title: Type I Interferon promotes MxA-dependent IL-1b release in SLE monocytes

first: Simone

last: Caielli

state: NY

address: 413 E 69th Street

department: Department of Pediatrics

institute: Weill Cornell Medicine

summary: Systemic lupus erythematosus (SLE) is characterized by upregulation of Type Ι Interferon (IFN) and widespread inflammation. However, blocking the IFN pathway benefits a fraction of patients, pointing to additional pathogenic players. Here we describe monocytes (Mo) undergoing erythrophagocytosis and co-expressing IFN-inducible genes (ISGs) and interleukin-1b (IL-1b) in patients with active disease. This phenotype is recapitulated in vitro upon internalization of red blood cells carrying mitochondria (Mito+ RBCs), a feature of SLE. While ISG expression requires the interaction between Mito+ RBC-derived mitochondrial DNA (mtDNA) and cGAS, the production of IL-1b entails Mito+ RBC-derived mitochondrial RNA (mtRNA) triggering RIG-I-like receptor (RLR) activation. This leads to the cytosolic release of Mo-derived mtDNA and activation of the NLRP3 inflammasome. Importantly, the Type I IFN-inducible protein myxovirus resistant protein 1 (MxA) enables IL-1b release by routing this cytokine into a trans-Golgi network (TGN)-mediated unconventional secretory pathway. As Type I IFN and IL-1b are thought to counter-regulate each other, our study highlights an unprecedented synergy between these two cytokine pathways in SLE.

overall_design: Human Monocytes (Mo) or BLaER1 Mo were cocultured with CFSE labeled red blood cells carrying (Mito+) or not carrying (Mito-) RBCs in complete RPMI medium. 18 hr later Mo or BLaER1 Mo internalizing RBCs were FACS sorted and processesed for RNAseq analysis

first: Preetha

last: Balasubramanian

first: Uthra

last: Balaji

first: Zurong

last: Wan

first: Jeanine

last: Baisch

first: Lynnette

last: Walters

first: Cynthia

last: Smitherman

first: Djamel

last: Nehar-Belaid

first: Radu

last: Marches

first: Lorien

last: Nassi

first: Katie

last: Stewart

first: Julie

last: Fuller

first: Jacques

last: Banchereau

first: Jinghua

last: Gu

first: Tracey

last: Wright

first: Virginia

last: Pascual


title: Viral triggers of Type 1 diabetes: Effects of Coxsackie B virus infection on transcriptome of trophoblast and pancreatic beta cell lines.

status: Public on Oct 06 2024

city: Albany

first: Salima

last: Lalani

state: New York

address: 47 New Scotland Avenue

department: Department of Molecular & Cellular Physiology

country: USA

institute: Albany Medical College

type: Expression profiling by high throughput sequencing

type: Non-coding RNA profiling by high throughput sequencing

summary: The epidemiological association of coxsackievirus B infection with type 1 diabetes suggests that therapeutic strategies that reduce viral load could delay or prevent disease onset.  Moreover, recent studies suggest that treatment with antiviral agents against coxsackievirus B may help preserve insulin levels in type 1 diabetic patients. In the current study, we performed small RNA-sequencing to show that infection of immortalized trophoblast cells with coxsackievirus caused differential regulation of several miRNAs. One of these, hsa-miR-AMC1, was similarly upregulated in human pancreatic β cells infected with coxsackievirus B4.  Moreover, treatment of β cells with non-cytotoxic concentrations of an antagomir that targets hsa-miR-AMC1 led to decreased CVB4 infection, suggesting a positive feedback loop wherein this microRNA further promotes viral infection. Interestingly, some predicted target genes of hsa-miR-AMC1 are shared with hsa-miR-184, a microRNA that is known to suppress genes that regulate insulin production in pancreatic β cells.  Consistently, treatment of coxsackievirus B4-infected β cells with the hsa-miR-AMC1 antagomir was associated with a trend toward increased insulin production. Taken together, our findings implicate novel hsa-miR-AMC1 as a potential early biomarker of coxsackievirus B4-induced type 1 diabetes and suggest that inhibiting hsa-miR-AMC1 may provide therapeutic benefit to type 1 diabetes patients. Our findings also support the use of trophoblast cells as a model for identifying microRNAs that might be useful diagnostic markers or therapeutic targets for coxsackievirus B-induced type 1 diabetes.

overall_design:
Samples 1-18: EndoC-βH1 cells were infected with CVB4-JVB and CVB4-E2 strains for 1 hour at a multiplicity of infection (MOI) of 0.1. Cells were pelleted for RNA extraction, library preparation, and RNA-Seq.
Samples 19-30: Sw.71 immortalized trophoblast cells were infected with CVB4-JVB strain for 24 hours at an MOI of 1. Cells were pelleted for RNA extraction, library preparation, RNA-seq, and small RNA-Seq.

first: Joseph

last: Knudsen

first: James

last: Kenney

first: Didier

last: Hober

first: C. Michael

last: DiPersio

first: Allen

last: Gerber


title: Bacterial purine signaling pathway modulates C. elegans development and stress tolerance via DAF-16

city: college station

first: Min

last: Feng

state: Texas

address: 200 Spence street

department: Chemical Engineering

institute: Texas A&M University

summary: The purine signaling pathway is crucial for cellular function and is a conserved metabolic network from prokaryotes to humans. While extensively studied in microorganisms like yeast and bacteria, the impact of perturbing dietary intermediates from the purine signaling pathway on animal development and growth remains poorly understood. We utilized Caenorhabditis elegans as the metazoan model to investigate the mechanisms underlying this deficiency. Through a high-throughput screening of an E. coli mutant library Keio collection, we identified 34 E. coli mutants that delay C. elegans development. Among these mutants, we found that E. coli purE gene is an essential genetic component that promotes host development in a dose-dependent manner. Further metabolites supplementation suggests that bacterial purE downstream metabolite 5-aminoimidazole-4-carboxamide ribotide (AICAR) can inhibit worm growth. Additionally, we found the FoxO transcription factor DAF-16 is indispensable in worm development delay induced by purE mutation, and observed increased nuclear accumulation of DAF-16 when fed E. coli purE- mutants, suggesting the role of DAF-16 in response to purE mutation. RNA-seq analysis and phenotypic assays revealed that worms fed the E. coli purE mutant exhibited elevated lifespan, thermotolerance, and pathogen resistance. These findings collectively suggest that certain intermediates in the bacterial purine signaling pathway can serve as a cue to modulate development and activate the defense response in the nematode C. elegans through DAF-16.

overall_design: Synchronized L1 stage C. elegans worms were grown on NGM agar plates seeded with E. coli BW25113 bacteria. After reaching the desired developmental stage, worms were transferred to NGM agar plates with either wild-type E. coli BW25113 or the E. coli purE mutant. Following a 48-hour feeding period, worms underwent a 2-hour diet switch.

first: Baizhen

last: Gao

first: Luis

middle: R

last: Garcia

first: Qing

last: Sun


title: G3BP2B stress granules regulate mRNA expression under ER stress [1]

status: Public on Oct 07 2024

city: San Francisco

first: José

middle: M.

last: Liboy-Lugo

state: CA

address: 513 Parnassus Ave.

institute: UCSF

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

overall_design: Refer to individual Series


title: TF ChIP-seq Caenorhabditis elegans II

city: Seattle

first: Louis

last: Gevirtzman

state: WA

address: 3720 15th Ave NE

department: Genome Science

institute: University of Washington

type: Genome binding/occupancy profiling by high throughput sequencing

summary: A catalog of transcription factor (TF) binding sites in the genome is critical for deciphering regulatory relationships.  Here we present the culmination of the efforts of the modENCODE (Model Organism ENCyclopedia Of DNA Elements) and modERN (model organism Encyclopedia of Regulatory Networks) consortia to systematically assay TF binding events in vivo in two major model organisms, Drosophila melanogaster (fly) and Caenorhabditis elegans (worm).  These datasets comprise 605 TFs identifying 3.6M sites in the fly and 356 TFs identifying 0.9 M sites in the worm, and represent the majority of the regulatory space in each genome. We demonstrate that TFs associate with chromatin in clusters termed “metapeaks”, that larger metapeaks have characteristics of high occupancy target (HOT) regions, and that the importance of consensus sequence motifs bound by TFs depends on metapeak size and complexity.  Combining ChIP-seq data with single cell RNA-seq data in a machine learning model identifies particular TFs with a prominent role in promoting target gene expression in specific cell types, even differentiating between parent-daughter cells during embryogenesis.  These data are a rich resource for the community that should fuel and guide future investigations into TF function.  To facilitate data accessibility and utility, all strains expressing GFP-tagged TFs are available at the stock centers for each organism.  The chromatin immunoprecipitation sequencing data are available through the ENCODE Data Coordinating Center, GEO, and through a direct interface (http://epic.gs.washington.edu/modERN/) that provides rapid access to processed data sets and summary analyses, as well as widgets to probe the cell type-specific TF-target relationships.

overall_design: Peak calling of sequence files using ENCODE TF/Histone ChIP-Seq pipeline version 1.3.6 https://github.com/ENCODE-DCC/chip-seq-pipeline2

first: Michelle

last: Kudron

first: Alec

last: Victorsen

first: Bridget

middle: C

last: Lear

first: Jiahao

first: Jinrui

last: Xu

first: Swapna

last: Samanta

first: Emily

last: Frink

first: Adri

last: Tran-Pearson

first: Chau

last: Huynh

first: Dionne

last: Vafeados

first: Ann

last: Hammonds

first: William

last: Fisher

first: Martha

last: Wall

first: Greg

last: Wesseling

first: Vanessa

last: Hernandez

first: Zhichun

last: Lin

first: Mary

last: Kasparian

first: Kevin

last: White

first: Ravi

last: Allada

first: Mark

last: Gerstein

first: LaDeana

last: Hillier

first: Susan

middle: E

last: Celniker

first: Valerie

last: Reinke

first: Robert

middle: H

last: Waterston


title: TF ChIP-seq Drosophila melanogaster II

first: Louis

last: Gevirtzman

first: Michelle

last: Kudron

first: Alec

last: Victorsen

first: Bridget

last: Lear

first: Jiahao

last: Gao

first: Jinrui

last: Xu

first: Swapna

last: Samanta

first: Emily

last: Frink

first: Adri

last: Tran-Pearson


title: Gene regulatory network inference from CRISPR perturbations in primary CD4+ T cells elucidates the genomic basis of immune disease

city: Baltimore

first: Josh

last: Weinstock

state: MD

address: 3100 Wyman Park Dr

institute: Johns Hopkins University

summary: The effects of genetic variation on complex traits act mainly through changes in gene regulation. Although many genetic variants have been linked to target genes in cis, the trans-regulatory cascade mediating their effects remains largely uncharacterized. Mapping trans-regulators based on natural genetic variation, including eQTL mapping, has been challenging due to small effects. Experimental perturbation approaches offer a complementary and powerful approach to mapping trans-regulators. We used CRISPR knockouts of 84 genes in primary CD4+ T cells to perturb an immune cell gene network, targeting both inborn error of immunity (IEI) disease transcription factors (TFs) and background TFs matched in constraint and expression level, but without a known immune disease association. We developed a novel Bayesian structure learning method called Linear Latent Causal Bayes (LLCB) to estimate the gene regulatory network from perturbation data and observed 211 directed edges among the genes that could not be detected in existing CD4+ trans-eQTL data. We used LLCB to characterize the differences between the IEI and background TFs, finding that the gene groups were highly interconnected, but that IEI TFs were much more likely to regulate immune cell specific pathways and immune genome-wide association study (GWAS) genes. We further characterized nine coherent gene programs based on downstream effects of the TFs and linked these modules to regulation of GWAS genes, finding that canonical JAK-STAT family members are regulated by KMT2A, a global epigenetic regulator. These analyses reveal the trans-regulatory cascade from upstream epigenetic regulator to intermediate TFs to downstream effector cytokines and elucidate the logic linking immune GWAS genes to key signaling pathways.

overall_design: CRISPR KO of inborn error of immunity transcription factors and non-immune disease related transcription factors in primary human CD4+ T cells

first: Joshua

middle: S

first: Maya

middle: M

last: Arce

first: Jacob

middle: W

last: Freimer

first: Mineto

last: Ota

first: Alexander

last: Marson

first: Alexis

last: Battle

first: Jonathan

middle: K

last: Pritchard


title: Binding profiles for 602 Drosophila and 350 C. elegans transcription factors reveal tissue specific regulatory relationships

first: Louis

sample_organism:
Caenorhabditis elegans
Drosophila melanogaster


title: The early transcriptional and post-transcriptional responses to fluconazole in sensitive and resistant Candida albicans (RNA-Seq)

city: Solna

first: Irene

last: Stevens

address: Tomtebodavägen 23A (Gamma5)

department: Department of Microbiology, Tumor and Cell Biology

country: Sweden

institute: Karolinska Institutet

summary: Candida albicans is a leading cause of fungal infections in immunocompromised patients. Management of candidemia relies on a few antifungal agents, with fluconazole being first line therapy. The emergence of fluconazole-resistant strains highlights the pressing need to improve our molecular understanding of the drug response mechanisms. By sequencing the 5’P mRNA degradation intermediates, we show that co-translational mRNA decay is common in C. albicans and characterize how in vivo 5´-3´ exonuclease degradation trails the last translating ribosome. Thus, the study of the 5'P mRNA degradome (5PSeq) offers a simple and affordable way to measure ribosome dynamics and identify codon specific ribosome stalls in response to drugs and amino acid deprivation. Building upon this, we combine RNA-Seq and 5PSeq to study the early response of sensitive and resistant C. albicans isolates to fluconazole. Our results highlight that transcriptional responses, rather than changes in ribosome dynamics, are the main driver of Candida resistance to fluconazole.

overall_design: Gene expression profiling by RNA-seq data for Candida albicans SC5314 (sensitive) versus PLC124 (clinical isolate, fluconazole resistant) treated with treated with Fluconazole (1 μg/ml, 1xMIC) or control (2% DMSO) for 30 minutes.

first: Fitz

middle: G

last: Silao

first: Susanne

last: Huch

first: Honglian

last: Liu

first: Kicki

last: Ryman

first: Adriana

last: Carvajal-Jimenez

first: Per

middle: O

last: Ljungdahl

first: Vicent

last: Pelechano


title: Ubiquitin-activating enzyme mediates immune escape via suppressing interferon signaling

city: Ann Arbor

first: Arul

last: Chinnaiyan

state: MICHIGAN

address: 1500 E. Medical Center Dr

institute: University of Michigan

summary: How cancer cells escape immune surveillance and resist immunotherapies remain to be elucidated. By screening candidate genes frequently amplified in cancer, we identified expression of ubiquitin-activating enzyme (UBA1) as being the most negatively correlated with signatures related to effector CD8+T cells. High expression of UBA1was strongly predictive of resistance to immune checkpoint blockade (ICB) and poor survival in ICB cohorts. In immunocompetent syngeneic models, but not in immunodeficient models, we found that overexpression ofUba1in cancer cells promoted tumor growth and reduced functional, intratumoral CD8+T cells. Conversely, depletion ofUba1impaired tumor progression, accompanied with increase of functional, intratumoral CD8+T cells. CD8+T cell depletion rescued tumor growth impairment mediated byUba1depletion. Additionally, a selective, small molecule inhibitor of UBA1, TAK-243, delayed tumor growth and augmented ICB in various syngeneic models, in a manner dependent on CD8+T cells. Intratumoral administration of TAK-243 and tumor rechallenge experiments revealed that TAK-243 treatment exhibited a remarkable abscopal effect and provided prolonged protection against cancer, accompanied with elevation of memory CD8+ T cells. Mechanistically, depletion or inactivation of UBA1 stabilized the short-lived interferon (IFN) pathway component, JAK1, upregulated both type-I and -II IFN signaling pathways, and resulted in elevation of key immune modulators, including CXCL9, CXCL10, and MHC-I. Taken together, our data supports that UBA1 mediates cancer immune escape via suppressing interferon signaling and suggests that targeting UBA1 may activate systemic cancer immunity.

overall_design: Tumors with Uba1 overexpression or depletion, or from mice treated by TAK-243 were dissociated for single cell RNA sequencing.

first: Yi

last: Bao

first: Gabriel

last: Cruz

first: Yuping

last: Zhang

first: Miriam

last: Shahine

first: Yuanyuan

last: Qiao

first: Rahul

last: Mannan

first: Jing

last: Hu

first: Mahnoor

last: Gondal

first: Fan

last: Yang

first: Sarah

last: Kang

first: Alec

last: Chu

first: Jae

last: Choi

first: Jiali

last: Yu

first: Heng

last: Lin

first: Stephanie

last: Miner

first: Dan

last: Robinson

first: Yi-Mi

last: Wu

first: Yang

last: Zheng

first: Xuhong

last: Cao

first: Fengyun

last: Su

first: Rui

last: Wang

first: Noshad

last: Hosseini

first: Marcin

last: Cieslik

first: Weiping

last: Zou


title: Decoding the immune checkpoint signatures in human atherosclerosis: implications of type 2 diabetes and lipid-lowering intervention [mouse_plaque_scRNAseq]

city: New York

first: Michael

last: Gildea

address: 550 1st avenue

institute: NYU Langone Health

summary: Immune checkpoint inhibitor (ICI) therapies in cancer accelerate atherosclerosis progression. Here, we have charted the landscape of immune checkpoint gene expression and defined the network of disease-relevant interactions with single-cell resolution. We found that signaling through PD-1 and CTLA4 is driven by a population of dendritic cells enriched for FSCN1 that can be derived from peripheral blood cells following anti-PD-1 or -CTLA4 treatment ex vivo. Type 2 diabetes dampened plaque PD-1 and CTLA4 signaling, showing that cardiometabolic comorbidities elicit unique responses to ICIs. Lipid-lowering therapy equalized the intensity and direction of immune checkpoint interactions in human blood, while atherosclerotic mice subjected to a lipid-lowering diet displayed both increased co-inhibitory signaling and a downregulation of inflammatory transcriptional programs in plaques. Our findings underscore the potential of lipid-lowering therapies in stabilizing immune checkpoint interactions and reducing plaque inflammation, offering new insights on atherosclerosis and cardiovascular risks in cancer patients undergoing ICI treatments.

overall_design: scRNAseq profiling of CD45+ cells from mouse aortic digestion before and after lipid lowering.

middle: A

first: Jose

last: Duran

first: Dayasagar

last: Das

first: Kathryn

middle: J

last: Moore

first: Chiara

last: Giannarelli


title: Cystatin F attentuates neuroinflammation and demyelination following murine coronavirus infection of the central nervous system

city: Salt Lake City

first: Amber

last: Syage

state: Utah

address: 2000 Circle of Hope

department: Pathology

institute: University of Utah

summary: Background: Cystatin F is a secreted lysosomal cysteine protease inhibitor that has been implicated in affecting the severity of demyelination and enhancing remyelination in pre-clinical models of immune-mediated demyelination. How cystatin F impacts neurologic disease severity following viral infection of the central nervous system (CNS) has not been well characterized and was the focus of this study. We used cystatin F null-mutant mice (Cst7-/-) with a well-established model of murine coronavirus-induced neurologic disease to evaluate the contributions of cystatin F in host defense, demyelination and remyelination. Methods: Wildtype controls and Cst7-/- mice were intracranially (i.c.) infected with a sublethal dose of the neurotropic JHM strain of mouse hepatitis virus (JHMV), with disease progression and survival monitored daily. Immune cell infiltration into the brain and spinal cord was determined by flow cytometry. Spinal cord demyelination was determined by luxol fast blue (LFB) and Hematoxylin/Eosin (H&E) staining and remyelination evaluated by electron microscopy (EM) and calculation of g-ratios. Immune cell infiltration into the CNS and microglia activation were determined by flow cytometry and 10X genomics chromium 3’ single cell RNA sequencing (scRNAseq). Results: JHMV-infected Cst7-/- mice were able to control viral replication within the CNS, indicating that cystatin F is not essential for an effective Th1 anti-viral immune response. Infiltration of T cells and monocytes/macrophages into the spinal cords of JHMV-infected Cst7-/- mice was increased compared to infected controls, and this correlated with amplified demyelination and impaired remyelination. Single-cell RNA-seq of CD45+ cells enriched from spinal cords of infected Cst7-/- and control mice at day 21 post-infection (p.i.) revealed enhanced expression of transcripts encoding macrophage chemoattractant, Ccl2, and T cell chemoattractants, Cxcl9 and Cxcl10, combined with elevated expression of interferon-g (Ifng) transcripts and activation markers in CD8+ T cells from Cst7-/- mice compared to controls. Conclusions: Cystatin F is not required for immune-mediated control of JHMV replication within the CNS. However, JHMV-infected Cst7-/- mice exhibited more severe clinical disease associated with increased demyelination and impaired remyelination. The increase in disease severity was associated with elevated expression of macrophage and T cell chemoattractant chemokines, concurrent with increased neuroinflammation. These findings support the theory that one mechanism by which cystatin F affects chronic disease in mice persistently infected with JHMV is through regulating expression of pro-inflammatory molecules that impact neuroinflammation and neurologic disease.

overall_design: Cst7 knock-out (KO) or wildtype (WT) littermate controls were injected intracranially (i.c.) in the right brain hemisphere with 1200 plaque forming units (PFU) of JHMV and CD45-positive (CD45+) cells were flow sorted from the spinal cords of uninfected and JHMV-infected Cst7 KO and WT mice at day 21 post-infection (p.i.)  FACS sorted CD45+ cells were CellPlex labeled and pooled, then underwent 3' single-cell RNA-sequencing analysis

first: Caden

last: Henningfield

first: Collin

last: Pachow

first: Annie

last: Du

first: Yuting

last: Cheng

first: Gema

last: Olivarria

first: Shimako

last: Kawauchi

first: Grant

last: MacGregor

first: Kim

middle: N

last: Green

first: Thomas

last: Lane


title: Perivascular fibrosis is mediated by a KLF10-IL-9 signaling axis in CD4+ T cells

city: Boston

first: Rulin

last: Zhuang

state: MA

address: 77 Avenue Louis Pasteur NRB-742F

department: Department of Medicine, Cardiovascular Division

institute: Brigham and Women’s Hospital/Harvard Medical School

summary:
Perivascular fibrosis, characterized by increased amount of connective tissue around vessels, is a hallmark for vascular disease. Angiotensin II (Ang II) contributes to vascular disease and end-organ damage via promoting T-cell activation. Despite recent data suggesting the role of T cells in the progression of perivascular fibrosis, the underlying mechanisms are poorly understood. Kruppel-like Factor 10 is a transcription factor expressed in T cell subsets. We sought to investigate the role of KLF10 in CD4+ T cells in regulating vascular damage in an AngII mouse model. CD4-targeted KLF10 deficient (TKO) and CD4-Cre (WT) mice were generated and subjected to 28 days of Ang II infusion. Differential gene expression profiling of non-stripped aorta, stripped aorta, isolated aortic fibroblasts from CD4-Cre (Cre) or CD4-Cre KLF10-fl/fl mice(TKO) mice after 28 days Angiotensin II (Ang II) treatment were performed to ascertain the underlying global transcriptomic changes involved in perivascular fibrosis. Since increased IL-9 is a profibrotic cytokine, we hypothesized that perivascular fibrosis might be rescued by administration of anti-IL-9 neutralizing monoclonal antibodies (mAbs) in Ang II-treated TKO mice. Then differential gene expression profiling of non-stripped aorta from TKO mice with or without anti-IL9-antibody treatment after 28 days Ang II infusion. To further understand the differences in phenotypic heterogeneity in fibroblasts during the progression of perivascular fibrosis between TKO and control mice in response to Ang II treatment, we performed single-cell RNA-sequencing (scRNA-seq) of the descending aorta. Collectively, comparative RNA-seq transcript profiling identified several pathways associated with periadventitial remodeling and that the calcium signaling pathway is dominantly upregulated in non-stripped aortas of TKO mice after Ang II treatment. Indeed, the profibrotic genes Col8a1, Mmp2, Fmod, and Angptl1, which were identified as significantly increased in TKO aortas and in isolated fibroblasts by both bulk RNA seq and scRNA seq, were all decreased after treatment with anti-IL-9 mAbs. These results indicate that the KLF10-IL-9 signaling axis in CD4+ T cells tightly regulate the processes of Ang II-induced pathological perivascular fibrosis and provide novel therapeutic opportunities for the treatment of hypertensive-associated diseases.
1) Differential gene expression profiling of non-stripped aotra, stripped aorta, aortic fibroblasts from CD4-Cre(Cre) or CD4-Cre KLF10-fl/fl mice(TKO) mice after 28 days Angiotensin II (Ang II) treatment. 2) Differential gene expression profiling of non-stripped aotra from TKO mice with or without anti-IL9-antibody treatment after 28 days Ang II infusion. 3) Single cell RNA-sequencing was perform on descending aorta from  Cre or TKO after Ang II treatment.

overall_design: 1) Stripped and non-stripped aorta from Cre or TKO after Ang II treatment were collected for RNA-sequencing and gene ontology pathway analysis.  2) Aortic fibroblasts isolated from Cre or TKO after Ang II treatment were subjected to RNA-sequencing and gene ontology pathway analysis. 3) Non-stripped aorta from TKO mice after Ang II treatment with/without anti-IL-9 antibodies were collected for RNA-sequencing and gene ontology pathway analysis 4) Non-stripped aorta from Cre or TKO after Ang II treatment were collected, digested, and subjected for Single cell RNA-sequencing.

first: Jingshu

last: Chen

first: Henry

first: Mark

last: Feinberg


title: Engineered extrachromosomal oncogene amplifications promote tumorigenesis  [shallow whole genome sequencing and ATACseq]

status: Public on Oct 05 2024

first: Andrea

last: Ventura

department: Cancer Biology and Genetics

institute: Memorial Sloan Kettering Cancer Center

summary: Focal gene amplifications are among the most common cancer-associated mutations, but their evolution and contribution to tumorigenesis have proven challenging to recapitulate in primary cells and model organisms. Here we describe a general strategy to engineer large (>1 Mbp) focal amplifications mediated by extrachromosomal circular DNAs (ecDNAs) in a spatiotemporally controlled manner in cells and in genetically engineered mice. By coupling ecDNA formation with expression of selectable markers, we track the dynamics of ecDNA-containing cells under physiological conditions and in the presence of specific selective pressures. We also apply this approach to generate mice harboring Cre-inducible Myc- and Mdm2-containing ecDNAs analogous to those occurring in human cancers. We show that the engineered ecDNAs spontaneously accumulate in primary cells derived from these animals, promoting their proliferation, immortalization, and transformation. Finally, we demonstrate the ability of Mdm2-containing ecDNAs to promote tumor formation in an autochthonous mouse model of hepatocellular carcinoma.

overall_design: Control neuronal stem cells and neuronal stem cells harboring engineered ecDNAs were analyzed by shallow whole genome sequencing and ATACseq.

first: Davide

last: Pradella

first: Howard

last: Chang

first: Paul

last: Mischel


title: Spatial Transcriptomics in Bone Mechanomics: Exploring the Mechanoregulation of Fracture Healing in the Era of Spatial Omics.

city: Zurich

first: Neashan

last: Mathavan

address: GLC H 21.1, Gloriastrasse 37 / 39

department: Institute for Biomechanics

country: Switzerland

institute: ETH Zurich

summary: In recent decades, the field of bone mechanobiology has sought experimental techniques to unravel the molecular mechanisms governing the phenomenon of mechanically-regulated fracture healing. Each cell within a fracture site resides within different local micro-environments characterized by different levels of mechanical strain - thus, preserving the spatial location of each cell is critical in relating cellular responses to mechanical stimuli. Our spatial transcriptomics based “mechanomics” platform facilitates spatially-resolved analysis of the molecular profiles of cells with respect to their local in vivo mechanical environment by integrating time-lapsed in vivo micro-computed tomography, spatial transcriptomics, and micro-finite element analysis. We investigate the transcriptomic responses of cells as a function of the local strain magnitude by identifying the differential expression of genes in regions of high and low strain within a fracture site. Our platform thus has the potential to address fundamental open questions within the field and to discover mechano-responsive targets to enhance fracture healing.

overall_design: Female 12-week-old mice (n = 4) received mid-diaphyseal femoral defects (0.68 ± 0.04 mm) using an established osteotomy surgical protocol. In vivo micro-CT imaging is performed weekly at the fracture site. Mice which exhibit bridging at 3 weeks post-surgery are then subdivided into Control and Loaded groups. Between weeks 3 – 5, mice receive either cyclic mechanical loading or sham-loading three times per week. All mice are euthanized at 5 weeks post-surgery. Spatial transcriptomics analyses are performed on the explanted femurs (n = 1 Control, n = 1 Loaded). Micro-finite element models - based upon in vivo micro-CT images - are used to generate tissue-scale 3D maps of the mechanical environment. We then correlated the molecular responses of individual cells to their local mechanical environment.

first: Amit

last: Singh

first: Francisco

last: Marques

first: Denise

last: Günther

first: Gisela

last: Kuhn

first: Esther

last: Wehrle

first: Ralph

last: Müller


title: Engineered extrachromosomal circular DNAs promote tumorigenesis (WGS)

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Control cells or cells harboring engineered ecDNAs were analyzed by shallow whole genome sequencing.

first: Minsi

last: Zhang

first: Rui

last: Gao


title: Engineered extrachromosomal oncogene amplifications promote tumorigenesis  (RNA-Seq)

overall_design: Control cells or cells harboring engineered ecDNAs were analyzed by RNAseq.


title: Engineered extrachromosomal oncogene amplifications promote tumorigenesis


title: Preclinical evaluation of aluminum hydroxide-tethered interleukin-12 (JEN-101) therapy in pet dogs with advanced malignant melanoma

status: Public on Oct 04 2024

city: Urbana

first: Timothy

last: Fan

state: Illinois

address: 1008 West Hazelwood Drive

department: Veterinary Clinical Medicine

institute: University of Illinois Urbana Champaign

summary: A Phase I study was conducted to determine the tolerability, activity, and immune responses of JEN-101 in dogs with advanced melanoma.  A 3+3 dose escalation design was used to evaluate intratumoral injection of JEN-101 at 1, 3, 10, or 20 μg/kg every three weeks for four cycles. A second course was allowable in the absence of disease progression or toxicity. Peripheral blood, serum, and tumor biopsies were collected at baseline and at pre-specified timepoints for pharmacokinetic and immune analyses, which included serum cytokines, immunohistochemistry, and gene expression assessment.  JEN-101 was well tolerated with adverse events being fever, lethargy, and isolated elevated liver enzymes. Five dogs experienced grade 3 events and no grade 4 events were observed. Pharmacokinetic analysis showed a trend towards dose-related Cmax within 8 hours of injection. Responding dogs demonstrated increased systemic interferon-γ and IL-10 AUC levels and local recruitment of CD3+ T cells. Increased pro-inflammatory and antigen processing gene expressions were identified in responding lesions.

overall_design: Pet dogs with melanoma  were enrolled into four cohorts of 1, 3,10, and 20 µg/kg administered by local injection of aluminum hydroxide-tethered interleukin-12 (JEN-101) every three weeks for four cycles. For dogs without disease progression or clinically significant toxicity, another four cycles of JEN-101 every three weeks were allowable. Dogs were followed for up to one year following completion of treatment. Baseline tumor biopsy was required in all dogs and post-treatment biopsies were collected when clinically feasibility or at the time of death.

first: Matheus

last: Passos Barbosa


title: Ubiquitin-activating enzyme mediates immune escape via suppressing interferon signaling [bulk RNA-seq]

first: Arul

last: Chinnaiyan

overall_design: We depleted Uba1 or inhibited UBA1 by TAK-243 in the murine Myc-CaP cells and performed RNA-sequencing. We also harvested tumors with Uba1 overexpression, depletion, or inactivation for RNA-sequencing.

first: Yi

last: Bao

first: Gabriel

last: Cruz

first: Yuping

first: Miriam

last: Shahine

first: Yuanyuan

last: Qiao

first: Rahul

last: Mannan

first: Jing

last: Hu

first: Mahnoor

last: Gondal

first: Fan

last: Yang

first: Sarah

last: Kang

first: Alec

last: Chu

first: Jae

last: Choi

first: Jiali

last: Yu

first: Heng

last: Lin

first: Stephanie

last: Miner

first: Dan

last: Robinson

first: Yi-Mi

last: Wu

first: Yang

last: Zheng

first: Xuhong

last: Cao

first: Fengyun

last: Su

last: Wang

first: Noshad

last: Hosseini

first: Marcin

last: Cieslik

first: Weiping

last: Zou


title: Aspartate signaling increases the aggressiveness of lung metastases by inducing eIF5A-mediated translation (Total RNA-Seq)

city: Leuven

first: Juan

last: Fernández-García

address: Campus Gasthuisberg, O&N4, Herestraat 49, box 912

department: VIB-KU Leuven Center for Cancer Biology

country: Belgium

institute: VIB/KU Leuven

summary: Lung metastases are detected in more than half of patients with metastatic tumors. However, it remains largely unknown why the lung environment is a permissive niche for metastases. Here, we discover that pulmonary aspartate triggers a cellular signaling cascade in disseminated cancer cells resulting in a translational program that boosts lung metastasis. Specifically, we observe that patients and mice with breast cancer have high concentrations of aspartate in their lung interstitial fluid. This extracellular aspartate activates the ionotropic N-methyl-D-aspartate (NMDA) receptor in cancer cells, which induces CREB-dependent mRNA expression of deoxyhypusine hydroxylase (DOHH). The latter is essential for hypusination, a posttranslational modification required for the activity of the non-classical translation initiation factor eIF5A. In turn, a translational program with TGF-β signaling as a central hub promotes collagen remodeling in the disseminated breast cancer cells. We detect key aspects of this mechanism in lung metastases from patients with breast cancer. In summary, we discover that pulmonary aspartate increases with breast cancer and induces a signaling cascade promoting the growth of lung metastases.

overall_design: RNA-seq of total mRNA fractions extracted from control (shSCR), GRIN2D-knockdown (shGRIN2D), and DHPS-knockdown (shDHPS) 4T1 spheroids cultured in lung-like medium (LLM) supplemented with 700 µM aspartate (ASP) and, in the case of control spheroids, also in the absence of aspartate supplementation (NoASPshSCR).

first: Ginevra

last: Doglioni

first: Marine

last: Leclercq

first: Ning

last: An

first: Arnaud

last: Blomme

first: Pierre

last: Close

first: S

last: Fendt


title: Listeria monocytogenes transcriptional response to low concentrations of paracetic acid (PAA)

city: Ithaca

first: Renato

middle: Hohl

last: Orsi

address: 324 Stocking Hall

department: Food Science

institute: Cornell University

summary: Peracetic acid (PAA), a strong oxidizing agent, has been widely used as a disinfectant in food processing settings as it does not produce harmful chlorinated by-products. In the present study, the transcriptional response of Listeria monocytogenes to 2.5 ppm of PAA was assessed using RNA-sequencing (RNA-seq). Our analysis revealed 12 differentially expressed genes, of which 9 were up-regulated (ohrR, ohrA, rpsN, lmo0637, lmo1973, fur, lmo2492, zurM, and lmo1007), and 3 were down-regulated (argG, lmo0604, lmo2156) in PAA treated samples compared to the control samples. A non-coding small RNA (rli32) was also found to be down-regulated. In detail, the organic peroxide toxicity protection (OhrA-OhrR) system, the metal homeostasis genes fur and zurM, the SbrE-regulated lmo0636-lmo0637 operon and a carbohydrate phosphotransferase system (PTS) operon component were induced under exposure of L. monocytogenes to PAA. Hence, this study identified key elements involved in the primary response of L. monocytogenes to oxidative stress caused by PAA. The investigation of the molecular mechanism of PAA response in L. monocytogenes is of utmost importance for the food industry, as this response can be induced in food-processing environments, as a result of inadequate rinsing during the disinfection process, that lead to PAA residues at low concentrations.

overall_design: To identify key elements that are involved in L. monocytogenes paracetic acid (PAA) stress response, RNA-sequencing was performed on rRNA-depleted total RNA extracted from cells exposed to 2.5 ppm  or 0.0 ppm (control) PAA for 0 (within 10 s), 5 and 15 min at 30oC. Differential expression analyses was performed between PAA-exposed and control samples for each time point (0, 5 and 15 minutes) separately. Three biological replicates were obtained for each condition at each time point.

first: Arvaniti

last: Marianna

first: Gabbala

last: Ahmed

first: Orsi

last: Renato

first: Wiedmann

last: Martin


title: Functional genomic analysis of the 68-1 RhCMV-Mycobacteria tuberculosis vaccine reveals an IL-15 response signature that is conserved with vector attenuation

city: Minneapolis

first: Michael

last: Gale

state: Minnesota

address: 689 23rd Ave SE

department: Microbiology and Immunology

institute: University of Minnesota

summary: Here we conducted a bulk RNA-seq analysis of whole blood samples collected from RMs administered with RhCMV vector vaccines using a prime-boost strategy. Two vaccines were used in this study, including 68-1 RhCMV/TB-6Ag (encoding 6 Mtb protein immunogens) and its attenuated variant, 68-1 RhCMV/Δpp71-TB-6Ag (a cell to cell spread-deficient vaccine vector lacking the Rh110 gene encoding the pp71 tegument protein).

overall_design: To examine the differences in the vaccine signatures between RhCMV/TB vaccines, we conducted a functional genomic analysis of whole blood collected from immunized rhesus macaques (RMs; Macaca mulatta) using bulk RNA sequencing (RNA-seq). A total of 24 RMs (males and females) were randomized and assigned to one of two vaccine groups (n=12 per group). Animals received a prime-boost vaccine regimen of 68-1 RhCMV/TB-6Ag (n=12) or 68-1 RhCMV/Δpp71-TB-6Ag vector (n=12) spaced 14 weeks apart that was followed by a second boost administered 92 weeks later (Figure 1a, Table S1). Four RMs of 68-1 RhCMV/Δpp71-TB-6Ag group were not included for the further transcriptional analyses due to lack of second boost data. Whole blood samples collected at pre-vaccination day 0 (D0) and 15 time points following prime immunization were processed for bulk RNA-seq analyses.

first: Cheng-Jung

last: Sung

first: Leanne

last: Whitmore

first: Jean

last: Chang

first: Elise

last: Smith

first: Jennifer

last: Tisoncik-Go

last: Gale Jr


city: Gainesville

first: Andrew

last: Bryant

state: Florida

address: 1345 Center Dr

department: Pulmonary

institute: University of Florida

summary:
Gene expression profile at single cell level of whole lung cells from C57BL/6J mice or global STING knock-out mice (STING-/-), exposed to either room air or chronic hypoxia (4-week 10% oxygen). Study described in manuscript "A non-inteferon role of STING in pulmonary hypertension".
We used scRNAseq to analyze the difference in pulmonary cells and different gene expression across populations between wild type and STING-/- underwent pulmonary hypertension induction with chronic hypoxia.

overall_design: Lung left lobes from 8-week-old male C57BL/6J and STING-/- mice exposed to chronic hypoxia or room air (normoxia) were collected. Lung tissues were digested and suspended into single cell suspension for scRNAseq.

first: Ann

last: Pham

first: Chunhua

last: Fu

first: Zadia

last: Dupee

first: Aline

last: Oliveira


title: APOE from astrocytes restores Alzheimer’s Aβ-pathology and DAM-like responses in APOE deficient microglia

city: Rotselaar

first: Daan

last: Moechars

address: Catharinalaan 43

institute: KULeuven

summary: The major genetic risk factor for Alzheimer’s disease (AD), APOE4, accelerates beta-amyloid (Aβ) plaque formation, but whether this is caused by APOE expressed in microglia or astrocytes is debated. We express here the human APOE isoforms in astrocytes in an Apoe-deficient AD mouse model. This is not only sufficient to restore the amyloid plaque pathology but also induces the characteristic transcriptional pathological responses in Apoe-deficient microglia surrounding the plaques. We find that both APOE4 and the protective APOE2 from astrocytes increase fibrillar plaque deposition, but differentially affect soluble Aβ aggregates. Microglia and astrocytes show specific alterations in function of APOE genotype expressed in astrocytes. Our experiments indicate a central role of the astrocytes in APOE mediated amyloid plaque pathology and in the induction of associated microglia responses.

overall_design: Compariative scRNA-seq study comparing expression profiles of astrocytic human APOE isoforms in an Alzheimer's mouse model

first: Pranav

last: Preman

first: Emre

last: Fertan

first: Leen

last: Wolfs

first: Lutgarde

last: Serneels

first: Disha

last: Shah

first: Jochen

last: Lamote

first: Suresh

last: Poovathingal

first: An

last: Snellinx

first: Renzo

last: Mancuso

first: Sriram

last: Balusu

first: David

last: Klenerman

first: Amaia

last: Arranz

first: Mark

last: Fiers

first: Bart

last: De Strooper


title: eIF4F controls ERK MAPK signaling in melanomas with BRAF and NRAS mutations

city: Brno

first: Core Facility

last: Bioinformatics

address: Kamenice 5

country: Czech Republic

institute: CEITEC - Masaryk University

summary: The eIF4F translation initiation complex plays a critical role in melanoma resistance to clinical BRAF and MEK inhibitors. In this study, we uncover a novel function of eIF4F in the negative regulation of the RAS/RAF/MEK/ERK mitogen-activated protein kinase (MAPK) signaling pathway. We demonstrate that eIF4F is essential for maintaining optimal ERK signaling intensity in treatment-naïve melanoma cells harboring BRAF or NRAS mutations. Specifically, the dual-specificity phosphatase DUSP6/MKP3, which acts as a negative feedback regulator of ERK activity, requires continuous production in an eIF4F-dependent manner to limit excessive ERK signaling driven by oncogenic RAF/RAS mutations. Treatment with small molecule eIF4F inhibitors disrupts the negative feedback control of MAPK signaling, leading to ERK hyperactivation and EGR1 overexpression in melanoma cells in vitro and in vivo. Furthermore, our quantitative analyses reveal a high spare signaling capacity in the ERK pathway, suggesting that eIF4F-dependent feedback keeps the majority of ERK molecules inactive under normal conditions. Overall, our findings highlight the crucial role of eIF4F in regulating ERK signaling flux and suggest that pharmacological eIF4F inhibitors can disrupt the negative feedback control of MAPK activity in melanomas with BRAF and NRAS activating mutations.

overall_design: We used next-generation RNA sequencing (RNA-seq) to perform unbiased analyses of transcriptome changes in two human melanoma cell lines in response to 20 h treatments with a small-molecule eIF4F inhibitor Rocaglamide A.

first: Stjepan

last: Uldrijan

first: Karolina

last: Smolkova

first: Boris

last: Tichy

first: Nicolas

last: Blavet


title: Aspartate signaling increases the aggressiveness of lung metastases by inducing eIF5A-mediated translation (Polysomal RNA-Seq)

first: Juan

last: Fernández-García

overall_design: RNA-seq of matched polysomal/sub-polysomal mRNA fractions extracted from control (shSCR), GRIN2D-knockdown (shGRIN2D), and DHPS-knockdown (shDHPS) 4T1 spheroids cultured in lung-like medium (LLM) supplemented with 700 µM aspartate (ASP) and, in the case of control spheroids, also in the absence of aspartate supplementation (NoASPshSCR).

first: Ginevra

last: Doglioni

first: Marine

last: Leclercq

first: Ning

last: An

first: Arnaud

last: Blomme

first: Pierre

last: Close

first: Sarah

last: Fendt


title: Aspartate signaling increases the aggressiveness of lung metastases by inducing eIF5A-mediated translation

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

overall_design: Refer to individual Series


title: Environmental alkalization suppresses deployment of virulence strategies in Pseudomonas syringae pv. tomato DC3000

status: Public on Oct 03 2024

first: Bryan

last: Swingle

address: 302 Plant Science Building

institute: USDA-ARS

summary: Plant pathogenic bacteria encounter a drastic increase in apoplastic pH during the early stages of plant immunity. The effects of alkalization on pathogen-host interactions have not been comprehensively characterized. Here we used a global transcriptomic approach to assess the impact of environmental alkalization on Pseudomonas syringae pv. tomato DC3000 in vitro. In addition to the Type 3 Secretion System (T3SS), we found expression of genes encoding other virulence factors such as iron uptake, and coronatine biosynthesis to be strongly affected by environmental alkalization. We also found activity of extracytoplasmic function sigma factor, AlgU, was induced at pH 5.5 and suppressed at pH 7.8, which are pH levels that this pathogen would likely experience before and during pattern triggered immunity, respectively. This pH-dependent control requires the presence of periplasmic proteases, AlgW and MucP, that function as part of the environmental sensing system that activates AlgU in specific conditions. This is the first example of pH-dependency of AlgU activity, suggesting a regulatory pathway model where pH affects the proteolysis-dependent activation of AlgU. These results contribute to deeper understanding of the role apoplastic pH has on host-pathogen interactions.

sample_organism: Pseudomonas syringae pv. tomato str. DC3000

overall_design: Comparative gene expression profiling analysis of RNA-seq data for Pst DC3000 WT and algU deletion strains across pH-modified KB media growth condition at pH 5.5, 6.5 and 7.8

first: Zichu

last: Yang

first: Haibi

last: Wang

first: Robert

last: Keebler

first: Amelia

last: Lovelace

first: Hsiao-Chun

last: Chen

first: Brian

last: Kvitko


title: The neurotrophic factor MANF regulates autophagy and lysosome function to promote proteostasis in Caenorhabditis elegans

city: Charleston

first: Jessica

last: Hartman

state: South Carolina

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

department: Biochemistry and Molecular Biology

institute: Medical University of South Carolina

summary: Mesencephalic astrocyte-derived neurotrophic factor (MANF) is the only neurotrophic factor to be conserved between C. elegans and humans. MANF is a unique neutrotophic factor, as it is a resident ER protein in cells that express it and has a role in regulating ER stress response to tunicamycin exposure and pathogenic Pseudomonas exposure. We sought to further understand the role of MANF by comparing a deletion allele, an overexpression line, and the wild-type in this RNAseq experiment.

overall_design: In order to test the role of MANF, we looked at aging animals - we age-synchronized eggs via bleaching gravid adults and grew worms to adulthood. Adult worms were transferred to new plates daily during reproduction in order to separate adults from their offspring. On 8 days post-hatch, we collected worms and isolated RNA, then the Duke Sequencing Core performed poly-A mRNA enrichment and RNA-sequencing.

first: Hartman

last: Jessica

first: Taylor

last: Shane

first: Gupta

last: Bhagwati


title: Aspartate signaling increases the aggressiveness of lung metastases by inducing eIF5A-mediated translation (scRNA-Seq)

first: Juan

last: Fernández-García

overall_design: scRNA-seq from lungs of female BALB/c mice treated with control medium (CM) or medium containing tumor-secreted factors (TSF) for 3 weeks (3 injections per week), and subsequently either euthanized (pre-metastatic niche, CM0/TSF0), or injected i.v. with 25,000 CD90.1-expressing 4T1 cancer cells and then euthanized 11 days (metastatic seeding, CM1/TSF1) or 16 days (metastatic colonization, CM2/TSF2) after cancer-cell injection.

first: Ginevra

last: Doglioni


title: The early transcriptional and post-transcriptional responses to fluconazole in sensitive and resistant Candida albicans (5P-Seq)

first: Irene

last: Stevens

summary: Candida albicans is a leading cause of fungal infections in immunocompromised patients. Management of candidemia relies on a few antifungal agents, with fluconazole being first line therapy. The emergence of fluconazole-resistant strains highlights the pressing need to improve our molecular understanding of the drug response mechanisms. By sequencing the 5’P mRNA degradation intermediates, we show that co-translational mRNA decay is common in C. albicans and characterize how in vivo 5´-3´ exonuclease degradation trails the last translating ribosome. Thus, the study of the 5'P mRNA degradome (5PSeq) offers a simple and affordable way to measure ribosome dynamics and identify codon specific ribosome stalls in response to drugs and amino acid deprivation. Building upon this, we combine RNA-Seq and 5PSeq to study the early response of sensitive and resistant C. albicans isolates to fluconazole. Our results highlight that transcriptional responses, rather than changes in ribosome dynamics, are the main driver of Candida resistance to fluconazole.

overall_design: 5' phosphate sequencing (5Pseq) of Candida albicans SC5314, PLC124, MAY7 and MAY478 treated with fluconazole (1 μg/ml, 1xMIC) for 30 min versus control (2% DMSO). Candida SC5314 treated with cycloheximide (0.067 mg/ml) for 15 min, and proline-arginine-ornithine depletion experiments with corresponding controls.

first: Fitz

last: Silao

first: Susanne

last: Huch

first: Honglian

last: Liu

first: Kicki

last: Ryman

first: Adriana

last: Carvajal-Jimenez

first: Per

last: Ljungdahl

first: Vicent

last: Pelechano


title: Polygenic Risk for Alcohol Use Disorder Affects Cellular Responses to Ethanol Exposure in a Human Microglial Cell Model II

city: Piscataway

first: Ronald

middle: P.

last: Hart

state: NJ

address: 604 Allison Rd Rm B430

department: Cell Biology & Neuroscience

institute: Rutgers University

summary: Polygenic risk scores (PRS) assess genetic susceptibility to Alcohol Use Disorder (AUD), yet their molecular implications remain underexplored. Neuroimmune interactions, particularly in microglia, are recognized as significant contributors to AUD pathophysiology. We investigated the interplay between AUD PRS and ethanol in human microglia derived from iPSCs from individuals with high-or low-PRS (HPRS or LPRS) of AUD. Ethanol exposure induced elevated CD68 expression and morphological changes in microglia, with differential responses between HPRS and LPRS microglial cells. Transcriptomic analysis revealed expression differences in MHCII complex and phagocytosis-related genes following ethanol exposure; HPRS microglial cells displayed enhanced phagocytosis and increased CLEC7Aexpression, unlike LPRS microglial cells. Synapse numbers in co-cultures of induced neurons with microglia after alcohol exposure were lower in HRPS co-cultures, suggesting possible excess synapse pruning. This study provides insights into the intricate relationship between AUD PRS, ethanol, and microglial function, potentially influencing neuronal functions in developing AUD.

overall_design: Cultures were prepared of iPS cells from multiple iPSC lines prepared from subjects with AUD and high polygenic risk (HPRS) and unaffected with low polygenic risk (LPRS).  Total cellular RNA was prepared for quality assurance assays.

first: Xindi

last: Li

first: Jiayi

first: Anat

last: Kreimer

middle: P

first: Zhiping

last: Pang


title: Comprehensive profiling of Roquin binding preferences for RNA stem-loops

city: Marburg

first: Julia

middle: E.

last: Weigand

address: Marbacher Weg 6

department: Pharmacy

country: Germany

institute: University of Marburg

summary: Roquin proteins target specific RNA stem-loop motifs with their unique ROQ domain for mRNA degradation. The cellular levels of mRNAs are controlled by these post-transcriptional cis-regulatory stem-loop elements in the 3'-untranslated region (3'-UTR). Here, we first provide an RBNS (RNA Bind-n-Seq) experiment using pre-structured RNA libraries as input. This sRBNS (structured RBNS) was used to map Roquin's RNA binding preferences at nucleotide resolution, revealing preferences for loop sizes and consensus motifs for CDEs and ADEs.

overall_design: Structured RNA Bind-n-Seq (sRBNS) experiment to test the ability of Roquin's unique ROQ domain to interact with specific RNA structures. Different pre-structured RNA pools (1 µM) with random loop nucleotides flanked by constant stem sequences were incubated with different concentrations of the coreROQ domain (10, 50, 500 nM).

first: Lasse

last: Oberstrass

first: Chiara

last: Lichtenthaeler

first: Jan-Niklas

last: Tants

first: Sara

last: Ali

first: David

last: Mathews

first: Andreas

last: Schlundt


title: Global coupling of R-loops with transcription dynamics modulates early development and survival of Drosophila

city: Wuhan

first: Shaobo

last: Liang

state: Hubei Sheng

address: BaYi Road, LUOJIA HILL

country: China

institute: Wuhan University

summary: R-loops are involved in many biological processes in cells, yet the regulatory principles for R-loops in vivo and their impact on development remain to be explored. Here, we modified the CUT&Tag strategy to profile R-loops in Drosophila at multiple developmental stages. While high GC content promotes R-loop formation in mammalian cells, it is not required in Drosophila. In contrast, RNAPII abundance on chromatin appears to be a universal inducing factor for R-loop formation in the Drosophila genome, including active promoters and enhancers, and H3K27me3 decorated repressive regions and intergenic repeat sequences. Importantly, such a regulatory relationship is dynamically maintained throughout development and many development-related transcription factors that may regulate RNAPII activation and R-loop dynamics were uncovered. By ablating the elongation factor Spt6, we further showed the global R-loop induction coupled with RNAPII pausing, genome instability, and defects in larvae survival and cuticle development. Overexpression of rnh1 largely reversed the upregulation or downregulation of gene expression depending on gene length, DNA damage, and defects in survival and cuticle development. Altogether, our findings indicate that dynamic R-loop regulation is dictated by RNAP pausing and transcription activity, and plays a feedback role in gene regulation, genome stability maintenance, and Drosophila development.

overall_design: To explore the regulatory mechanism of R-loop in Drosophila and embryonic development, we constructed a multi-temporal R-loop map of embryonic development. As well as pol2-chip-seq and cut&tag before and after interfering transcription factor spt6. RNA-seq experiments were performed on da-GAL4, da-GAL4>Spt6-RNAi, da-GAL4>rnh1 and da-GAL4>rnh1, Spt6-RNAi Drosophila in 14-16h embryos

first: Liang

last: Chen

first: Xianhong

last: Zhang


title: Oncogenic KRAS Mutations Confer a Unique Mechanotransduction Response to Peristalsis in Colorectal Cancer Cells

city: Houston

first: Abigail

middle: Jones

last: Clevenger

address: 6500 Main Street

department: Biomedical Engineering

institute: Rice University

summary: Colorectal cancer (CRC) tumors start as precancerous polyps on the inner lining of the colon or rectum, where they are exposed to the mechanics of colonic peristalsis. Our previous work leveraged a custom-built peristalsis bioreactor to demonstrate that colonic peristalsis led to cancer stem cell enrichment in colorectal cancer cells. However, this malignant mechanotransductive response was confined to select CRC lines that harbored an oncogenic mutation in the KRAS gene. In this work, therefore, we explored the involvement of activating KRAS mutations on peristalsis-associated mechanotransduction in CRC. Peristalsis enriched the cancer stem cell marker LGR5 in KRAS mutant (G13D, etc.) lines, in a Wnt-independent manner. Conversely, LGR5 enrichment in wild type KRAS lines exposed to peristalsis were minimal. LGR5 enrichment downstream of peristalsis translated to increased tumorigenicity in vivo in KRAS mutant vs. wild type lines. Differences in mechanotransduction response was additionally apparent via unbiased gene set enrichment analysis, where many unique pathways were enriched in wild type vs. mutant lines, in response to peristalsis. Interestingly, peristalsis also triggered β-catenin nuclear localization independent of Wnt, particularly in KRAS mutant lines. The central involvement of KRAS in the mechanotransductive responses was validated via gain and loss of function strategies. β-catenin activation and LGR5 enrichment downstream of peristalsis converged to the activation of the MEK/ERK kinase cascade, that remains active in cells that harbor oncogenic KRAS mutations. Taken together, our results demonstrated that oncogenic KRAS mutations conferred a unique peristalsis-associated mechanotransduction response to colorectal cancer cells, resulting in cancer stem cell enrichment and increased tumorigenicity. These mechanosensory connections can be leveraged in improving the sensitivity of emerging therapies that target oncogenic KRAS.

overall_design: To investigate the difference in KRAS mutant CRC cellular response to peristalsis, three cell lines were used: 1) HCT116 KRAS mutant, 2) RKO KRAS wild type and 3) RKO KRAS mutant. We performed gene expression analysis on all three cell lines in static and peristalsis conditions.

first: Shreya

last: Raghavan


city: Hong Kong

first: Feng

last: Yang

address: Room 503, 5/F, Li Ka Shing Medical Sciences Building, Prince of Wales Hospital

department: Chemical Pathology

institute: The Chinese University of Hong Kong

summary: To identify the YTHDF1 binding changes upon DHX36 loss, we conducted and analyzed YTHDF1 CLIP-seq data in WT and DHX36-KO HEK293T cells.

overall_design: YTHDF1 CLIP-seq in WT and DHX36-KO HEK293T cells with 2 replicates

first: Yuwei

first: Jieyu

last: Zhao

first: Chun

last: Kwok

first: Huating

last: Wang


title: CD99 contributes to the EWS:FLI transcriptome by specifically affecting FOXM1-targets involved in the G2/M cell cycle phase

city: Bologna

first: Emanuela

last: Palmerini

address: via G.C.Pupilli, 1

country: Italy

institute: IRCCS Istituto Ortopedico Rizzoli

summary: Ewing sarcoma (EwS), a highly aggressive malignancies affecting children and young adults, is primarily driven by a distinctive oncogenic fusion (EWSR1-ETS), whose activity is a principal source of epigenetic and clinical heterogeneity. However, another molecule (CD99) is constantly present in EWS cells. Despite the fact that this molecule is widely reported to modulate EWS genetic profile and tumor malignancy, the relevance of CD99 alone or in association with the chimera has been largely ignored. In this study, we explored the dynamic relationship between EWS::FLI1, the main fusion observed in EWS, and CD99 through experimental inducible models for the expression of one or the other molecule. The transcriptome of cells with or without expression of EWS::FLI or CD99 were analyzed in dynamics and associated with in tumor cell growth. CD99-associated EWS gene profile was found to have commonalities with that induced by EWS::FLI but also some peculiar diversities. In particular, both EWS::FLI and CD99 regulates targets of the DREAM complex but CD99 expression specifically impacts on those genes that are targets of FOXM1 and involved in the setting of G2/M phase of cell cycle. Most of these CD99-regulated genes were found to be associated to worst clinical prognosis in two different clinical datasets on the freely available R2 platform , further supporting the clinical relevance of CD99-mediated regulation of EWS gene expression.

overall_design: We developed two inducible cell models using a tetracycline-inducible shRNA system targeting EWS::FLI1 or CD99 transcripts (named as A673pTERshEWS::FLI1 or A673pTERshCD99 respectively)to determinate whether these two molecules work in a dependent or synergistic manner.We performed the gene expression profiles associated with their silencing and recovery by SurePrint G3 Human Gene Expression v3 8x60K Microarray Kit, cat# G4851C, Agilent Technologies in two biological replicate fro both models.

first: Michela

last: Pasello

first: Maria

last: Laginestra

middle: C

last: Manara

first: Lorena

last: Landuzzi

first: Francesca

last: Ruzzi

first: Margherita

last: Maioli

first: Evelin

last: Pellegrini

first: Alessandra

last: De Feo

first: Pier-Luigi

last: Lollini

first: Katia

last: Scotlandi


title: CaClust: linking genotype to transcriptional heterogeneity of follicular lymphoma using BCR and exomic variants (K6B)

city: Leiden

first: Cornelis

last: van Bergen

address: Albinusdreef 2

department: Hematology

country: Netherlands

institute: Leiden University Medical Center

summary: Tumours exhibit high genotypic and transcriptional heterogeneity. Both affect cancer progression and treatment, but have been predominantly studied separately in follicular lymphoma. To comprehensively investigate the evolution and genotype-to-phenotype maps in follicular lymphoma, we introduce CaClust, a probabilistic graphical model integrating deep whole exome, single-cell RNA and B-cell receptor sequencing data to infer clone genotypes, cell-to-clone mapping, and single-cell genotyping. CaClust outperforms a state-of-the-art model on simulated and patient data. In-depth analyses of single cells from four samples showcase effects of driver mutations, follicular lymphoma evolution, possible therapeutic targets, and single-cell genotyping that agrees with an independent targeted resequencing experiment.

overall_design: Dataset K6B: 10X Genomics of flowcytometry purified follicular lymphoma cells, 5' chemistry, full transcriptome and BCR enrichment; targeted amplification of selected genes on sc cDNA samples and full length single molecule PacBio sequencing

first: Hendrik

last: Veelken

first: Kazimierz

last: Oksza-Orzechowski

first: Ewa

last: Szczurek


city: Trondheim

first: Kristine

last: Misund

address: Prinsesse Kristinas gt 1

department: Department of Clincal and Molecular Medicine

country: Norway

institute: Norwegian University of Science and Technology

summary: This RNAseq analysis is performed as a part of the work "Heterogeneous expression of cell surface markers in human myeloma cell lines: implications for disease modelling and therapeutic targeting", where expression levels both at the cell surface level and transcriptomic level are analysed.

overall_design:
To analyze the gene expression levels in our in-house Multiple Myeloma (MM) cell lines, VOLIN, KJON, FOLE, URVIN, IH-1 and OH-2 was RNA sequenced.We also included the MM cell lines ANBL-6, INA-6 and JJN-3 as controls, and to check that they were similar to already published data. 2 biological replicates were sequenced for each cell line.

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Raw data are not included, due to regulations in the Norwegian law.
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first: Toril

last: Holien


title: CaClust: linking genotype to transcriptional heterogeneity of follicular lymphoma using BCR and exomic variants (K5B)

overall_design: Dataset K5B: 10X Genomics of flowcytometry purified follicular lymphoma cells, 5' chemistry, full transcriptome and BCR enrichment.


title: CaClust: linking genotype to transcriptional heterogeneity of follicular lymphoma using BCR and exomic variants (K7B)

type: Genome variation profiling by high throughput sequencing

overall_design: Dataset K7B: 10X Genomics of flowcytometry purified follicular lymphoma cells, 5' chemistry, full transcriptome and BCR enrichment; targeted amplification of selected genes on sc cDNA samples and full length single molecule PacBio sequencing


title: Epigenomic maps of the rainbow trout reveal epiQTL signatures of genetic selection and regulatory elements involved in controlling gene expression and genome evolution

city: Murfreesboro

first: Rafet

last: Al-Tobasei

state: TN

address: 358 KOM

department: Computer Science Department

institute: Middle Tennessee State University

type: Methylation profiling by high throughput sequencing

summary:
Over the last century, rainbow trout (RBT) has been widely used as a biological model in many biological disciplines and has become one of the best-studied fish.  For example, RBT is an excellent model to study gene evolution as it is a partially tetraploid organism that underwent a whole-genome duplication (salmonid-specific 4th WGD) followed by a partial re-diploidization and significant genome rearrangements.  In the meantime, RBT domestication spread quickly and globally to six continents for aquaculture and recreation.
Efforts to sequence a pangenome reference have begun, and genome sequence is available for at least three rainbow trout clonal lines.  However, epigenome reference annotations are needed to understand the functional genomic basis of RBT's phenotypic, environmental, and evolutional variations.
This study provides a comprehensive catalog and epigenome annotation tracks of the rainbow trout.  Gene regulatory elements, including chromatin histone modifications, chromatin accessibility, and DNA methylation, were identified by integrating data from ChIP-seq, ATAC-seq, and Methyl Mini-seq across RBT tissues together with RNA-seq gene expression data sets.

overall_design:
Two biological replicates for six tissues (Brain, Intestine, Kidney, Liver, Spleen, and White muscle) were used for ChIP-Seq (H3K4me3, H3K4me1, H3K27ac, H3K27me3, input control)

Two biological replicates for six tissues (Brain, Intestine, Kidney, Liver, Spleen, and White muscle) were used for Bisulfite Methylation and Three tissue with two bioloical replicates (Brain, Liver, and Spleen) used for ATAC seq.

first: Mohamed

last: Salem


title: Translation of zinc finger domains induces ribosome collision and Znf598-dependent mRNA decay in zebrafish [Ribo-seq]

city: Kita-ku, Kyoto

first: Kota

last: Ishibashi

state: Kyoto

address: Kamigamo-motoyama

department: Department of Frontier Life Sciences

country: Japan

institute: Kyoto Sangyo University

summary: Quality control of translation is crucial for maintaining cellular and organismal homeostasis. Obstacles in translation elongation induce ribosome collision, which is monitored by multiple sensor mechanisms in eukaryotes. The E3 ubiquitin ligase Znf598 recognizes collided ribosomes, triggering ribosome-associated quality control (RQC) to rescue stalled ribosomes and no-go decay (NGD) to degrade stall-prone mRNAs. However, the impact of RQC and NGD on maintaining the translational homeostasis of endogenous mRNAs has remained unclear. In this study, we investigated the endogenous substrate mRNAs of NGD during the maternal-to-zygotic transition (MZT) of zebrafish development. RNA-Seq analysis of zebrafish znf598 mutant embryos revealed that Znf598 downregulates mRNAs encoding the C2H2-type zinc finger domain (C2H2-ZF) during the MZT. Reporter assays and disome profiling indicated that ribosomes stall and collide while translating tandem C2H2-ZFs, leading to mRNA degradation by Znf598. Our results suggest that NGD maintains the quality of the translatome by mitigating the risk of ribosome collision at the abundantly present C2H2-ZF sequences in the vertebrate genome.

overall_design: Ribosome profiling

first: Yuichi

last: Shichino

first: Penxun

last: Han

first: Shintaro

last: Iwasaki

first: Yuichiro

last: Mishima


title: Translation of zinc finger domains induces ribosome collision and Znf598-dependent mRNA decay in zebrafish [RNA-seq]

overall_design: RNA-seq

first: Seisuke

last: Kimura


title: CaClust: linking genotype to transcriptional heterogeneity of follicular lymphoma using BCR and exomic variants (K4B)

overall_design: Dataset K4B: 10X Genomics of flowcytometry purified follicular lymphoma cells, 5' chemistry, full transcriptome and BCR enrichment.


title: Discovery of BBO-8520, a first-in-class direct and covalent dual inhibitor of GTP-bound (ON) and GDP-bound (OFF) KRASG12C

status: Public on Oct 02 2024

city: Palo Alto

first: Jin Hyun

last: Ju

address: 3160 Porter Dr, Suite 250

institute: BridgeBio

summary: Approved inhibitors of KRASG12C prevent oncogenic activation by sequestering the inactive, GDP-bound (OFF) form rather than directly binding and inhibiting the active, GTP-bound (ON) form of the protein. This suboptimal mechanism provides no direct target coverage against KRASG12C(ON). Expectedly, adaptive resistance to KRASG12C (OFF)-only inhibitors is observed in association with increased expression and activity of KRASG12C(ON). To provide optimal KRASG12C target coverage, we have developed BBO-8520, a first-in-class, direct dual inhibitor of KRASG12C(ON) and (OFF) forms. BBO-8520 binds in the Switch-II/Helix3 pocket, covalently modifies the target cysteine and disables effector binding to KRASG12C(ON). BBO-8520 exhibits potent signaling inhibition in growth factor activated states where current (OFF)-only inhibitors demonstrate little measurable activity. In vivo, BBO-8520 demonstrates rapid target engagement and inhibition of downstream signaling, resulting in durable tumor regression in multiple models, including those resistant to KRASG12C (OFF)-only inhibitors.

overall_design:
To investigate the transcriptional regulation of BBO-8520, a dual KRAS G12C (OFF) and (ON)  inhibitor, compared to KRAS G12C (OFF) inhibitors in the the KRAS G12C homozygous pancreatic cell line MIA PaCa-2
MIA PaCa-2 cells were treated with DMSO, BBO-8520 (30nM), adagrasib (300nM) or sotorasib (1uM ) for 24 hours with 5 replicates/condition and collected for RNA-seq analysis
We then performed gene expression profiling anlysis using data obtained from RNA-seq of the MIA PaCa-2 cell line treated with the 4 different compounds

first: James

last: Stice

first: Devansh

last: Singh

first: Stevan

last: Jeknic


title: Translation of zinc finger domains induces ribosome collision and Znf598-dependent mRNA decay in zebrafish

overall_design: Refer to individual Series


title: Gene expression and DNA methylation changes in response to hypoxia in toxicant-adapted Atlantic killifish (Fundulus heteroclitus)

city: Woods Hole

first: Neelakanteswar

last: Aluru

address: 45 Water Street

department: Biology

institute: Woods Hole Oceanographic Institution

summary: Hypoxia in coastal waters is an increasing concern, with the frequency of hypoxic events rising because of climate change and human-driven impacts such as eutrophication. The Atlantic killifish (Fundulus heteroclitus), an estuarine species known for its resilience to environmental stressors, provides a valuable model to investigate the physiological, transcriptional, and epigenetic mechanisms underlying adaptation. Some killifish populations, like those in New Bedford Harbor (NBH), Massachusetts, have evolved resistance to dioxin-like polychlorinated biphenyls (PCBs) due to chronic exposure. These populations offer a unique opportunity to investigate how toxicant resistance might influence responses to secondary stressors like hypoxia. The objective of this study is to characterize the impacts of evolved resistance to toxicants on the ability to cope with acute hypoxia. We compared hepatic gene expression and DNA methylation patterns in response to two levels of hypoxia in killifish from NBH and Scorton Creek (SC), a reference population from a relatively pristine environment. We hypothesized that NBH fish would exhibit altered molecular responses to hypoxia due to trade-offs associated with toxicant resistance. Our results revealed significant differences between the two populations. SC fish demonstrated a dose-dependent increase in gene expression in response to hypoxia, while NBH fish exhibited a muted transcriptional response to severe hypoxia, indicating potential impairment in their ability to cope with hypoxic stress. Interestingly, NBH fish showed significant DNA methylation changes in response to hypoxia, while SC fish did not exhibit notable epigenetic alterations. These findings suggest that toxicant-adapted killifish may face trade-offs in their response to additional environmental stressors, with potential consequences for their ability to survive in increasingly hypoxic coastal habitats. Further research is needed to elucidate the functional implications of these epigenetic modifications and their role in adaptive stress responses.

overall_design: Juvenile fish from two different populations (New Bedford Harbor and Scorton Creek) were exposed to either normoxia (control group) or two different concentrations of hypoxia (5 and 10%) for 6 hours. At the end of the exposure, liver samples were dissected and RNA/DNA was isolated simultaneously.

first: Yaamini

last: Venkataraman

first: Christopher

last: Murray

first: Veronica

last: DePascuale


title: Microarray analysis of human cord blood derived mast cells (hCBMC) activated by Interleukin-33 (IL-33)

city: Jeddah

first: Sherin

last: Bakhashab

address: Abdullah Sulyman

department: Biochemistry

country: Saudi Arabia

institute: King Abdulaziz University

summary:
Mast cells respond to acute and chronic exposure to IL-33 by differing the expression of their receptors, cytokines and proteases.
We aim to characterize the distinct acute and chronic responses of mast cells to IL-33 by analyzing hCBMC expression profile of pro- and anti- inflammatory mediators.

overall_design: CD34+ cells were isolated from human umbilical cord blood and differentiated into mast cells using rhIL-6 and rh-SCF for 6-8 weeks. hCBMCs were stimulated with 10 and 20 ng/mL of IL-33 for 6 and 24 hours each. Total RNA was extracted from hCBMCs and untreated controls and hybridized using Affymetrix Human Gene ST 1.0 arrays.

first: Ghalya

last: Banafea

first: Farid

last: Ahmed

first: Nadia

last: Bagatian

first: Ohoud

last: Subhi

first: Kalamegam

last: Gauthaman

first: Mahmood

last: Rasool

first: Hans-Juergen

last: Schulten

first: Peter

last: Pushparaj


title: Muscle-specific rescue of Bmal1 impacts survival and health span of the Bmal1-KO mouse

city: Gainesville

first: Karyn

last: Esser

address: 1345 Center Drive M5-44

department: Department of Physiology and Aging

institute: University of Florida

summary: This dataset was generated to investigate the muscle and systemic effects of muscle-specific Bmal1 expression in the Bmal1-KO mouse model

overall_design: Adeno-associated virus were used to rescue the expression of the clock gene Bmal1 in the Bmal1 KO. Skeletal muscle, lung, heart, liver, and white fat tissues from 10 and 40-week-old WT, KO, and KO+AAV male mice were collected at ZT1 to characterize the muscle and systemic effects of muscle-specific expression of Bmal1.

first: Miguel

last: Gutierrez-Monreal

last: Wolff

first: Zhiguang

last: Huo


title: Organ-specific Sympathetic Innervation Defines Visceral Functions

city: Pasadena

first: Tongtong

last: Wang

address: 1200 E California Blvd

institute: California Institute of Technology

summary: We performed single-cell RNA-seq (10x Chromium 3' v.3.1) on dissociated single cell suspensions from mouse (C57BL/6) celiac and superior mesenteric sympathetic ganglia to determine transcriptomic cell types. We identified sympathetic neuron types that specifically innervate unique combinations of visceral organs and control physiological functions.

overall_design: We used 20 animals in total, with male and female mice in equal measure. All animals were 7-8 weeks of age.

first: Yuki

last: Oka


title: Effect of autophagy inhibition (ATG7/13 KD) on the transcriptome of proliferating and senescent cells

city: Seoul

first: Jaejin

last: Kim

address: Gwanak-ro 1

country: South Korea

institute: Seoul National University

summary: To identify differentially expressed genes by autophagy inhibition during cellular senescence, we performed transcriptional profiling by RNA sequencing (RNA-seq) with or without autophagy inhibition (using a shRNA targeting either ATG7 or ATG13), before and after senescence induction using a stress-induced senescence model (bleomycin-induced senescence)

overall_design: IMR90 cells were treated with bleomycin (10 ug/ml for 24 h) to induce senescence. Autophagy was inhibited by expression of a shRNA targeting either ATG7 or ATG13 in proliferating cells and senescent cells. Total RNA was isolated using the acid phenol extraction method. One microgram of high-quality RNA samples (RIN > 7.0) was used to construct RNA-seq libraries using the TruSeq Stranded mRNA LT Sample Prep Kit (Illumina). RNA-seq was performed on an Illumina NovaSeq 6000 sequencer.

first: Yeonghyeon

last: Lee

first: Taerang

last: Jeon

first: Chanhee

last: Kang


title: Alternative lipid synthesis in response to phosphate limitation promotes antibiotic tolerance in Gram-negative ESKAPE pathogens

city: Richardson

first: Joseph

middle: Michael

last: Boll

state: TX

address: 800 W Campbell

department: Biological Sciences

institute: University of Texas at Dallas

summary: Acinetobacter baumannii is an ESKAPE pathogen that rapidly develops resistance to antibiotics and persists for extended periods in the host or on abiotic surfaces. Survival in environmental stress such as phosphate scarcity, represents a clinically significant challenge for nosocomial pathogens. In the face of phosphate starvation, certain bacteria encode adaptive strategies, including the substitution of glycerophospholipids with phosphorus-free lipids. In bacteria, phosphatidylethanolamine, phosphatidylglycerol, and cardiolipin are conserved glycerophospholipids that can form lipid bilayers, particularly in the presence of other lipids. Here, we demonstrate that in response to phosphate limitation, conserved regulatory mechanisms induce alternative lipid production in A. baumannii. Specifically, phosphate limitation induces formation of three lipids, including amine-containing ornithine and lysine aminolipids. Mutations that inactivate aminolipid biosynthesis exhibit fitness defects relative to wild type in colistin growth and killing assays. Furthermore, we show that other Gram-negative ESKAPE pathogens accumulate aminolipids under phosphate limiting growth conditions, suggesting aminolipid biosynthesis may represent a broad strategy to overcome cationic antimicrobial peptide-mediated killing.

overall_design: Differential gene expression in A. baumannii strain ATCC 17978 in minimal medium supplemented with excess (1mM) vs. limiting (50 μM) phosphate at OD600 of ~0.5 in triplicate

first: Roberto

last: Olea-Ozuna

first: Melanie

last: Campbell

first: Samantha

middle: Y

last: Quintanilla

first: Sinjini

last: Nandy

first: Jennifer

last: Brodbelt


title: PPARγ-dependent Remodeling of Translational Machinery in Adipose Progenitors is Impaired in Obesity [Polysome-seq]

city: Los Angeles

first: Xia

last: Yang

state: California

address: 2000 Terasaki Life Sciences Bldg

department: Department of Integrative Biology and Physiology

institute: University of California, Los Angeles

summary:
Adipose tissue plays a pivotal role in energy homeostasis and metabolic regulation. However, in obesity, the remarkable adaptability of adipose tissue becomes impaired. The underlying mechanisms behind this limited adaptability remain poorly understood. In this study, we investigate a novel layer of regulation involving translation in the adipose stromal vascular fraction, examining its response to obesity and acute PPAR? agonist treatment with rosiglitazone. Using single-cell RNA sequencing, we establish a transcriptional profile atlas of cellular remodeling from obese to lean-like states in inguinal and epididymal adipose tissue following rosiglitazone treatment. Notably, both stromal fractions exhibit a downregulation of inflammation-related transcripts and an upregulation of lipid-related metabolism and ribosomal transcripts. Adipocyte progenitor and preadipocyte populations display enhanced ex-vivo differentiation potential and upregulation in ribosome and peptide chain elongation pathways. This ribosomal remodeling is directly driven by PPAR? binding to gene promoters of ribosomal factors. Furthermore, we have characterized the translatome in the epididymal stromal fraction, highlighting a buffering response and fat-exclusive preferential translation after rosiglitazone treatment. Enhanced translation efficiency in rosiglitazone-elicited polysomes promotes the translation of transcripts containing G-rich sequences in their 5? untranslated regions. Our findings shed light and provide a resource on how rosiglitazone remodels the adipose stromal vascular fraction, both dependent and independent of PPAR?. Importantly, we uncover translatome remodeling as a major new mechanism for maintaining translation homeostasis and preserving adipose tissue health in obesity.
PLEASE NOTE: the raw fastq files for sample inputVeh3 were corrupted and, therefore, no longer retrievable. However, the processed data file (inputVeh3_quant.sf) for this sample is still usable.

overall_design:
Stromal vascular fraction (SVF) of Lepob/Lepob mice were isolated and primary cells were differentiated for 3 days in vitro. Cell lysates were used to perfome polysome profilling, and fractions containing 3+ribosomes per mRNA  were used for  RNA extraction and sequencing.

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Submitter states that missing raw files are due to file corruption.
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first: Mirian

last: De Siqueira

first: Yutian

last: Zhao

first: Gaoyan

last: Li

first: Siqi

first: Xinshu

last: Xiao

first: Claudio

last: Villanueva


title: single nuclei combined ATAC/RNA sequencing (10X Multiome) of airway-resident cells isolated from bronchoalveolar lavage fluid (BALF) of naive mice or mice recovered from SARS-CoV-2 (mouse-adapted strain MA10)

city: New York

first: Michael

last: Bale

address: 1300 York Ave

institute: Weill Cornell

summary: Pathogen encounter results in long-lasting epigenetic imprinting that shapes diseases caused by heterologous pathogens. The breadth of this innate immune memory is of particular interest in the context of respiratory pathogens with increased pandemic potential and wide-ranging impact on global health. Here, we investigated epigenetic imprinting across cell lineages in a disease relevant murine model of SARS-CoV-2 recovery. Past SARS-CoV-2 infection resulted in increased chromatin accessibility of type I interferon (IFN-I) related transcription factors and transcriptionally poised antiviral genes in airway-resident macrophages. Mechanistically, viral pattern recognition and canonical IFN-I signaling were required for establishment of this innate immune memory and resulting augmented secondary antiviral responses. SARS-CoV-2-associated innate immune memory in airway-resident macrophages was necessary and sufficient to ameliorate secondary disease caused by the heterologous respiratory pathogen influenza A virus. Insights into how innate immune memory shapes outcome of heterologous secondary diseases could facilitate the development of broadly effective therapeutic strategies.

overall_design: single nuclei combined ATAC/RNA sequencing of cells isolated from bronchoalveolar lavage fluid (BALF) of naive mice or mice recovered from SARS-CoV-2 (mouse-adapted strain MA10) (n = 3, pooled per condition).

first: Alexander

last: Lercher

first: Jin-Gyu

last: Cheong

first: Chenyang

last: Jiang

first: Steven

middle: Z

last: Josefowicz

first: Charles

last: Rice


title: ATAC-seq of alveolar macophages sorted from bronchoalveolar lavage fluid (BALF) isolated from naive mice or mice recovered from SARS-CoV-2 (mouse-adapted strain MA10)

summary: Pathogen encounter results in long-lasting epigenetic imprinting that shapes diseases caused by heterologous pathogens. The breadth of this innate immune memory is of particular interest in the context of respiratory pathogens with increased pandemic potential and wide-ranging impact on global health. Here, we investigated epigenetic imprinting across cell lineages in a disease relevant murine model of SARS-CoV-2 recovery. Past SARS-CoV-2 infection resulted in increased chromatin accessibility of type I interferon (IFN-I) related transcription factors and transcriptionally poised antiviral genes in alveolar macrophages. Mechanistically, viral pattern recognition and canonical IFN-I signaling were required for establishment of this innate immune memory and resulting augmented secondary antiviral responses. SARS-CoV-2-associated innate immune memory in alveolar macrophages was necessary and sufficient to ameliorate secondary disease caused by the heterologous respiratory pathogen influenza A virus. Insights into how innate immune memory shapes outcome of heterologous secondary diseases could facilitate the development of broadly effective therapeutic strategies.

overall_design: Alveolar macrophages (CD45+CD11c+SiglecF+) were sorted from BALF of naive or SARS-CoV-2 (mouse adapted strain MA10) recovered C57B/6J wild type (n = 3) and chromatin accessibility was assesses via ATAC-seq.

middle: M


title: PPARγ-dependent Remodeling of Translational Machinery in Adipose Progenitors is Impaired in Obesity

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


title: CUT&RUN of alveolar macophages sorted bronchoalveolar lavage fluid (BALF) isolated from naive mice or mice recovered from SARS-CoV-2 (mouse-adapted strain MA10)

overall_design: Alveolar macrophages (CD45+CD11c+SiglecF+) were sorted from BALF of naive or SARS-CoV-2 (mouse adapted strain MA10) recovered C57B/6J wild type (n = 2-3) and chromatin modifications (H3K4me1, H3K4me3, H3K27Ac, IgG isotype ctrl) was assesses via CUT&RUN.


title: PPARγ-dependent Remodeling of Translational Machinery in Adipose Progenitors is Impaired in Obesity [scRNA-seq]

summary: Adipose tissue plays a pivotal role in energy homeostasis and metabolic regulation. However, in obesity, the remarkable adaptability of adipose tissue becomes impaired. The underlying mechanisms behind this limited adaptability remain poorly understood. In this study, we investigate a novel layer of regulation involving translation in the adipose stromal vascular fraction, examining its response to obesity and acute PPARγ agonist treatment with rosiglitazone. Using single-cell RNA sequencing, we establish a transcriptional profile atlas of cellular remodeling from obese to lean-like states in inguinal and epididymal adipose tissue following rosiglitazone treatment. Notably, both stromal fractions exhibit a downregulation of inflammation-related transcripts and an upregulation of lipid-related metabolism and ribosomal transcripts. Adipocyte progenitor and preadipocyte populations display enhanced ex-vivo differentiation potential and upregulation in ribosome and peptide chain elongation pathways. This ribosomal remodeling is directly driven by PPARγ binding to gene promoters of ribosomal factors. Furthermore, we have characterized the translatome in the epididymal stromal fraction, highlighting a buffering response and fat-exclusive preferential translation after rosiglitazone treatment. Enhanced translation efficiency in rosiglitazone-elicited polysomes promotes the translation of transcripts containing G-rich sequences in their 5’ untranslated regions. Our findings shed light and provide a resource on how rosiglitazone remodels the adipose stromal vascular fraction, both dependent and independent of PPARγ. Importantly, we uncover translatome remodeling as a major new mechanism for maintaining translation homeostasis and preserving adipose tissue health in obesity.

overall_design: Lepob/Lepob  & C57Bl/6J mice were treated with 30mg/Kg of rosiglitazone or vehicle for 3 days. Stromal vascular fraction was isolated and used for scRNA- sequencing.


title: A microbiota-derived bile acid controls hepatic cholesterol levels in cachectic mice

city: Brussels

first: Laure

last: Bindels

address: Avenue E. Mounier, 73 bte B1.73.11

department: Metabolism and Nutrition Research Group

institute: Louvain Drug Research Institut

summary:
Our work showed decreased hepatic levels of TDCA in the C26 model of cancer cachexia. We also found gut microbiota composition alterations in cachectic C26 mice, as well as reductions in microbial 7α-dehydroxylase activity, and activity of 7α-HSDH and 12α-HSDH, which drive the production of secondary bile acids. Finally, we showed that TDCA administration induces lower hepatic levels of cholesterol in C26 mice.
Published in Thibaut et al, Gut Microbes 2025

overall_design: Hepatic RNA profiles of sham-injected mice (CT, n=6), untreated C26-transplanted mice (C26, n=8) and C26-injected mice receiving daily intraperitoneal injection of taurodeoxycholic acid (C26-TDCA, 10 mg/kg, n=8)

first: Morgane

last: Thibaut

first: Martin

last: Roumain

first: Axelle

last: Loriot

middle: B


title: HKDC1 promotes liver cancer stemness under hypoxia via stabilizing β-catenin

city: MEMPHIS

first: Li

last: Fan

state: Tennessee

address: 262 DANNY THOMAS PLACE, MAIL STOP 313, CHILI'S CARE CENTER

institute: St.jude Children's Research Hospital

summary: Hexokinases (HKs), a group of enzymes catalyzing the first step of glycolysis, have been shown to play important roles in liver metabolism and tumorigenesis. Our recent studies identified HKDC1 as a top candidate associated with liver cancer metastasis. We aimed to compare its cell type specificity with other HKs upregulated in liver cancer and investigate the molecular mechanisms underlying its involvement in liver cancer metastasis. We found that, compared to HK1 and HK2, the other two commonly upregulated HKs in liver cancer, HKDC1 was most strongly associated with the metastasis potential of tumors and organoids derived from two liver cancer mouse models we previously established. RNA in situ hybridization and single-cell RNA-seq analysis revealed that HKDC1 was specifically upregulated in malignant cells in hepatocellular carcinoma (HCC) and cholangiocarcinoma (CCA) patient tumors, whereas HK1 and HK2 were widespread across various tumor microenvironment lineages. An unbiased metabolomic profiling demonstrated that HKDC1 overexpression in HCC cells led to metabolic alterations distinct from those from HK1 and HK2 overexpression, with HKDC1 particularly impacting the tricarboxylic acid (TCA) cycle. HKDC1 was prometastatic in HCC orthotopic and tail vein injection mouse models. Molecularly, HKDC1 was induced by hypoxia and bound to glycogen synthase kinase 3β to stabilize β-catenin, leading to enhanced stemness of HCC cells. Overall, our findings underscore HKDC1 as a prometastatic HK specifically expressed in the malignant compartment of primary liver tumors, thereby providing a mechanistic basis for targeting this enzyme in advanced liver cancer.

overall_design: To investigate the function of HKDC1 in liver cancer, we established PLC/PRF/5 cell lines in which HKDC1 has been knocked out and rescued. We performed gene expression profiling analysis from RNA-seq of 3 groups including HKDC1-control, KO and KO/RES

first: Wentao

first: Liqin

last: Zhu


title: HOT3/eIF5B1 confers Kozak motif-dependent translational control of photosynthesis-associated nuclear genes for chloroplast biogenesis

city: Shanghai

first: Chenjiang

last: You

state: Shanghai

address: 2005 Songhu Rd

institute: Fudan University


title: Chronic social stress induces p16-mediated senescent cell accumulation in mice

city: Minneapolis

first: Maria

last: Razzoli

department: Integrative Biology and Physiology

institute: University of Minnesota

summary: Life stress can shorten lifespan and increase risk for aging-related diseases, but the biology underlying this phenomenon remains unclear. We assessed the effect of chronic stress on cellular senescence — a hallmark of aging. Exposure to restraint stress, a psychological non-social stress model, increased p21Cip1 exclusively in the brains of male, but not female mice, and in a p16Ink4a-independent manner. Conversely, exposure to chronic subordination stress (CSS; males only were tested) increased key senescent cell (SNC) markers in peripheral blood mononuclear cells, adipose tissue and brain, in a p16Ink4a-dependent manner. p16Ink4a-positive cells in the brain of CSS-exposed mice were primarily hippocampal and cortical neurons with evidence of DNA damage that could be reduced by p16Ink4a cell clearance. Clearance of p16Ink4a-positive cells was not sufficient to ameliorate the adverse effects of social stress on measured metrics of healthspan. Overall, our findings indicate that social stress induces an organ-specific and p16Ink4a-dependent accumulation SNCs, illuminating a fundamental way by which the social environment can contribute to aging.

overall_design: Brain sections from 4 control (ctrl), and 6 chronic social stress (CSS) exposed mice were prepared using the Nanostring GeoMx Digital Spatial Profiling with the mouse Whole Transcriptome Atlas. Matching areas of interest (AOIs) were selected from each sample in areas CA1 and CA3 of the hippocampus, the somatosensory cortex (SS), motor cortex (MC) and choroid plexus (CP).

first: Carey

middle: E

last: Lyons

first: Jean Pierre

last: Pallais

first: Seth

last: McGonigle

first: Rachel

last: Mansk

middle: W

last: Collinge

first: Matthew

last: Yousefzadeh

first: Darren

last: Baker

first: Patricia

middle: R

last: Schrank

first: Jesse

last: Williams

first: Laura

last: Niedernhofer

first: Jan

last: van Deursen

first: Alessandro

last: Bartolomucci


title: Transcriptional profiling of single cells isolated from bronchoalveolar lavage of naive mice or mice recovered from SARS-CoV-2 (mouse-adapted strain MA10) infected with influenza A/PR/8/34 virus at 7 days post infection (PR8).

first: Alexander

last: Lercher

state: NY

address: 1230 York Avenue

institute: The Rockefeller University

overall_design: single cell RNA-sequencing of cells isolated from bronchoalveolar lavage fluid (BALF) of naive mice or mice recovered from SARS-CoV-2 (mouse-adapted strain MA10) infected with influenza A/PR/8/34 virus at 7 days post infection (PR8).

first: Emma

last: DeGrace

first: Brad

last: Rosenberg

first: Charles

last: Rice