-HiC, CTCF Chip-seq, 4C-seq, single-cell RNA-sequencing, and bulk RNA-seq to identify the change in the 3D genome and expression of the MusD mRNA. We further confirm that the MusD retroviral proteins and particles are produced and lead to cell death, causing limb phenotypes.


title: Spatially separated intraepithelial and lamina propria neutrophils present distinct functional identities in the inflamed colon mucosa

first: Parambir

last: Dulai

state: IL

address: 259 E Erie St Ste 1600

institute: Northwestern University

summary: We used GeoMx DSP technology to profile the gene expression information of intraepithelial neutrophils (IENs) and lamina propria neutrophils (LPNs) from moderate-severe UC patients's colon.

overall_design: FFPE blockes were obtained from moderate-severe UC patients colon. Slides were processed for GeoMx processing under the manufactor instruction; panCK and Elastase were used to mark the area of interest.


title: Macrophage phenotypes modulate neoangiogenesis and fibroblast profiles in synovial-like organoid cultures.

city: Palo Alto

first: Qi

last: Gao

address: 240 Pasteur Dr., BMI building Room 0200E, Department of Orthopaedic Surgery, Stanford University, BMI building Room 0200E, Stanford University

institute: Stanford University

summary: To investigate the interaction among the cells thought to be foundational to inflammation, fibrosis, and angiogenesis in the synovial membrane.

overall_design: We encapsulated fibroblasts, polarized macrophages, and endothelial cells in a 3D culture system. We used this model to determine the cellular transcriptional profiles, cytokine secretion, and vascular formation associated with different macrophage phenotype conditions.

first: Xiurui

last: Zhang

first: Meagan

middle: J

last: Makarcyzk

first: Laurel

last: Wong

first: Madison

last: Quig

first: Issei

last: Shinohara

first: Masatoshi

last: Murayama

first: Simon

last: Chow

first: Bruce

last: Bunnell

first: Hang

last: Lin

first: Stuart

middle: B

last: Goodman


title: Unraveling cysteine deficiency-associated rapid weight loss [liver II]

city: New York

first: Ilya

last: Shamovsky

state: New York

address: 450 E 29th Street

department: Biochemistry and Molecular Pharmacology

institute: NYU School of Medicine

summary: Forty percent of the US population and 1 in 6 individuals worldwide are obese, with the incidence surging globally1,2. Various dietary interventions, including carbohydrate, fat and more recently amino acid restriction, have been explored to combat this epidemic3-6. We investigated the impact of removing individual amino acids on the weight profiles of mice. Here, we show that conditional cysteine restriction resulted in the most dramatic weight loss when compared to essential amino acid restriction, amounting to 30% within one week, which was readily reversed. We found that cysteine deficiency activated the integrated stress response and oxidative stress response, which amplify each other, leading to the induction of GDF15 and FGF21, which partly explained the phenotype7-9. Surprisingly, we observed lower tissue coenzyme A (CoA), which has been considered to be extremely stable10, resulting in reduced mitochondrial functionality and metabolic rewiring. This results in energetically inefficient anaerobic glycolysis and defective TCA cycle, with sustained urinary excretion of pyruvate, orotate, citrate, α-ketoglutarate, nitrogen rich compounds, and amino acids In summary, our investigation reveals that cysteine restriction, by depleting GSH and CoA, exerts a maximal impact on weight loss, metabolism, and stress signaling compared to other amino acid restrictions. These findings suggest novel strategies for addressing a range of metabolic diseases and the growing obesity crisis.

overall_design: Various tissues were collected from either WT or cse KO mice kept on different diets and total RNA was isolated by trizol method.

first: Alan

last: Varghese

first: Ivan

last: Gusarov


title: Unraveling cysteine deficiency-associated rapid weight loss

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

overall_design: Refer to individual Series


title: Transcriptional changes of the extracellular matrix in chronic thromboembolic pulmonary hypertension governs right ventricle remodeling and recovery [ext300_rnaseq]

city: Bad Nauheim

first: Carsten

last: Kuenne

address: Ludwigstrasse 43

department: Bioinformatics

institute: Max Planck Institute for Heart and Lung Research

summary: Tissue samples from healthy controls were collected from the same patient (n = 10) for both Right ventricle and septum. The tissue samples were collected in the Department of Heart Failure and Transplantology, National Institute of Cardiology, Warsaw, Poland.

overall_design: To decipher the molecular mechanisms underlying RV remodeling in CTEPH, the transcriptomic profile of the RV and septum, were compared with those of a normal control group.

first: Christoph

last: Liebetrau

first: Soni

last: Pullamsetti

first: Eckhard

last: Mayer

first: Peter

last: Dorfmüller

first: Leili

last: Jafari

first: Christian

last: Troidl

first: Stefan

last: Guenther


title: Adult Sprague-Dawley Rat intestinal epithelium

city: New Haven

first: Kaelyn

last: Sumigray

state: CT

address: 333 Cedar St, SHM I 336A; Yale School of Medicine

department: Genetics

institute: Yale School of Medicine

summary: The proximal small intestine of humans and rats contain a distinct cell type not represented in mouse intestine. We used scRNA-seq to analyze the transcriptional profile of CFTR High Expressers (CHEs) from adult male Sprague Dawley rat jejunal epithelium compared to other cells that arise from the secretory lineage.

overall_design: The epithelial cells of the proximal jejunum of 2 150-200 g male Sprague Dawley rats were isolated using EDTA and dispase and analyzed using scRNA-seq.

first: Zachary

last: Smith


title: ZEB2 is a Master Switch Controlling the Tumor-Associated Macrophage Program

first: Ido

last: Amit

address: 234 Herzl st.

department: Immunology

institute: Weizmann Institute of Science


title: ZEB2 is a Master Switch Controlling the Tumor-Associated Macrophage Program [in vivo]

summary: Tumor-associated macrophages (TAMs) are major players in regulating the immunosuppressive tumor environment, and their abundance is highly correlated with poor clinical outcomes. Here, we constructed a TAM regulatory network by integrating scRNA-seq data across human solid tumors with a dedicated CRISPR knockout screen. Using a deep generative model capable of learning a local representation for each candidate regulator, we constructed a gene perturbation network that linked individual target genes with prototypical functional modules in TAMs. We identified non-redundant pathways regulating distinct TAM functions, showing modular circuitry. For instance, the complement module is repressed by Stat6, Zeb2, Gpnmb, and Spp1, while the Tgfbr1-Smad2/4 pathway induces this program in TAMs. Importantly, we identified Zeb2 as the master regulator of pro-tumor functions in TAMs, orchestrating the suppression of type I interferon response and antigen presentation alongside the activation of immune suppression programs. Genetic ablation of Zeb2 reprogrammed TAMs identity on chromatin, RNA, and protein levels. In human tumors with high macrophage content, ZEB2 expression was associated with poor prognosis in solid cancers, including lung and bladder. Functional macrophage coculturing assays defined Zeb2 as a critical regulator of TAM immunosuppression activity by inhibiting T cell proliferation and activation. Selective in vivo targeting of Zeb2 in macrophages using a CpGsiRNAZeb2 DNA hybrid reprogrammed TAMs and mobilized systematic anti-tumoral T cell responses, achieving complete tumor clearance as a monotherapy. Overall, our study generated a detailed genetic roadmap of TAM gene circuits and identified ZEB2 as a master switch of TAMs with potential therapeutic implications for macrophage-based immunotherapies.

overall_design: WT C57BL/6 mice were s.c. implanted with MC38 cells at the flanks. When tumors reached 80-100 mm^3, mice either received PBS or CpG-siRNA conjugates via intratumoral injections. Tumor-bearing mice were sacrificed at indicated timepoints and tumor-infiltrating leukocytes were extracted and single-cell suspensions were prepared for flow cytometry antibody staining and single-cell FACS index sorting into 384-well-plates. Single-cell RNAseq libraries were prepared with SPID-seq protocol  (Yofe et al., 2023) and sequenced with Illumina Novaseq.

first: Fadi

last: Sheban

first: Truong

last: San Phan

first: Ken

last: Xie

first: Florian

last: Ingelfinger

first: Chamutal

last: Gur

first: Yuval

last: Shapir Itai

first: Ronnie

last: Blecher-Gonen

first: Chunsong

last: Yu

first: Roberto

last: Avellino

first: Paulina

last: Chalan

last: Yofe

first: Vladimir

last: Yutkin

first: Pierre

last: Boyeau

first: Can

last: Ergen-Behr

first: Justin

last: Hong

first: Kfir

last: Mazuz

first: Rony

last: Dahan

first: Marcin

last: Kortylewski

first: Nir

last: Yosef

first: Assaf

last: Weiner


title: Unraveling cysteine deficiency-associated rapid weight loss [liver]

first: Ilya

last: Shamovsky

first: Alan

last: Varghese

first: Ivan

last: Gusarov


title: ZEB2 is a Master Switch Controlling the Tumor-Associated Macrophage Program [KO_screen]

first: Ido

last: Amit

overall_design: 8 weeks old CAS9-expressing C57BL/6 mice were sacrificed and bone marrow was isolated and cultured in the presence of human MCSF to generate BMDMs. Day 2 of culture, cells were transduced with lentiviruses expressing different guides in an arrayed screen fashion. At day 6, cells were treated with murine IL4 and isolated on day 7 for FACS sorting to SPID-seq scRNA-seq protocol.

first: Fadi

last: Sheban

first: Truong

last: San Phan

first: Ken

last: Xie

first: Florian

last: Ingelfinger

first: Chamutal

last: Gur

first: Yuval

last: Shapir Itai

last: Blecher-Gonen


title: Rapid and Inducible Mislocalization of Endogenous TDP43 in a Novel Human Model of Amyotrophic Lateral Sclerosis [Nanopore]

status: Public on Mar 07 2025

city: Exeter

first: Akshay

last: Bhinge

address: Stocker Road

department: CBS

institute: Living Systems Institute, University of Exeter

summary: In this study, we describe the development of a new model of TDP43 proteinopathy using human induced pluripotent stem cell (iPSC)-derived neurons. Utilizing a genome engineering approach, we induced the mislocalization of endogenous TDP43 from the nucleus to the cytoplasm without mutating the TDP43 gene or using chemical stressors

overall_design:
RNA was harvested from iPSC-derived motor neurons (MNs) displaying mislocalized or nuclear TDP43 at day 40 of the differentiation process.
RNA-seq was performed using ONT long-read sequencing.

first: Johanna

last: Ganssauge

first: Seema

last: Namboori

first: Szi Kay

last: Leung


title: Rapid and Inducible Mislocalization of Endogenous TDP43 in a Novel Human Model of Amyotrophic Lateral Sclerosis [RNA-seq]

overall_design: RNA was harvested from iPSC-derived motor neurons (MNs) displaying mislocalized or nuclear TDP43 at day 40 of the differentiation process.


title: Unraveling cysteine deficiency-associated rapid weight loss [adipose]


title: A subcellular map of translational machinery composition and regulation at the single-molecule level [ALIBi_Nmd3_Gnl3_KD]

status: Public on Mar 05 2025

first: Maria

last: Barna

address: 240 Pasteur Dr, BMI 4500

summary: Millions of ribosomes are packed within mammalian cells, yet we lack tools to visualize them in toto and characterize their subcellular composition. Here, we present ribosome expansion microscopy (RiboExM) to visualize individual ribosomes and an optogenetic proximity-labeling technique (ALIBi) to probe their composition. We generated a super-resolution ribosomal map, revealing unexpected subcellular translational hotspots and enrichment of 60S subunits near polysomes at the endoplasmic reticulum (ER). We found that Lsg1 tethers 60S to the ER and regulates translation of select proteins. Additionally, we discovered ribosome heterogeneity at mitochondria guiding translation of metabolism-related transcripts. Finally, we visualized ribosomes in neurons, revealing a dynamic switch between monosomes and polysomes in neuronal translation. Together, these approaches enable exploration of ribosomal localization and composition at unprecedented resolution.

overall_design: Rpl31-FLAG-TEV-AviTag-containing ribosomes were proximity biotinylated using ALIBi targeted to either the cytosol or endoplasmic reticulum. Biotinylated ribosomes were affinity purified and subjected to poly-A-enriched RNA-seq. Whole-cell RNA from the same cell lysates (input for affinity purification) was sequenced in parallel. In some cases, cells were treated with either a targeting or scrambled (control) siRNA prior to performing ALIBi-mediated enrichment of subcellular ribosome populations.

first: Adele

middle: F

last: Xu


title: Concurrent loss of the Y chromosome in cancer and T cells impacts outcome [scRNA-seq]

city: Baltimore

first: XINGYU

last: CHEN

address: 3501 St Paul St, Baltimore, MD 21218

institute: Cedars-Sinai Medical Center

summary: Loss of the Y chromosome (LOY) in peripheral blood mononuclear cells (PBMCs) is the most common somatic alteration in normal men and is associated with greater mortality from epithelial cancers. In tumors, epithelial cell LOY is also associated with worse survival. This raises the fundamental question of whether a relationship between LOY in PBMCs, PBMC-derived immune cells, and cancer cells exists, and if so, what the consequences are. We addressed this through a comprehensive pan-cancer analysis of bulk and single-cell RNA sequencing data from 29 distinct human tumor types, and from autochthonous and syngeneic murine models of cancer. In both human and murine tumors, malignant epithelial cells had the highest LOY prevalence, which was associated with significant phenotypic changes. Remarkably, LOY was also found in stromal and immune cells of the tumor microenvironment (TME) of both species, with the presence of LOY in malignant epithelial cells predicting LOY in benign cells. In addition, LOY was correlated between paired tumor and PBMC samples from patients. Among benign cells, LOY drove the strongest phenotypic shift in CD4+ T and CD8+ T cells, with both showing signs of immunosuppression. The magnitudes of LOY in epithelial cells, CD4+ T cells, and CD8+ T cells were independent predictors of overall survival, with tumors exhibiting concurrent epithelial and T cell LOY having the worst outcomes. These findings provide a novel model linking LOY in immune and malignant epithelial cells and explain in part why LOY in PBMCs and tumors are both associated with greater cancer mortality.

overall_design: Mouse single-cell RNA sequencing for hepatocellular carcinoma (HCC) studies was conducted using MUP-uPA+ mice. The mice were fed a Western Diet (Teklad, TD.88137) for 8 months, starting at 8 weeks of age. HCC development was assessed at 10 months of age.

first: Xingyu

last: Chen

first: Yiling

last: Shen

first: Suhyeon

last: Choi

first: Mukta

last: Basu

first: Lena

last: Hoelzen

first: Martina

last: Tufano

first: Hany

last: Abdel-Hafiz

first: Saravana

last: Kailasam Mani

first: Maryam

last: Ranjpour

first: Jiani

last: Zhu

first: Krishnan

last: Ramanujan

first: Ekaterina

last: Koltsova

first: Vinicius

last: Calsavara

first: Simon

last: Knott

first: Dan

last: Theodorescu


title: Unraveling cysteine deficiency-associated rapid weight loss [muscle]

first: Ilya

last: Shamovsky

first: Alan

last: Vaghese

first: Ivan

last: Gusarov


title: Concurrent loss of the Y chromosome in cancer and T cells impacts outcome [snRNA-seq]

first: XINGYU

last: CHEN

overall_design: Mouse single-nuclei RNA sequencing for hepatocellular carcinoma (HCC) studies was conducted using MUP-uPA+ mice. The mice were fed a Western Diet (Teklad, TD.88137) for 8 months, starting at 8 weeks of age. HCC development was assessed at 10 months of age.

first: Xingyu

last: Chen

first: Yiling

last: Shen


title: A subcellular map of translational machinery composition and regulation at the single-molecule level [ALIBi_ER_vs_cyto]

first: Maria

last: Barna

first: Adele

last: Xu


title: Enhancer adoption by an LTR retrotransposon generates viral-like particles causing developmental limb phenotypes

first: Juliane

last: Glaser


title: Immune and tumor cell landscape in pediatric high grade gliomas

city: Boston

first: Carlos Alberto

middle: Oliveira de

last: Biagi Junior

address: 360 Longwood Ave

department: Pediatric Oncology

institute: Dana Farber Cancer Institute

summary: Single cell RNA-seq profiling of tumor, myeloid, and T cells from fresh pediatric high grade glioma samples

overall_design:
We used Xenium Analyzer Spatial Transcriptomics to analyze 4 sections from 4 samples.

UPDATE: [July-24-2025] The raw and processed data files were renamed. The content of the files did not change.

first: Jenna

last: LaBelle

first: Rebecca

last: Poetschke

first: Alexander

last: Beck

first: Jacob

last: Haase

first: Bernhard

last: Englinger

first: Ilon

last: Liu

last: Trissal

first: Li

last: Jiang

first: Andrezza

last: Nascimento

first: Carlos

last: Alberto Oliveira de Biagi-Junior

first: Nathan

last: Mathewson

first: Orr

last: Ashenberg

first: Olivia

last: Hack

first: McKenzie

last: Shaw

first: Sina

last: Neyazi

first: Cuong

last: Nguyen

first: Gustavo

last: Alencastro Veiga Cruzeiro

first: Tom

last: Rosenberg

first: Jane

last: Vogelzang

first: Jason

last: Pyrdol

first: Sascha

last: Marx

first: Adrienne

last: Luomo

first: Anze

last: Godicelj

first: Alicia

last: Baumgartner

first: Sophia

last: Castellani

first: Daeun

last: Jeong

last: Rozowsky

first: Sibylle

last: Madlener

first: Lisa

last: Mayr

first: Andreas

last: Peyrl

first: Rene

last: Geyeregger

first: Daniela

last: Loetsch

first: Christian

last: Dorfer

first: Christine

last: Haberler

first: Natalia

last: Stepien

first: Irene

last: Slavc

last: Davidson

first: Robert

last: Prins

first: Kee

last: Yeo

first: Tabitha

last: Cooney

first: Keith

last: Ligon

first: Hart

last: Lidov

first: Sanda

last: Alexandrescu

first: Lissa

last: Baird


title: Inference of secreted-protein signaling activities in intercellular communication

status: Public on Mar 04 2025

city: Bethesda

first: Peng

last: Jiang

state: Maryland

address: 41 Center Drive

department: Center for Cancer Research

institute: National Cancer Institute

summary: The SecAct (Secreted Activity) model infers secreted-protein signaling activity from spatial, single-cell, and bulk transcriptomics data. Using SecAct, LY86 was identified and validated in vivo as a promising immunotherapy target.

overall_design: B16F10-Ly86 and B16F10-Vector tumors from mice that received 4 rounds of anti-PD-1 + anti-CTLA4 treatment were harvested and immediately snap-frozen in liquid nitrogen.

first: Beibei

last: Ru

first: Lanqi

last: Gong


title: Development and comprehensive evaluation of synthetic scarless circular RNAs

status: Public on Mar 01 2025

city: City

first: Lianhao

last: Song

state: Guangzhou

address: Jinsui road

department: Zhongshan Ophthalmic Center

institute: Sun Yat-sen University

summary: Circular RNAs (circRNAs) are a unique class of single-stranded RNA molecules with a closed-loop structure that confer enhanced stability, extended protein expression, and resistance to exonucleases, making them promising candidates for RNA therapeutics. In recent years, several methods have been developed to generate circRNAs, including the traditional, scar-containing Anabaena Permuted Intron-Exon (Ana-PIE) method and newer “scarless” circularization approaches. This study introduces a novel scarless circularization method, Split-Coxsackievirus B3-Anabaena Permuted Intron-Exon (Split-CVB3 Ana-PIE, SCAP). Scarless circular RNAs generated using the SCAP system were systematically compared to their scarred counterparts produced by the Ana-PIE system in terms of circularization efficiency, protein expression, stability, and immune response. The SCAP system achieved circularization efficiencies comparable to those of the Ana-PIE system while significantly enhancing protein expression. Scarless circular RNAs exhibited similar stability to scarred circular RNAs and did not trigger significant immune responses. These findings highlight the potential of scarless circular RNAs in gene therapy and vaccine development, demonstrating that removing extraneous sequences improves translation efficiency without compromising stability or immunogenicity. This study provides a foundation for the rational design of circular RNAs, with future efforts focusing on diverse target genes, optimized delivery platforms, and in vivo validation.

overall_design: RNA-seq profiling of HEK293T cells transfected with linear RNA and circRNA at 24 hours

first: Linfeng

last: Chen

first: Zhi

last: Xie


title: Separate brainstem circuits for fast steering and slow exploratory turns

city: Shanghai

first: Lulu

last: Xu

address: No.500 Zhennan Raod

institute: TongJi university

summary: During locomotion, trajectory changes necessitate precise tuning of descending commands to scale turning movements according to specific tasks or objectives, resulting in either rapid steering turns during prey pursuit or routine shallow turns associated with exploration. We show that these two types of turning are controlled by separate brainstem circuits that encode rapid steering turning versus slow exploratory turns. The circuit for rapid steering is widely distributed across different brainstem nuclei, involving specific excitatory V2a and inhibitory commissural V0d neurons. The steering V2a and V0d neurons are furthermore coupled via gap junctions and simultaneously recruited to ensure rapid steering through an asymmetrical recruitment of spinal motor neurons. The recruitment of these steering neurons is primarily associated with the degree of the direction change, rather than the locomotor frequency. The brainstem steering neurons are, in turn, controlled by a subset of V2a neurons in the pretectum activated by salient visual input. Conversely, the circuit controlling swim-related slow exploratory turns comprises a different set of V2a neurons localized in fewer brainstem nuclei. These findings demonstrate a modular organization of the brainstem circuits that control rapid steering and slow exploratory turning during locomotion.

overall_design: CaD neurons were taken fromTg ( gly2: GFP) zebrafish brain stem, one cell from each fish, a total of eight fish

first: Bing

last: Zhu

first: Zhiqiang

first: Xingyu

last: Tao

first: Tianrui

last: Zhan

first: Abdel

last: El Manira

first: Jianren


title: Metformin attenuates skeletal muscle transcriptional and mitochondrial respiratory adaptations to aerobic exercise training.

status: Public on Mar 03 2025

city: Madison

first: Matthew

middle: D

last: Bruss

state: WI

address: 201 West Lakeside Street

institute: UW-Madison

summary: We have previously demonstrated that the inhibitory effect of metformin on skeletal muscle complex-I linked mitochondrial respiration was associated with attenuated improvements in whole-body insulin sensitivity and cardiorespiratory fitness after aerobic exercise training (AET) in older adults. However, the mechanism(s) by which metformin impacts skeletal muscle adaptations to aerobic exercise remains unclear. To understand potential mechanisms associated with the inhibitory effect of metformin on mitochondrial adaptations to AET, we evaluated the skeletal muscle transcriptome, mitochondrial respiration, and hydrogen peroxide (H2O2) emissions in 7-month-old male C57BL6/J mice after 8-weeks of non-exercise sedentary control (SED) or progressive AET with and without metformin treatment. Metformin attenuated the number of differentially expressed genes by AET (975 vs. 449). Pathway analysis suggests that metformin suppressed several signal transduction pathways involved in oxidative capacity and cellular metabolism, including VEGF, AMPK, HIF-1α, and PI3K-AKT. Notably, the addition of metformin to AET suppressed 16 downstream targets of HIF-1α and was accompanied by metformin ameliorating the increase in mitochondrial respiratory capacity and ADP sensitivity after AET. However, neither AET nor AET+MET impacted mitochondrial H2O2 emitting potential. Given the central role of HIF-1α as an upstream transcriptional regulator of mitochondrial function and biogenesis, these results suggest that suppression of HIF-1α pathway by metformin could serve as a transcriptional regulatory node associated with the inhibitory role of metformin on skeletal muscle mitochondrial adaptations to aerobic exercise training.

overall_design: Mice were randomized to treatment groups (n=8-9/group) to sedentary (SED), aerobic exercise training (AET), aerobic exercise training and metformin (AET+MET). Mice allocated to AET (or AET + MET) were exercised on a 6-lane motorized treadmill (Columbus Instruments Exer 3/6 treadmill) 5 days/week, 60min/day for 8 weeks. Metformin (Cayman chemical 13118) was provided in the drinking water at 2mg/mL and replaced weekly.  Left gastrocnemius was collected and snap frozen.

first: Christian

last: Elliehausen

first: Dennis

last: Minton

first: Audrey

middle: L

last: Spiegelhoff

first: Adam

last: Konopka