title: The G Protein-Coupled Receptor GPR31 Promotes Proinflammatory Responses in Pancreatic Islets and Macrophages II

city: Chicago

first: Sarah

last: Tersey

state: IL

department: Medicine

institute: University of Chicago

summary: In type 1 diabetes (T1D), the innate and adaptive immune systems attack and eventually destroy the insulin-secreting islet β cells. During this process, β cells activate inflammatory signaling pathways that augment the dysfunction and destruction imposed by cellular autoimmunity. The 12-lipoxygenase (12-LOX) pathway produces the pro-inflammatory eicosanoid 12-HETE, which induces oxidative and endoplasmic reticulum stress and results in diminished insulin secretion and apoptosis. The G protein-coupled receptor 31 (GPR31) has been identified as a putative receptor for 12-HETE. In this study, we generated conventional GPR31 knockout mice. To interrogate the role of GPR31 in β cells, we treated islets from wildtype and Gpr31b-/- mice with proinflammatory cytokines and subjected the islets to RNA sequencing. Differentially expressed genes in Gpr31b-/- islets included those pertaining to receptor signaling, inflammation, oxidative stress, and macrophage migration — effects that are reminiscent of 12-LOX inhibition. Bone-marrow derived macrophages from Gpr31b-/- mice had reduced macrophage migration compared to wildtype macrophages. To mimic islet and macrophage inflammation as seen in T1D, wildtype and Gpr31b-/- mice were treated with the pro-diabetic toxin streptozotocin. Compared to wildtype, Gpr31b-/- mice had improved glucose tolerance and preserved β-cell mass. These results are consistent with previously published data using 12-LOX knockout mice and suggests that GPR31 mediates the proinflammatory responses of 12-HETE in the β cell.

overall_design: To investigate the knockout of Gpr31b in macrophages, bone marrow bone marrow was harvested from WT and Gpr31b KO mice and bone marrow devired macrophages were isolated to M0 macrophages. M0 macrophages were stimulated to M1 macrophages with LPS and IFN-γ. RNA was isolated and processed for RNA-sequencing.

first: Sarah A

first: Raghavendra G

last: Mirmira


title: Multi-modal investigation reveals pathogenic features of diverse DDX3X missense mutations

status: Public on Jan 06 2025

city: Durham

first: Federica

last: Mosti

state: NC

address: 213 Research Dr

country: USA

institute: Duke University

type: Expression profiling by high throughput sequencing

summary: De novo mutations in the RNA binding protein DDX3X cause neurodevelopmental disorders including DDX3X syndrome and autism spectrum disorder. Amongst ~200 mutations identified to date, half are missense. While DDX3X loss of function is known to impair neural cell fate, how the landscape of missense mutations impacts neurodevelopment is almost entirely unknown. Here, we integrate transcriptomics, proteomics, and live imaging to demonstrate clinically diverse DDX3X missense mutations perturb neural development via distinct cellular and molecular mechanisms. Using mouse primary neural progenitors, we investigate four recurrently mutated DDX3X missense variants, from clinically severe (2) to mild (2). While clinically severe mutations impair neurogenesis, mild mutations have only a modest impact on cell fate. Moreover, expression of severe mutations leads to profound neuronal death. Using a proximity labeling screen in neural progenitors, we discover DDX3X missense variants have unique protein interactors. We observe notable overlap amongst severe mutations, suggesting common mechanisms underlying altered cell fate and survival. Transcriptomic analysis and subsequent cellular investigation highlights new pathways associated with DDX3X missense variants, including upregulated DNA Damage Response. Notably, clinically severe mutations exhibit excessive DNA damage in neurons, associated with aberrant cytoplasmic DNA:RNA hybrids and formation of stress granules. These findings highlight aberrant RNA metabolism and DNA damage in DDX3X-mediated neuronal cell death. In sum our findings reveal new mechanisms by which clinically distinct DDX3X missense mutations differentially impair neurodevelopment.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for wildtype and Ddx3x missense mutations (R326H, R376C, R488H, T532M) expressed through lentiviral delivery in primary culture from mouse E12.5 cortex. The cells were harvested for analysis 48hrs after transduction. Three biological replicates have been collected for each condition. Each replicate consist of 6 embryos cortecis pooled together. Conditions: no virus, WT, R326H, R376C, R488H, T532M

first: Debra

last: Silver


title: A mutant ASXL1-EHMT complex contributes to heterochromatin dysfunction in clonal hematopoiesis and chronic monomyelocytic leukemia [RNAseq_mouse]

city: La Jolla

first: Zhen

last: Dong

state: California

address: 9420 Athena Circle

department: Department of Signaling and Gene Expression

institute: La Jolla Institute for Immunology

summary: ASXL1 is one of the three most frequently mutated genes in age-related clonal hematopoiesis (CH), with the others being DNMT3A and TET2. CH can progress to myeloid malignancies including chronic monomyelocytic leukemia (CMML), and is also strongly associated with inflammatory cardiovascular disease and all-cause mortality in humans. DNMT3A and TET2 regulate DNA methylation and demethylation pathways respectively, and DNMT3A and TET2 loss-of-function mutations in CH reduce DNA methylation in heterochromatin, allowing de-repression of silenced elements in heterochromatin. In contrast, the mechanisms that connect mutant ASXL1 and CH are not yet fully understood. CH/CMML-associated ASXL1 mutations encode C-terminally truncated proteins that enhance the deubiquitinase activity of the ASXL-BAP1 “PR-DUB” deubiquitinase complex, which removes mono-ubiquitin from H2AK119Ub. Here we show that ASXL1 mutant proteins interact with the EHMT1-EHMT2 methyltransferase complex, which generates H3K9me1 and me2, the latter a repressive modification in constitutive heterochromatin. Compared to cells from age-matched wildtype mice, we found that expanded myeloid cells from old Asxl1tm/+ mice, a heterozygous knock-in mouse model of CH, display genome-wide decreases of H3K9me2, H3K9me3 and H2AK119Ub as well as an associated increase in expression of transposable elements (TEs) and satellite repeats. Increased TE expression was also observed in monocytes from ASXL1-mutant CMML patients compared to monocytes from healthy control individuals. Our data suggest that mutant ASXL1 proteins compromise the integrity of both constitutive and facultative heterochromatin in an age-dependent manner, by reducing the levels of H3K9me2/3 and H2AK119Ub respectively. The resulting increase in TE expression can alter the expression of nearby genes and promote the expression of inflammation-associated and interferon-inducible genes (ISGs).

overall_design: To study the functions of ASXL1 mutant proteins in clonal hematopoiesis and chronic monomyelocytic leukemia (CMML), we isolated myeloid cells (CD11b+) and hematopoietic progenitor or stem cells (LK+) from mice. We performed RNA sequencing (RNA-seq) to identify differentially expressed genes in different cell types.

first: Hugo

last: Sepulveda

first: Leo

last: Josue Arteaga

first: Anjana

last: Rao


title: Detection of DNA double-strand breaks in T-47D cells using DSBCapture-seq

city: London

first: Chi

last: Zhang

address: 15 Cotswold Road

department: Division of Cancer Therapeutics

country: United Kingdom

institute: The Institute of Cancer Research

summary: Apolipoprotein B mRNA-editing enzyme catalytic polypeptide-like 3 (APOBEC3B, A3B) is a key molecular driver inducing mutations in multiple human cancer.  A3B belongs to the APOBEC3 enzyme family, which consists seven closely related DNA deaminases that catalyse cytosine-to-uracil (C>U) editing of single-stranded DNA (ssDNA). Here, we used DSBCapture-seq to study the formation of DNA double-strand breaks (DSB) by estrogen receptor activation upon 17-estradiol (E2) induction. The effect of A3B-depletion on DSB formation was investigated using RNA interference.

overall_design: T-47D cells were divided into three groups and treated with siNT+DMSO, siNT+E2 and siA3B+E2, where siNT and siA3B and denotes non-targeting and A3B-targeting siRNA respectively. Experiments were carried out with two biological repeats.

first: Yu-jing

last: Lu

first: Jianbin

last: Yan

first: Bingjie

last: Chen

first: Konstantinos

last: Mitsopoulos

first: Marcello

last: Totoricci

first: Olivia

last: Rossanese

first: Paul

middle: A

last: Clarke

last: Workman


title: IL-23 tunes inflammatory functions of human MAIT cells

city: Paris

first: Victoire

last: Baillet

address: 25-28 rue du Dr Roux

country: France

institute: Institut Pasteur

type: Genome binding/occupancy profiling by high throughput sequencing

summary: IL-23 signaling plays a key role in the pathogenesis of chronic inflammatory and infectious diseases, yet the cellular targets and signaling pathways affected by this cytokine remain poorly understood. We show that IL-23 receptors are expressed on the large majority of human MAIT, but not of conventional T cells, suggesting that these innate-like T cells are critical mediators of IL-23 functions. In this study, we investigated the effects of IL-23 on human MAIT cell transcriptional and chromatin accessibility profiles, using RNA- and CITE-seq and ATAC-seq, respectively. Protein and transcriptional profiling at the population and single cell level demonstrates that stimulation with IL-23 or the structurally related cytokine IL-12 drives distinct functional profiles, revealing a high level of plasticity of MAIT cells. IL-23, in particular, affects key molecules and pathways related to autoimmunity and cytotoxic functions. Integrated analysis of transcriptomic and chromatin accessibility shows that AP-1 transcription factors constitute a key regulatory node of the IL-23 pathway in MAIT cells.

overall_design: PBMC were isolated from healthy donors. MAIT cells were activated for 6 days in PBMC cultures using MR1 tetramers loaded with 5-OP-RU (MR1/5-OP-RU), in the absence or presence of IL-23, or IL-12. After 6 days, MAIT cells were sorted and we performed RNA-seq, ATAC-seq, and CITE-seq. To identify the effects of IL-23 or IL-12 on MAIT cells, we compared cells activated in the presence of IL-23 or IL-12 with those stimulated with MR1/5-OP-RU only.

first: Laetitia

last: Camard

first: Tharshana

last: Stephen

first: Hanane

last: Yahia-Cherbal

first: Vincent

last: Guillemot

first: Sébastien

last: Mella

first: Hélène

last: Lopez-Maestre

first: Daniele

last: Capocefalo

first: Laura

last: Cantini

first: Claire

last: Leloup

first: Julie

last: Marsande

first: Juan

last: Sienes Bailo

first: Ambre

last: Dangien

first: Natalia

last: Pietrosemoli

first: Milena

last: Hasan

first: Huimeng

last: Wang

first: Sidonia

middle: B

last: Eckle

first: Anne

middle: M

last: Fourie

first: Carrie

last: Greving

first: Barbara

middle: J

last: Shaikh

first: Raphaelle

last: Parker

first: Daniel

last: Cua

first: Elisabetta

last: Bianchi

first: Lars

last: Rogge


title: Strand-specific DNA:RNA immunoprecipitation sequencing on T-47D breast cancer cells.

first: Chi

last: Zhang

summary: Apolipoprotein B mRNA-editing enzyme catalytic polypeptide-like 3 (APOBEC3B) is a key molecular driver inducing mutations in multiple human cancer.  A3B is a member of the APOBEC3 enzyme family, which consists seven closely related DNA deaminases that catalyse cytosine-to-uracil (C>U) editing of single-stranded DNA (ssDNA). Using a cell model that lacks base excision repair function, we sampled A3B editing sites in bulk. Analyses conducted on these A3B editing sites point to a function of A3B in editing DNA:RNA hybrid structure known as R-loops. In order to verify this result, we conducted strand-specific DNA:RNA immunoprecipitation sequencing (ssDRIP-seq; S9.6, KeraFAST, ENH001) on T-47D breast cancer cells.

overall_design: DNA:RNA immunoprecipitation (DRIP) were carried out on T-47D samples using an optimised method that minimise sampling bias (PMID: 28341774). Strand-specific libraries were constructed using input or immunoprecipitated DNA:RNA hybrids and compared.

first: Yu-jing

last: Lu

first: Jianbin

last: Yan

first: Bingjie

last: Chen

first: Konstantinos

last: Mitsopoulos

first: Marcello

last: Totoricci

first: Olivia

last: Rossanese

first: Paul

last: Clarke

last: Workman


title: R-loop editing by DNA cytosine deaminase APOBEC3B determines activities of estrogen receptor enhancers.

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

overall_design: Refer to individual Series


title: The Role of BAZ2-dependent Chromatin Remodeling in Suppressing G4 DNA Structures and Associated Genomic Instability [G4]

status: Public on Jan 05 2025

city: Boston

first: Tovah

middle: A.

last: Day

state: MA

address: 330 Huntington Avenue, Northeastern University Mugar Life Science Building, Room 134

department: Biology

institute: Northeastern University

summary: DNA G-quadruplexes (G4s) are secondary structures with significant roles in regulating genome function and stability. Dysregulation of the dynamic formation of G4s is linked to genomic instability and disease, but the underlying mechanisms are not fully understood. In this study, we conducted a screen of chromatin-modifying enzymes and identified nine potential inhibitors of G4 formation, including seven that were not previously characterized. Among these, we highlight the role of BAZ2 chromatin remodelers as key suppressors of G4 DNA and G4-related genome instability. Depletion of BAZ2 subunits led to increased G4 formation, especially at transcriptional regulatory elements. BAZ2B was found to associate with G4 loci, suggesting that it plays a direct role in suppressing G4s. While BAZ2-deficient cells exhibited modest genomic instability, treatment with the G4-stabilizing ligand BRACO19 exacerbated double-strand breaks (DSBs), highlighting its utility as a tool to study G4-dependent genome instability. DSB profiling using INDUCE-seq uncovered distinct breakage patterns around G4s, further underscoring the impact of G4s on genome integrity. Notably, we found that within G4s, G repeats were more susceptible to DSBs than loops. These results establish BAZ2 chromatin remodeling complexes as direct regulators of G4 dynamics and provide new insights into G4-dependent genome instability.

overall_design: CUT&Tag performed for G4 DNA using BG4 following different siRNA treatments as well as H3K4me3 CUT&Tag controls.

first: Adrianna

middle: L

last: Vandeuren


title: Co-regulation of plant defense  by paired Trihelix transcript factors in Arabidopsis

city: Wuhan

first: Ying

last: Wang

address: No.1 Shizishan Street

country: China

institute: Huazhong Agricultural University

summary: Transcriptomic analysis of Col-0 and AITF1 overexpressing transgenic plants was used to screen the differentially expressed genes between AITF1 overexpressing transgenic plants and Col-0. Further, transcriptomic analysis of Col-0 and aitf1-1asr3-1 plants was used to screen the differentially expressed genes between aitf1-1asr3-1 and Col-0 after flg22 treatment for indicated times, we analyzed the  ASR3/AITF1-regulated target genes to further elucidate the function of ASR3 and AITF1 in plant immune response.

overall_design: 22 RNA-seq libraries were sequenced


title: LncRNA IRF1-AS1/Gm12216 regulates the interferon response to viral challenge [ChIRP-Seq]

city: New York

first: Sofie

last: Delbare

state: NY

address: 435 East 30th Street

institute: NYU Langone

summary: Long non-coding RNAs (lncRNAs) are important regulators of the immune response. Here we study lincRNA IRF1-AS1 (human) and its mouse ortholog Gm12216. We established that IRF1-AS1/Gm12216 is involved in the antiviral response.

overall_design: Three replicate samples of THP cells were used to perform ChIRP-Seq and appropriate controls

first: Yannick

last: Cyr

first: Florencia

last: Schlamp

first: Kathryn

last: Moore

first: Coen

last: van Solingen


title: Next Generation Sequencing Facilitates Quantitative Analysis of WT and cry1 in dark and Blue light Transcriptomes

city: Shanghai

first: Yuhan

last: Hao

address: #300 Fenglin Road

institute: CAS Center for Excellence in Molecular Plant Science

summary: Analysis of differential differential gene expression regulated by Blue light.

overall_design: mRNA profiles of 10 days of WT and cry1 seedlings  grown in the dark and then exposed to continuous Blue light (40 μmol·m-2·s-1) for 1 hour were generated by deep sequencing, in triplicate, using Illumina Hiseq4000

first: Hongtao

last: Liu


title: Mapping the genomic occupancy by flag-tagged APOBEC3B protein

first: Chi

last: Zhang

summary: Apolipoprotein B mRNA-editing enzyme catalytic polypeptide-like 3 (APOBEC3B) is a key molecular driver inducing mutations in multiple human cancers.  APOBEC3B belongs to the APOBEC3 enzyme family, which consists seven closely related DNA deaminases that catalyse cytosine-to-uracil (C>U) editing of single-stranded DNA (ssDNA). In order to investigate the genomic binding sites of APOBEC3B, high through-put sequencing experiments were conducted using DNA samples acquired by standard chromatin immunoprecipitation (ChIP) and ssDNA protein immunoprecipitation (SPI) samples. Substrate preference by APOBEC3B were characterised by comparing these results.

overall_design: T-47D breast cancer cells transduced with a lentiviral vector that allows inducible expression of flag-tagged APOBEC3B protein. After induction of the lentiviral APOBEC3B cassette by doxycycline, cells were transfected with plasmid that allow transient expression of human RNase H, or treated with transfection agent as control. Treated cells were subjected to ChIP-seq and SPI-seq with anti-flag antibody (M2, Sigma, F1804), which detect APOBEC3B-bound dsDNA or ssDNA respectively.

first: Yu-jing

last: Lu

last: Yan


title: Metabolic Deficiencies Underlie Reduced Plasmacytoid Dendritic Cell IFN-I Production following Viral Infection [RNA-Seq]

first: Trever

last: Greene

state: CA

address: 9500 Gilman Dr,

institute: University of California San Diego

summary: T ype I Inter ferons (IFN-I) are central to host protection against viral infections, with plasmacytoid de ndritic cells (pDC) being the most significant source, yet pDCs lose their exceptional IFN-I production capacity following an initial burst of IFN-I, resulting in susceptibility to secondary infections.  The underlying mechanisms of these dynamics are not well understood. Here we find that viral infection reduces the capacity of pDCs to engage both oxidative and glycolytic metabolism. Mechanistically, we identify lactate dehydrogenase B (LDHB) as a positive regulator of pDC IFN-I production in mice and humans; meanwhile, LDHB deficiency is associated with suppressed IFN-I production, pDC metabolic capacity, and viral control following infection. In addition, preservation of LDHB expression is sufficient to partially retain the function of otherwise exhausted pDCs, both in vitro and in vivo. Furthermore, restoring LDHB in vivo in pDCs from infected mice increases IFNAR-dependent, infection-associated pathology. Our work identifies a mechanism for balancing immunity and pathology during viral infections, while also providing insight into the highly preserved infection-driven pDC inhibition.

overall_design: C57Bl/6 Mice were infected with LCMV or left uninfected. At the indicated timepoints (24 hours, 8 days, 30 days) post infection, spleens were harvested, and pDCs sorted .


title: Metabolic Deficiencies Underlie Reduced Plasmacytoid Dendritic Cell IFN-I Production following Viral Infection

summary: Type I Inter ferons (IFN-I) are central to host protection against viral infections, with plasmacytoid dendritic cells (pDC) being the most significant source, yet pDCs lose their exceptional IFN-I production capacity following an initial burst of IFN-I, resulting in susceptibility to secondary infections.  The underlying mechanisms of these dynamics are not well understood. Here we find that viral infection reduces the capacity of pDCs to engage both oxidative and glycolytic metabolism. Mechanistically, we identify lactate dehydrogenase B (LDHB) as a positive regulator of pDC IFN-I production in mice and humans; meanwhile, LDHB deficiency is associated with suppressed IFN-I production, pDC metabolic capacity, and viral control following infection. In addition, preservation of LDHB expression is sufficient to partially retain the function of otherwise exhausted pDCs, both in vitro and in vivo. Furthermore, restoring LDHB in vivo in pDCs from infected mice increases IFNAR-dependent, infection-associated pathology. Our work identifies a mechanism for balancing immunity and pathology during viral infections, while also providing insight into the highly preserved infection-driven pDC inhibition.

overall_design: Mixed bone marrow chimeras were infected with LCMV Cl13, and WT or TLR7ko pDC were isolated from the spleens of these mice at day 8 post infection prior to RNA extraction. 2 independent biological replicates were performed.


title: Multi-Omic Single-Cell Dissection of Leukemic T-Cell Lymphoma Following CAR T-Cell Therapy [Spatial Transcriptomics]

city: Cologne

first: Till

last: Braun

address: Kerpener Strasse 62

department: Clinic I for Internal Medicine

country: Germany

institute: University Hospital of Cologne

summary: A 63-year-old male who received ciltacabtagene autoleucel (cilta-cel) CAR-T cells and the GPRC5DxCD3 bispecific talquetamab for early relapse of his multiple myeloma (MM) developed a leukemic peripheral T-cell lymphoma (PTCL) with cutaneous and intestinal involvement. Longitudinal single-cell RNA and T-cell receptor sequencing of peripheral blood and bone marrow revealed two hyperexpanded CAR-carrying, exhausted effector-memory T-cell clones with marked immunophenotypic as well as transcriptional alterations and different susceptibilities towards treatment with dexamethasone. Spatial transcriptomes of skin lesions confirmed the aberrant CAR-expressing T cells. Whole genome sequencing revealed three distinct integration sites, within the introns of ZGPAT, KPNA4, and polycomb-associated non-coding RNAs. Pre/post-CAR-T whole-genome analyses implicated clonal outgrowth of a TET2-mutated precursor propelled by additional sub-clone specific LOH and other secondary mechanisms

overall_design: Two tissue sections of a formalin-fixed paraffin-embedded (FFPE) skin sample, from a patient treated with CAR-T cells, of the left dorsal region. Three custom probe sets were developed to detect CAR-positive (cilta-cel) spots. To demonstrate the specificity of the cilta-cel custom probes, one slide with a skin sample was subjected to the Visium Spatial Protocol WITHOUT custom probes, another slide with a skin sample was subjected to the Visium Spatial Protocol WITH custom probes and an extramedullary disease sample (sample excision from the lumbar vertebral body) from a patient not treated with CAR-T cells was subjected to the Visium Spatial Protocol WITH custom probes.

first: Michael

last: Rade

first: Maximilian

last: Merz

first: Hanna

last: Klepzig

first: Florian

last: Große

first: David

last: Fandrei

first: Nhu-Nguyen

last: Pham

first: Markus

last: Kreuz

first: Christina

middle: K

last: Kuhn

last: Kuschel

first: Dennis

last: Löffler

first: Jörn

last: Meinel

first: Eva

last: Heger

first: Viola

last: Schweinsberg

first: Natali

last: Pflug

first: Uwe

last: Platzbecker

last: Hallek

first: Udo

last: Holtick

first: Ulrike

last: Köhl

first: Christof

last: Scheid

first: Kristin

last: Reiche

first: Marco

last: Herling

first: Tim

last: Richardson


title: Chronic cigarette smoke exposure masks pathological features of Helicobacter pylori infection while promoting tumor initiation

city: RTP

first: Xiaojiang

last: Xu

state: North Carolina

address: 111 T. W. Alexander Dr.,

institute: NIEHS

summary: Gastric cancer is the fifth most common cancer and the fifth leading cause of cancer deaths worldwide. Chronic infection by the bacterium Helicobacter pylori is the most prominent gastric cancer risk factor, but only 1-3% of infected individuals will develop gastric cancer. Cigarette smoking is another independent gastric cancer risk factor, and H. pylori-infected smokers are at a 2-11-fold increased risk of gastric cancer development, but the direct impacts of cigarette smoke on H. pylori pathogenesis remain unknown. In this study, male C57BL/6 mice were infected with H. pylori and began smoking within one week of infection. The mice were exposed to cigarette smoke (CS) five days/week for 8 weeks. CS exposure had no notable impact on gross gastric morphology or inflammatory status compared to filtered-air (FA) exposed controls in mock-infected mice. However, CS exposure significantly blunted H. pylori-induced gastric inflammatory responses, reducing gastric atrophy and pyloric metaplasia development. Despite blunting these classic pathological features of H. pylori infection, CS exposures increased DNA damage within the gastric epithelial cells and accelerated H. pylori-induced dysplasia onset in the INS-GAS gastric cancer model. These data suggest that cigarette smoking may clinically silence classic clinical symptoms of H. pylori infection but enhance the accumulation of mutations and accelerate gastric cancer initiation.

overall_design: Mice were infected with H. pylori and exposed to filtered air or  cigarette smoke 5 days/week for 8 weeks.

first: Jonathson

last: Busada


title: Mangrove endophytes enhance yield and convey salt tolerance to Nipponbare rice

city: Jeddah

first: Sabiha

last: Parween

address: Thuwal

department: Hirt lab

country: Saudi Arabia

institute: KAUST

summary: Global climate change increasingly challenges agriculture with flooding and salinity. Among strategies to enhance crop resilience to these stresses, we showed two mangrove strains that can enhance flooding and salinity tolerance in rice plants. Two strains massively enhanced the growth and yield of Oryza sativa cv. Nipponbare under hydroponic growth conditions with and without salt treatment. The bacteria-induced transcriptome changes in O. sativa roots related to ABA-signaling with suberin deposition in root tissues explain the altered responses of colonized rice plants to hypoxic and saline stress conditions. While enhancing yield and grain quality, bacterially colonized rice plants also show much earlier flowering, thereby massively shortening the life cycle of rice plants and opening the possibility for an additional harvest per year. These results show that microbes can be a powerful tool for enhancing the yield and resilience of rice to hypoxic and saline stress conditions.

overall_design: Oryza sativa plants were grown in growth chambers at 27/25°C (day/night) with a 12:12 h light/dark cycle, the light intensity of 200 µmol photons m−2 s−1, and 70% humidity. For RNASeq analysis, the 5-days-old seedlings were transferred to buckets containing Hoagland solution supplemented with 0 or 100 mM NaCl. After 24h, the shoots and roots were collected separately and flash-frozen in liquid N2 before RNA extraction. Total RNA was extracted from roots and shoots of 6-day-old mock and AK164, AK171 colonized plants using Direct-zol RNA Miniprep Plus Kit following the manufacturer’s instructions (ZYMO RESEARCH; USA). RNA libraries were prepared for sequencing using ....protocol. RNA-seq was carried out on three independent biological replicates.

first: Amal

last: Khalaf Alghamdi

first: Anamika

last: Ravat

first: Arun

middle: P

last: Nagarajan

first: Maged

last: Saad

first: Heribert

last: Hirt


title: TSPYL5 induces cancer stem cells stemness by G3BP1/RanBP2-mediated p53 modification in Neuroblastoma

city: Chengdu

first: Wang

last: Zhaokun

address: Chuanda Road Section 2

institute: Sichuan university

summary: TSPYL5 induces cancer stem cells stemness by G3BP1/RanBP2-mediated p53 sumoylation modification in Neuroblastoma

overall_design: Comparative gene expression profiling analysis of RNA-seq data for SK-N-SH and SH-SY5Y cells and their KD derivatives (shTSPYL5).

first: Zhaokun

last: Wang

first: Yangwei

last: Zhang

first: Yunqiang

last: Liu

first: Yuan

last: Yang


title: CD9-dependent extracellular vesicles mediate the propagation of ferroptosis

city: Hangzhou

first: Tianyu

last: Guo

state: Zhejiang

address: 866 Yuhangtang Road

institute: Zhejiang University

summary: Ferroptosis is a form of regulated cell death that is mediated by accumulation of lipid peroxidation. Emerging evidences report dispersion of ferroptosis between neighboring cells. However, the molecular mechanism underlying the propagation of ferroptosis remains unclear. Here we provide evidences that ferroptotic cells elicit cellular projections that produce massive extracellular vesicles (EVs). Proteomic analysis reveals that these EVs are CD9 positive and contain major components of mitogen-activated protein kinase (MAPK) signaling. MAP2K1/2 that is transferred by EVs increases the sensitivity to ferroptosis of target cells. We further show that a MAP2K-interacting transcription factor, peroxisome-proliferator activator receptor gamma (PPARG), activates various components of ferroptosis defense systems. Moreover, perturbation of EV-based transport protects adjacent tissue from ferroptosis in vivo. Our study reveals an EV-dependent transport between ferroptotic cells and surrounding neighbors and establishes that MAPK-responsive PPARG is essential for ferroptosis defense in recipient cells.

overall_design: U-2 OS cells expressing PPARG-EGFP were used CUT&Tag-sequencing to analyze PPARG regulating genes.  Antibody against EGFP was used and three replicates were analysed.

first: Huanle

last: Li

first: Pengzhen

last: Dong

first: Dandan

first: Yu

last: Ran

first: Yuying

first: Tianheng

last: Xiao

first: Tianfeng

last: Lu

first: Honggang

last: Wu

first: Fudi

first: Bing

first: Hai

last: Huang


title: Mapping and quantification of single-strand DNA bound by flag-tagged APOBEC3B

first: Chi

summary: Apolipoprotein B mRNA-editing enzyme catalytic polypeptide-like 3 (APOBEC3B, A3B) is a key molecular driver inducing mutations in multiple human cancer.  A3B belongs to the APOBEC3 enzyme family, which consists seven closely related DNA deaminases that catalyse cytosine-to-uracil (C>U) editing of single-stranded DNA (ssDNA). Here, we adopted single-strand DNA protein-immunoprecipitation sequencing (SPI-seq) to capture the ssDNA bound by flag-tagged A3B protein. Quantification on sequence reads was carried out to investigate the effect of 17-beta estradiol (E2) on A3B binding to DNA.

overall_design: T-47D breast cancer cells transduced with a lentiviral vector that allows inducible expression of flag-tagged APOBEC3B protein. After induction of the lentiviral APOBEC3B cassette by doxycycline, SPI-seq experiments were carried out using T-47D cells treated with DMSO or E2 (anti-flag antibody (M2, Sigma, F1804)) Quantification was carried out using n = 2 biological repeats to investigate the effect of E2 stimulation on A3B binding.

first: Yu-jing

first: Jianbin

last: Yan

first: Bingjie

last: Chen

first: Konstantinos

last: Mitsopoulos

first: Marcello

last: Totoricci

first: Olivia

last: Rossanese

first: Paul

last: Clarke

last: Workman


title: Aminoglycoside tolerance in Vibrio cholerae engages translational reprogramming associated to queuosine tRNA modification

city: Orsay

first: Olivier

last: Namy

address: Rue Gregor Mendel, bat 400

institute: CNRS, Université Paris-Sud

summary: Tgt is the enzyme modifying the guanine (G) in tRNAs with GUN anticodon to queuosine (Q). tgt is required for optimal growth of Vibrio cholerae in the presence of sub-lethal aminoglycoside concentrations. We further explored here the role of the Q in the efficiency of codon decoding upon tobramycin exposure. We characterized its impact on the overall bacterial proteome, and elucidated the molecular mechanisms underlying the effects of Q modification in antibiotic translational stress response. Using molecular reporters, we showed that Q impacts the efficiency of decoding at tyrosine TAT and TAC codons. Proteomics analyses revealed that the anti-SoxR factor RsxA is better translated in the absence of tgt. RsxA displays a codon bias towards tyrosine TAT and overabundance of RsxA leads to decreased expression of genes belonging to SoxR oxidative stress regulon. We also identified conditions that regulate tgt expression. We propose that regulation of Q modification in response to environmental cues leads to translational reprogramming of genes bearing a biased tyrosine codon usage. In silico analysis further identified candidate genes possibly subject to such translational regulation, among which DNA repair factors. Such transcripts, fitting the definition of modification tunable transcripts, are plausibly central in the bacterial response to antibiotics.

overall_design: RiboSeq of three replicates of two Vibrio Cholerae strains (one WT and  the other deleted for the tgt gene) in presence or absence of tobramycin, to identify which genes are impacted at the translational level by the antibiotic.

first: Louna

last: Fruchard

first: Anamaria

last: Babosan

first: Andre

last: Carvalho

first: Manon

last: Lang

first: Blaise

first: Magalie

last: Duchateau

first: Quentin

last: Giai-Gianetto

first: Mariette

last: Matondo

first: Frédéric

last: Bonhomme

first: Isabelle

last: Hatin

first: Hugo

last: Arbes

first: Céline

last: Fabret

first: Virgine

last: Marchand

first: Iouri

last: Motorine

first: Valérie

last: de Crécy-Lagard

first: Didier

last: Mazel

first: Zeynep

last: Baharoglu


title: Whole genome sequencing of T-47D cell colonies overexpressed with APOBEC3B and hUGI

first: Chi

last: Zhang

summary: Apolipoprotein B mRNA-editing enzyme catalytic polypeptide-like 3 (APOBEC3B, A3B) is a key molecular driver inducing mutations in multiple human cancer.  A3B belongs to the APOBEC3 enzyme family, which consists seven closely related DNA deaminases that catalyse cytosine-to-uracil (C>U) editing of single-stranded DNA (ssDNA). Here, we used a lentiviral inducible system to overexpress A3B and hUGI in T-47D cells. This system will allow augmented genomic editing by A3B, while preserving the C>U editing sites from DNA repair by inhibiting base excision repair mechanism.

overall_design: T-47D cell colonies were separated after induced with wild-type or inactive mutant A3B expression for 120 hours, which were subjected to whole genome sequencing and mutation calling.

first: Yu-jing

last: Lu

first: Jianbin

last: Yan

first: Bingjie

last: Chen

first: Konstantinos

last: Mitsopoulos

first: Marcello

last: Totoricci

first: Olivia

last: Rossanese

first: Paul

last: Clarke

last: Workman


title: Quantified strand-specific DNA:RNA immunoprecipitation sequencing on T-47D breast cancer cells.

summary: Apolipoprotein B mRNA-editing enzyme catalytic polypeptide-like 3 (APOBEC3B) is a key molecular driver inducing mutations in multiple human cancer.  A3B belongs to the APOBEC3 enzyme family, which consists seven closely related DNA deaminases that catalyse cytosine-to-uracil (C>U) editing of single-stranded DNA (ssDNA). Using a cell model that lacks base excision repair function, we sampled A3B editing sites in bulk. Analyses conducted on these A3B editing sites point to a function of A3B in editing DNA:RNA hybrid structure known as R-loops. In order to verify this result, we conducted strand-specific DNA:RNA immunoprecipitation sequencing (ssDRIP-seq; S9.6, KeraFAST, ENH001) on T-47D breast cancer cells.

overall_design: T-47D cells were divided into four sample groups and treated with siNT+DMSO, siNT+E2, siA3B+DMSO and siA3B+E2, where siNT and sA3B denote non-targeting and A3B-targeting siRNA respectively and E2 denotes 17-beta estradiol (E2). For each of the treatment group, three biological replicates were conducted, resulting a total of 12 samples. The effect of E2 and A3B knockdown on R-loop formation was investigated with quantified ssDRIP-seq.


title: Salivary miRNA profiles in relationship with occupational stress

status: Public on Jan 04 2025

city: Orbassano

first: Giulio

last: Ferrero

state: TO

address: Regione Gonzole, 10

department: Dept. Clinical and Biological Sciences

country: Italy

institute: University of Turin

type: Non-coding RNA profiling by high throughput sequencing

summary: Work-related stress in police officers has been recognized as a risk factor for mental and physical illness, compromising work performance and public safety. While genetic, individual and environmental factors may contribute to stress, basic pathophysiological mechanisms are still not completely understood. Previous studies have shown the potential of microRNAs (miRNAs) as biomarkers and/or mediator for stress responses. In this study, we aimed at identifying possible dysregulation of salivary miRNAs associated with work-related stress in police officers. The study was conducted in a cohort composed of police officers in service at the mobile unit of Genoa in 2009, whose response to occupational stress has been continuously monitored since 2009 using the Demand-Control and the Effort-Reward Imbalance models. 113 male police officers in service in 2022 were recruited and divided in three groups according to their response to stress (low, intermediate and high stress response). miRNAs were extracted from saliva samples and analyzed by small RNA sequencing followed by an integrated bioinformatics/biostatistics analysis. The high stress response group was characterized by 18 differentially expressed miRNAs compared with the low stress response group, with four miRNAs (miR-10400-5p, miR-1290, miR-6074-5p, and miR-9902) at higher levels and fourteen miRNAs (miR-203a-3p, miR-7-5p, miR-143-3p, miR-181a-5p, miR-142-5p, miR-223-5p, let-7f-5p, let-7g-5p, miR-26b-5p, miR-27a-3p, miR-30e-5p, miR-26a-5p, miR-142-3p, and miR-21-5p) at lower levels. We also discovered sets of miRNAs that exhibited dynamic expression changes in response to intermediate and low stress. Functional enrichment analysis identified processes related to apoptosis, cellular response to stress, negative regulation of gene expression, responses to abiotic stimuli, positive regulation of catabolism as commonly regulated by the modulated miRNAs. Work-related stress response was associated with specific salivary miRNA expression patterns, with a general decrease of miRNA levels in the highly distressed police officers. These results suggest salivary miRNA profiling as a non-invasive and useful biomarker of stress and further characterize the link between chronic psychosocial stress and cellular and behavioral response mechanisms.

overall_design: Small RNA-Seq salivary miRNA profiles of 113 subjects with different levels of occupational stress

first: Segio

last: Garbarino

first: Nicola

last: Magnavita

first: Barbara

last: Pardini

first: Sonia

last: Tarallo

first: Fabrizio

last: Ciprani

first: Egeria

last: Scoditti

first: Alessio

last: Naccarati


title: The SinR•SlrR heteromer internally attenuates a long transcript of flagellar genes in B. subtilis [RNA-seq]

city: Bloomington

first: Ayushi

last: Mishra

state: Indiana

address: Biology Building, BB432, 1001 E 3rd St

institute: Indiana University

summary: Bacillus subtilis differentiates into two cell types during growth, dictated by the activity of the alternative sigma factor SigD. The frequency of SigD-ON motile cells is increased when the heteromeric transcription factor SwrA•DegU activates the promoter of the long fla/che operon that encodes SigD. Conversely, the frequency of SigD-ON cells is decreased by the heteromeric transcription factor SinR•SlrR but the mechanism and location of inhibition is poorly understood. Using ChIP-Seq analysis, we determine the binding sites of the SinR•SlrR heteromer, including five sites within the fla/che operon. The first two sites were found to be both necessary and sufficient to attenuate transcript abundance within the operon upstream of the gene that encodes SigD. We conclude that the SinR•SlrR heteromer inhibits SigD accumulation by binding within the operon by causing RNA polymerase to pause and prematurely terminate. Thus, the development of motile cells, and the motility-to-biofilm transition, depends on the expression of a long operon governed by two opposing heteromeric transcription factors that operate at two different stages of the transcription cycle.

sample_organism: Bacillus subtilis subsp. subtilis NCIB 3610 = ATCC 6051 = DSM 10

overall_design: RNA extraction and sequencing was performed on wild type and mutant cells of Bacillus subtilis 3610 grown in LB at 37C. Before sequencing, ribosomal RNA was depleted.


title: The SinR•SlrR heteromer internally attenuates a long transcript of flagellar genes in B. subtilis [ChIP-seq]

platform_organism: Bacillus subtilis subsp. subtilis NCIB 3610 = ATCC 6051 = DSM 10

overall_design: ChIP-Seq experiments were performed on wildtype and other mutants using anti-SinR antibody. All strains were grown in LB and crosslinked with 3% formaldehyde.


title: Cadmium-induced oxidative stress, histopathology, and transcriptome changes in the hepatopancreas of freshwater crayfish (Procambarus clarkii)

city: Harbin

first: Yu

last: Zhang

address: Huanghe Street No.73

institute: Harbin Institute of Technology

summary: The transcriptome results, including siginificantly enriched GO terms, KEGG pathways, and DEGs involved in the response to Cd, provided further evidence indicating the effects of Cd on the P. clarkii hepatopancreas at the molecular level.

overall_design: Crayfish (P. clarkii) were exposed to 2, 5, and 10 mg/L Cd for 72h

first: Kai

last: Sun


title: The Role of BAZ2-dependent Chromatin Remodeling in Suppressing G4 DNA [HA] Structures and Associated Genomic Instability

first: Tovah

last: Day

summary: DNA G-quadruplexes (G4s) are secondary structures with significant roles in regulating genome function and stability. Dysregulation of the dynamic formation of G4s is linked to genomic instability and disease, but the underlying mechanisms are not fully understood. In this study, we conducted a screen of chromatin-modifying enzymes and identified nine potential inhibitors of G4 formation, including seven that were not previously characterized. Among these, we highlight the role of BAZ2 chromatin remodelers as key suppressors of G4 DNA and G4-related genome instability. Depletion of BAZ2 subunits led to increased G4 formation, especially at transcriptional regulatory elements. BAZ2B was found to associate with G4 loci, suggesting that it plays a direct role in suppressing G4s. While BAZ2-deficient cells exhibited modest genomic instability, treatment with the G4-stabilizing ligand BRACO19 exacerbated double-strand breaks (DSBs), highlighting its utility as a tool to study G4-dependent genome instability. DSB profiling using INDUCE-seq uncovered distinct breakage patterns around G4s, further underscoring the impact of G4s on genome integrity. Notably, we found that within G4s, G repeats were more susceptible to DSBs than loops. These results establish BAZ2 chromatin remodeling complexes as direct regulators of G4 dynamics and provide new insights into G4-dependent genome instability.

overall_design: CUT&Tag performed for endogenously HA tagged BAZ2B using an HA antibody as well as H3K4me3 CUT&Tag controls.

first: Adrianna

last: Vandeuren


title: Small cell carcinoma of the ovary, hypercalcemic type (SCCOHT) cell lines treated with triptolide for 6 or 16h -  H3K27ac CUT&RUN

city: Madison

first: Jessica

middle: Diane

last: Lang

state: Wisconsin

address: 1111 HIGHLAND AVE WIMR 2766

department: Pathology and Laboratory Medicine

institute: University of Wisconsin-Madison

summary: Small cell carcinoma of the ovary-hypercalcemic type (SCCOHT) is a rare form of ovarian cancer affecting young women and girls. Survival rates remain poor despite aggressive treatment of these patients with high-dose chemotherapy and radiation. SCCOHT is driven by loss of both SWI/SNF ATPases SMARCA4 and SMARCA2. Loss of SWI/SNF complex activity alters chromatin state, particularly at enhancers, which is believed to be crucial for oncogenesis. Super-enhancers are a distinct subset of enhancer clusters frequently associated with oncogenes in cancer. Here we discovered key distinctions between SWI/SNF binding following SMARCA4 restoration at enhancer vs. super-enhancer sites and characterized putative oncogene expression driven by super-enhancer activity. We found high sensitivity of SCCOHT cell lines to triptolide, a small molecule derived from the naturally occurring "thunder god vine" (Tripterygium wilfordii) that targets the XPB subunit of the transcription factor II H (TFIIH) complex, found at super-enhancers. Triptolide inhibits expression of many super-enhancer associated genes, and in particular, oncogenes. Notably, SALL4 expression, which is high in SCCOHT relative to other ovarian cancers and normal cell types, is significantly decreased following short triptolide treatment. In SCCOHT patient-derived xenograft models, triptolide and its prodrug derivative minnelide are particularly effective in inhibiting tumor growth. These results demonstrate the key oncogenic role of super-enhancer activity following epigenetic dysfunction in SCCOHT, which can be effectively targeted through inhibition of its functional components, such as TFIIH inhibition with triptolide.

overall_design: H3K27ac & IgG CUT&RUN on BIN67, and COV434 SCCOHT cell lines treated with triptolide

middle: D

first: Jeffrey

last: Trent


title: Small cell carcinoma of the ovary-hypercalcemic type (SCCOHT) tumor gene expression

summary: Small cell carcinoma of the ovary-hypercalcemic type (SCCOHT) is a rare form of ovarian cancer that has not previously been characterized at the gene expression level. The goal of this study was to generate gene expression profiles from SCCOHT tumor samples by RNA-Seq.

overall_design:
RNA-Seq on SCCOHT tumors.

**The raw data require controlled access and have been deposited in dbGaP under accession number phs001528.v2.p1**

first: William

last: Hendricks

first: Winnie

last: Liang

first: Patrick

last: Pirrotte

first: Elizabeth

last: Raupach


title: Wild type and tumor-derived mutant p53 share a non-canonical mode of gene occupancy [RNA-seq]

city: WEST HOLLYWOOD

first: Moray

last: Campbell

address: 8687 Melrose Avenue, Office B227.5G

institute: Cedars-Sinai Medical Center

summary: To directly examine the interplay between mutant p53 or Mdm2 and wild type p53 in gene occupancy and expression, an integrated RNA-seq and ChIP-seq analysis was performed in vivo using isogenically matched mouse strains. Response to radiation was used as an endpoint to place findings in a biologically relevant context. Unexpectedly, mutant p53 and Mdm2 only inhibit a subset of wild type p53 gene expression. In contrast to a dominant-negative or inhibitory role, the presence of either mutant p53 or Mdm2 actually enhances the occupancy of wild type p53 on many canonical targets. The C-terminal 19 amino acids of wild type p53 suppresses the p53 response allowing for survival at sublethal doses of radiation. Further, the mutant 172H p53 is shown to occupy genes and regulate their expression via a non-canonical means that is shared with wild type p53. This results in the heterozygous 172H/+ genotype having an expanded transcriptome compared to wild type p53+/+.

overall_design: Expression profiling by high-throughput sequencing of bone marrow in wt p53 mice and mice with a deletion of the C-terminus of p53. Bone marrow from the indicated genotype mice was treated as indicated (9.5 or 6 Gy) from 8-week-old mice (8w). Tissue were immediately placed in RNAlater during dissection and stored at 4oC until RNA is extracted using RNeasy Mini Kit (Qiagen) and following manufacturer’s instructions. The isolated RNA is quantitated and either stored at -80o in preparation for RNA-seq. Sequencing libraries prepared with the TruSeq Stranded Total RNA kit (Illumina Inc), from 1ug total RNA.

first: M

first: J

last: Manfredi


title: A developmentally-inspired hypoxia condition promotes kidney organoids differentiation from human pluripotent stem cells

city: Ulsan

first: Tae-Eun

last: Park

address: 50, UNIST-gil, Eonyang-eup, Ulju-gun

department: Biomedical Engineering

country: South Korea

institute: Ulsan National Institute of Science and Technology (UNIST)

summary: Kidney organoids derived from human pluripotent stem cells (hPSCs) lack a contiguous network of collecting ducts, which limits their utility in modeling kidney development and disease. Here, we report the generation of kidney organoids containing ureteric bud (UB)-derived collecting ducts connected to metanephric mesenchyme (MM)-derived nephrons using a developmentally-inspired hypoxic differentiation condition. Hypoxia promotes a reiterative process of branching morphogenesis and nephron induction through reciprocal interactions between co-induced MM and UB, which lead to a higher-order kidney organogenesis in vitro. The resulting kidney organoids demonstrate greater maturity, as indicated by higher levels of functional markers and a more realistic micro-anatomy. Additionally, these hypoxic-enhanced kidney organoids show a great potential as in vitro models for renal cystic diseases, as they efficiently generate cystic formations and display high sensitivity to drugs. This hypoxia approach may open new avenues for an enhanced understanding of kidney development and diseases.

overall_design: Single-cell RNA sequencing was conducted to profile the cellular composition of kidney organoids differentiated under normoxic and hypoxic conditions.

first: Hyeonji

last: Lim

first: Dong

middle: S

last: Kim


title: Small cell carcinoma of the ovary, hypercalcemic type (SCCOHT) cell lines treated with triptolide for 6h -  RNA-seq

overall_design: RNAseq on CUT&RUN on BIN67, and COV434 SCCOHT cell lines treated with triptolide


title: Advancing the genetic engineering toolbox by combining AsCas12a knock-in mice with ultra-compact screening

status: Public on Jan 03 2025

city: Parkville

first: John

middle: E

last: La Marca

state: VIC

address: 1G Royal Parade

department: Blood Cells and Blood Cancer

country: Australia

institute: Walter and Eliza Hall Institute

summary: Cas12a is a next-generation gene-editing tool that enables multiplexed gene targeting. Here, we present a mouse model that constitutively expresses enhanced Acidaminococcus sp. Cas12a (enAsCas12a) linked to an mCherry fluorescent reporter. We demonstrate efficient single and multiplexed gene-editing in vitro, using primary and transformed cells from enAsCas12a mice. We further demonstrate successful in vivo gene-editing, using normal and cancer-prone enAsCas12a stem cells to reconstitute the haematopoietic system of wild-type mice. We also present compact, genome-wide Cas12a knockout libraries, with four crRNAs per gene encoded across one (Scherzo) or two (Menuetto) vectors, and demonstrate the utility of these libraries across methodologies: in vitro enrichment and drop-out screening in lymphoma cells and immortalised fibroblasts, respectively, and in vivo screens to identify lymphoma-driving events. Finally, we demonstrate CRISPR multiplexing via simultaneous gene knockout (via Cas12a) and activation (via dCas9-SAM) using primary T cells and fibroblasts. Our enAsCas12a mouse and accompanying crRNA libraries enhance genome engineering capabilities and complement current CRISPR technologies.

overall_design: NGS of mouse cells containing CRISPR/Cas12 libraries (Menuetto/dual and Scherzo/quad). Sequences are of the sgRNA plasmids. Three screens were performed: 1) enrichment screen in mouse EuMyc lymphoma cells using S63845 and Nutlin-3a as selection pressures, 2) drop out screen in immortalised mouse dermal fibroblasts (MDFs), and 3) in vivo enrichment screen for drivers of lymphomagenesis in mice.

first: Sarah

middle: T

last: Diepstraten

first: Yexuan

last: Deng

first: Wei

last: Jin

first: Marco

last: Herold

first: Jean-Philippe

last: Fortin

first: Benjamin

last: Haley


title: Bovine turbinate cells as an in vitro model for the study of influenza D virus

city: Nottingham

first: Meshach

last: Maina

address: University Of Nottingham, Sutton Bonington Campus

department: One Virology, Wolfson Center for Global Virus Research, Vet. School

institute: University of Nottingham

summary: Although it causes only mild respiratory disease, influenza D virus (IDV) is implicated in bovine respiratory disease complex (BRDC), which is a major cause of economic concern for cattle producers globally. The virus replicates in the upper and lower respiratory tract with virus titre highest in nasal turbinates. We therefore hypothesised that the bovine turbinate (BT) cell line could provide a useful in vitro model to study viral–host interactions. Host gene expression was determined using RNA-Seq. Significantly differentially expressed genes with respect to mock-infected cells were identified using DESeq2 and KEGG pathway analysis used to determine enriched pathways. Psuedotyped viruses were used to show the involvement of intrinsic innate immune responses including IFN-inducible transmembrane (IFITM) proteins in restricting the entry of IAV, ICV and IDV. Localization of bovine IFITM3 was determined using immunofluorescence assay. Pull-down and immunoprecipitation assays were used to detect binding of IDV NS1 to p85β subunit of the PI3K pathway. RNA-Seq demonstrated the involvement of genes related to type I or type III interferon responses in IDV infection of BT cells. Intrinsic innate immune responses including IFN-inducible transmembrane (IFITM) proteins restricted the entry of IAV, ICV and IDV pseudotyped viruses. Determination of the subcellular localization of bovine IFITM3 showed that it has an intracellular cytoplasmic distribution. KEGG pathway analysis of the transcriptomic data showed activation of the PI3K/Akt pathway. This led to the hypothesis that IDV activates the pathway in a similar way to influenza A virus (IAV), by non-structural NS1 protein binding the PI3K p85β subunit. However, pull-down and immunoprecipitation assays did not detect binding of IDV NS1 to p85β; this may be due to amino acid differences in the tyrosine-phosphorylated YXXXM motif that is important in the binding of the IAV NS1 to the p85β subunit. This suggests a different mechanism in preventing apoptosis after IDV infection. In conclusion, this study has shown the efficient replication of IDV in bovine turbinate cells. The study has also shown the involvement of type I interferon responses in immune response to IDV infection of cells of the upper respiratory tract of the cow. There is the involvement of different signalling pathways including the FoXO and PI3K/Akt pathway in IDV infection. Bovine IFITM3 is localised intracellularly and is involved in the restriction of IDV entry.

overall_design: To determine the transcriptomic profile of bovine turbinate cells following influenza D virus infection (IDV), we extracted total RNA from 3 biological replicates of mock infected BT cells (time 0, no virus control) and IDV infected BT cells at 24 and 48 hours post infection. Host gene expression was determined using data from RNA-Seq.

first: Toshana

last: Foster

first: Ken

last: Mellits

first: Mariette

last: Ducatez

first: Stephen

last: Dunham

first: Janet

last: Daly


title: SIRPa ablation overcomes exhaustion and enhances the antibody- and CAR-mediated antitumor activities of iPSC-derived CAR Macrophages

first: Igor

last: Slukvin

address: 1220 Capital Ct

department: Wisconsin National Primate Research Center

summary: SIRPa-CD47 “don’t eat me” checkpoint axis plays a critical role in defining anti-tumor activities of macrophages within the tumor microenvironment. However, targeting this axis with anti-CD47 antibodies to improve anti-tumor responses in clinical trials has proven challenging. Here, we demonstrated that iPSC-derived macrophages (iMacs) with ablated SIRPa yield a superior antitumor effect in conjunction with cancer-targeted antibodies (Ab) or chimeric antigen receptor (CAR) against a variety of CD47-expressing tumors in vitro. Moreover, SIRPA-KO protected macrophages from Ab- or CAR-driven exhaustion, allowing for efficient phagocytosis of tumors after multiple rounds of cancer re-challenges.  Ablation of SIRPa in iMacs improved survival of mice grafted ovarian carcinoma and treated with anti-HER2 Abs, while anti-GD2 CAR iMacs with KO SIRPA demonstrated reduction of initial tumor burden in mice with metastatic neuroblastoma xenograft. Overall, our studies support the feasibility of using the iPSC platform to generate SIRPα-ablated iMacs that are resistant to CD47-mediated inhibition for therapeutic applications.

overall_design: RNA-seq data of SIRPA-KO, SIRPA-ITAM, and WT iMacs either cultured alone, with SKOV-3 ovarian cancer and anti-HER2 for 24 hours, or for 96 hours. All co-culture samples were sorted for CD45-bead selection prior to total RNA isolation.

first: Portia

last: Smith

first: Mark

last: Berres


title: Endogenous fine-mapping of functional regulatory elements in complex genetic loci in HAP1 [4C]

city: Tianjin

first: Ke

last: Zhao

address: Qixiangtai Road, No.22

institute: Tianjin Medical University

summary: The majority of genetic loci linked to polygenic complex traits are found in non-coding regions of the human genome. These loci often exhibit complex gene regulatory relationships and linkage disequilibrium (LD) configurations, making it challenging to accurately identify causal variants and their target genes. We used multiplexed single-cell CRISPR interference and activation perturbations to investigate cis-regulatory element (CRE) and target gene expression relationships within tight LD in the endogenous chromatin context. We demonstrated the prevalence of multiple causality in perfect LD (pLD) for independent expression quantitative trait locus (eQTL) and uncovered fine-grained genetic effects on gene expression within pLD, which are difficult to decipher using traditional eQTL fine-mapping or existing computational methods. We found that over one third of the causal CREs lack classical epigenetic markers, and we functionally validated one of these hidden regulatory mechanisms. Leveraging Multiome single-cell epigenetic and sequence perturbations, we highlighted the regulatory plasticity of the human genome. Our study will guide the exploration of missing causal mechanisms in molecular trait formation and disease development.

overall_design: Nine scRNA-seq datasets under varying multiplicity-of-infection (MOI) conditions, three single-cell multiome datasets (ATAC + Gene Expression) from modified HAP1 clones(dCas9-KRAB-blast, dCAS9-VP64-blast and Cas9-blast stably expressed HAP1 clones), four ATAC-seq datasets from wild-type and CRISPR-edited HAP1 cells(64bp deletion, 17bp deletion and CRISPRi around rs73156934-tagged cis-regulate element(CRE)), six CUT&TAG histone modification datasets and 1 Micro-C dataset from wild-type HAP1 cells, and three 4C-seq datasets for validating CRE-gene regulatory relationships in HAP1 cells.

first: Yao

last: Zhou

first: Mulin

last: JunLi


title: Direct lysine dimethylation of IRF3 by the methyltransferase SMYD3 attenuates antiviral innate immunity [II]

city: 武汉

first: zixuan

last: wang

state: 湖北省

address: 湖北省武汉市东湖南路7号

institute: 中国科学院水生生物研究所

summary: Interferon regulatory factor 3 (IRF3) is the key transcription factor in the type I IFN signaling pathway, whose activation is regulated by multiple posttranslational modifica_x0002_tions. Here, we identify SMYD3, a lysine methyltransferase, as a negative regulator of IRF3. SMYD3 interacts with IRF3 and catalyzes the dimethylation of IRF3 at lysine 39. This modification reduces IRF3 phosphorylation, dimerization, and subsequent nuclear translocation, leading to the inhibition of downstream type I interferon production. In addition, Smyd3-deficient mice are more resistant to RNA and DNA viral infections. Zebrafish lacking smyd3 or treated with the inhibitor BCI121 are also more resistant to viral infection. Our findings reveal a role for SMYD3 in the regulation of antiviral innate immunity and provide insight into a unique modulation of IRF3 that affects its activation.

overall_design: Whole RNA from MEF cells was purified using the RNeasy Mini Kit (QIAGEN, #74104). To ensure the quality of the information analysis, fastp software was used to remove the splice sequences, filter the low quality, N bases (indicating that the base information could not be determined), and obtain high quality clean data. The clean data and the number of bases and sequences of the clean data were counted. At the same time, the GC, Q20, Q30 contents of the clean data were calculated (Benagen Technology, Wuhan, China). Volcano Plot were generated using the ggplot2 software. Heatmaps were generated using the Multi Experiment Viewer (MeV) software. Gene Ontology (GO) enrichment analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses for the differentially expressed genes (DEGs) was performed using Cluster Profiler version 3.8.

first: Zixuan

last: Wang

first: Xiaoyun

last: Chen

first: Xing

last: Liu

first: Wuhan

last: Xiao


title: Wild type and tumor-derived mutant p53 share a non-canonical mode of gene occupancy [ChIP-seq]

first: Moray

last: Campbell

overall_design: Briefly, approximately cells were crosslinked with 1% formaldehyde solution, quenched with glycine (0.125 M) and harvested in cold PBS. Sonication of crosslinked chromatin was performed using a Bioruptor® UCD-200TM Sonicator (Diagenode) with optimized cycles for each cell type. Immunoprecipitated DNA fragments (using CM5 antibody, or IgG (Rabbit IgG D1 antibody – Santa Cruz)) were purified the following day using the Qiagen Qiaquick PCR purification Kit Protocol and antibody/bead complexes isolated with Magna ChIPTM Protein A+G magnetic beads (Millipore). Complexes were washed, reverse crosslinked, and treated sequentially with RNase and proteinase K prior to DNA isolation. The sample and input DNA can be used for PCR or measured using the Qubit reader and used for ChIP-seq.

first: M

first: J

last: Manfredi


title: Innate immune pathways drive early inflammation in the intravascular space in stroke patients with large vessel occlusion

city: Hamburg

first: Berenike

last: Rutz

address: Martinistrasse 52

institute: University Medical Center Hamburg-Eppendorf

summary: Neuroinflammation contributes substantially to stroke pathophysiology. This study investages the transcriptome of human peripheral blood monocytes from individuals with acute ischemic stroke.

overall_design: Peripheral blood mononuclear cells (PBMCs) from the intravascular compartment of the ischemic hemisphere were compared to systemic arterial controls from ischemic stroke patients and analyzed using scRNAseq. Furthermore cells were incubated with TotalSeq-C Human Universal Cocktail (Biolegend #399905) according to the manufacturers instructions.


title: Commensal protists regulate intestinal CD8+ T cells to shape mucosal immune memory

city: Stanford

first: Colin

last: Raposo

address: 240 Pasteur Drive, BMI 4600

department: Pathology

institute: Stanford University

summary: Tissue and microbial cues regulate the abundance and function of CD8+ T cells at barrier sites, yet the impact of specific microbes on their long-term durability remains unclear. Here, we show that the commensal protist Tritrichomonas musculus (T. mu) depletes intestinal CD8+ T cells, particularly tissue resident memory (TRM) cells, through activation of localized type 2 immunity. Colonization with T. mu or administration of its major secreted metabolite, succinate, led to the rapid decline of intestinal CD8+ T cells but left systemic memory T cells unaffected. The purinergic receptor, P2RX7, is highly expressed by intestinal TRMs and chemical antagonism of this receptor markedly restored CD8+ T cells during succinate feeding. Using lymphocytic choriomeningitis virus (LCMV) infection to track antigen-specific CD8+ memory T cells, we found viral-specific CD8+ TRMs repopulate the intestine independent of LCMV reinfection after removal of succinate treatment. These findings highlight how commensal protists and their metabolites reset homeostatic CD8+ T cell carrying capacity through damage-independent stimulation of TRM apoptosis and regulate mucosal memory.

overall_design: 6-week-old WT C56BL6/J mice were colonized with 1e6 T. mu or control (n=2 for control, n=5 for T. mu colonized). After 21 days, mice were sacrificed and the distal small intestine was excised. Intraepithelial CD8a+ T cells were sorted and pooled scRNA-seq was performed.

first: Katherine

middle: F

last: Nico

first: Ansuman

last: Satpathy

first: Michael

middle: R

last: Howitt


title: E3 ubiquitin ligase DTX2 reinforces innate antiviral immune responses by modulating K63-linked ubiquitination of TBK1

first: Zexing

last: Li

address: No.92 Weijin road

department: School of Life Sciences

institute: Tianjin University

summary: TBK1 plays a pivotal role in innate antiviral immunity, serving as a critical mediator downstream of various pattern recognition receptors (PRRs). TBK1 activity is tightly regulated by post-translational modifications (PTMs) including ubiquitination to maintain innate immune homeostasis. Here, we identify the Deltex E3 ubiquitin ligase 2 (DTX2) as a positive regulator of innate antiviral signaling in response to RNA viruses. We find that DTX2 overexpression enhances antiviral signaling, whereas loss of DTX2 evidently reduces antiviral responses. Mechanistically, we demonstrated that DTX2 directly interacts with TBK1 and promotes TBK1 K63-linked polyubiquitination to regulate antiviral signaling. Collectively, our study reveals that DTX2 plays a novel positive regulatory role in antiviral signaling by modulating the K63-linked ubiquitination of TBK1 to maintain the balance of innate antiviral immune responses.

overall_design: RNA-seq profiling of wildetype and Dtx2 knockout  MEF cells 8 hours after Sendai virus infection.

first: Di

last: Xin


title: Transcriptome Sequencing Screened Differentially Expressed Genes in GES-1 intervened with Ulcer-Healing Decoction

first: Xiaohong

last: Shi

address: No.280 Xiangshan Road, Xihu District

department: Department of Traditional Chinese Medicine

summary: To analyze the gene expression of human gastric epithelial cells (GES-1) regulated by paeonifl or in based on transcript tome sequencing; to identify and analyze key genes; to study the mechanism of paeoniflorin on Leuk-1cells; and to explore its medicinal value. Method: Different doses of Ulcer-healing Decoction were used to treat GES-1, and its activity was assessed by MTT assay. Transcriptome sequencing analysis was then performed to identify differentially expressed genes (DEGs). The function and pathways of the DEGs were annotated using GO and KEGG enrichment analysis. Result: The survival rate of GES-1 cells was significantly increased under different concentrations of ulcer-healing decoction intervention (P < 0.05 or P < 0.01). After 24 hours of treatment with 80 µ g/mL ulcer-healing decoction, a total of 312 crucial DEGs were observed in GES-1 cells, of which 233 DEGs were upregulated and 79 DEGs were downregulated. The top five differentially expressed genes include CYP1A1, IL24, IKBKGP1, GDF15, SCUBE1 and OASL, ADAMTS5, SULF1, ID2, and DDX60. The functions of these genes are mainly related to promoting wound,cell proliferation and migration,immunity and inflammation.GO enrichment analysis showed that the DEGs were significantly enriched in the extracellular region and involved in biological processes such as cytokine production and response, stimulus response, and exerting molecular functions such as proliferation factor receptor activity. KEGG pathway enrichment showed that the DEGs were significantly enriched in various signaling pathways. Immunohistochemistry and other methods were used to detect differentially expressed proteins in GES-1 cells induced by ulcer-healing decoction, and the results were consistent with those from RNA-seq, thereby validating the accuracy of sequencing technologies. Conclusion: The role of ulcer-healing decoction in GES-1 is primarily reflected in regulating inflammatory factors and growth factors, promoting cell proliferation, and supporting the regeneration and repair of gastric mucosal epithelial cells.

overall_design: RNA-seq analyzing differentiation of human gastric epithelial cell intervened with Ulcer-Healing Decoction of adipogenic

first: Xiao

last: Lv

first: Ying

last: Yu


title: Purified Astragalus Polysaccharide Combined with Inactivated Vaccine Markedly Prevent Infectious Haematopoietic Necrosis Virus Infection in Rainbow Trout (Oncorhynchus mykiss)

city: Lan zhou

first: Pan

middle: Yu

last: Cai

state: Gansu

address: Yin Tan Road Street, An Ning District

department: College of Animal Science and Technology

institute: Gansu Agricultural University

summary: Rainbow trout (Oncorhynchus mykiss) is experiencing a catastrophic pandemic. In recent years, infectious hematopoietic necrosis virus (IHNV) has spread nationwide, resulting in significant mortality. Currently, there are no available treatments or vaccines for IHNV in China. Here, Astragalus extract was purified and characterized. Then, we developed an inactivated IHNV vaccine with purified Astragalus polysaccharide (P-APS) as an adjuvant. Safety assays showed that IHNV was successfully inactivated. After a serious IHNV challenge, the cumulative mortality rates were 76.0%, 38.0% and 22.08% in control, vaccine and P-APS+vaccine group, respectively. P-APS+vaccine was effective in reducing head kidney damage and apoptosis after IHNV challenge by histopathological and TUNEL analyses. P-APS+vaccine group showed better results in enhancing specific antibodies (IgM) and immune enzyme activities (C3, LZM, GOT and GPT). RNA-seq revealed that many immune-related pathways were significantly enriched. TLR2, TLR7, C3, IFN-γ, IgM, MHC1, MHC2, MX1, and VIG1 identified as core genes based on RNA-seq and PPI networks. Mechanistic investigations showed that the P-APS+vaccine activates the immune pathway by upregulating the expression of these genes. P-ASP+vaccine induced effective innate and adaptive immune responses that were stronger than single vaccines after vaccination and IHNV challenged. Our findings will provide a promising vaccine candidate against IHNV.

overall_design:
Fish were divided into three experimental groups (P-APS+vaccine, vaccine and control) to test the immunoprotective effect of the vaccine and P-APS+vaccine. Each experimental group had 150 fish, 100 of which were used for mortality measurements and the remaining 50 for sampling. The injection volume was 150 μL (intraperitoneal) in both vaccine and P-APS+vaccine groups, and 150 μL of PBS was injected under the same conditions in the control group. At 14 days (D 14) after vaccination, 150 fish per group were injected intraperitoneally with 50 µl IHNV (107 TCID50 ml-1) and monitored for mortality for two weeks. During vaccination and sampling, fish were anesthetized with 50 mg/L MS-222 (Sigma, St. Louis, MO, USA). Four fish from each treatment group were sacrificed for head kidney and blood collection after immunization at D1, D3, D7, D14, D15, D21 and D30 for assaying immune-related gene expression and non-specific enzyme activity. Notably, a total of seven fish head kidneys were collected from each group at D7 and D21, four of which were used for the gene expression assays described above, and the other three were used for RNA-seq. In addition, the head kidneys of three fish for each group were collected at D1, D3, D7, D14, D15, D21, D30 and D60 for histopathological examination.
Research and Demonstration of Integrated Prevention and Control Technologies for Rainbow Trout with IHN and IPN (Grant No. GSLK-2022-11), and Discipline Team Project of Gansu Agricultural University (NO: GAU-XKTD-2022-23)

first: Zhe

first: Yucai

last: Pan

first: Jinqiang

last: Quan

first: Wei

last: Gu

first: Junwei


title: Elucidation and Engineering of Sphingolipid Biosynthesis Pathway inYarrowia lipolyticaforProduction of Sphingoid Bases and Glycosylceramides

city: Seoul

first: Seo Hyeon

last: Shin

state: Seoul

address: Seoul 동작구 흑석로 84

institute: Chung-ang University

summary: This study focused on understanding and modifying the sphingolipid biosynthesis pathway in the yeast Yarrowia lipolytica. By altering specific biological processes within the yeast, the creation of new types of lipid molecules became possible. The research findings highlight the high potential of this yeast in producing non-acetylated sphingoid bases and human-type glucosylceramides, which could serve as valuable components in cosmetics or nutritional supplements.

overall_design: To investigate the profiles of gene expression in C4-hyroxylase, SUR2, disrupted mutant strain, we performed mRNA seq analysis of sur2 deleted mutant strain compared to the Yarrowia lipolytica wild-type strain which were cultivated in YPD medium. The experiment collected data through two replicates.

first: Seo

middle: H

first: Hye

middle: Y

last: Moon

first: Hae

last: Park

first: Hyun

last: Kang


title: Tumor and immune environment changes in EGFR-TKI treated EGFR-mutant non-small cell lung cancer

first: Soomin

last: Kim

address: 101, Daehak-ro, Jongno-gu

institute: Seoul National University Cancer Research Institute

summary: Patients with EGFR mutations exhibit immunosuppressive microenvironments, limiting responsiveness to immunotherapy. In our analysis, 25 Korean patients diagnosed with NSCLC harboring common EGFR mutations (L858R and exon 19 microdeletion) that received EGFR-TKI treatment was compared before and after treatment. 25 patients were analyzed overall, and 14 patients were analyzed in the paired analysis who received first-line osimertinib. Among 171 ROIs, 68ROIs were used for tumor paired analysis, and 51 ROIs for immune region.

overall_design: In this study, clinical specimens from 25 patients were selected for spatial and whole transcriptome analysis. To decipher the molecular changes that occur with EGFR-TKI (especially, osimertinib) treatment, we conducted DEG analysis by categorizing the patients into pre-treated and post-treated groups. This study was approved by the Institutional Review Board of the Seoul National University Hospital and performed in accordance with the Declaration of Helsinki.

first: Jaemoon

last: Koh

first: Songji

last: Oh

first: Tae Min


title: Small cell carcinoma of the ovary, hypercalcemic type (SCCOHT) cell line H3K27ac CUT&RUN

first: Jessica

last: Lang

overall_design: BIN67, SCCOHT, and COV434 SCCOHT cell line H3K27ac & IgG CUT&RUN

first: Jeffrey

last: Trent


title: SARS-CoV-2 infection of human pluripotent stem cell-derived vascular cells reveals smooth muscle cells as key mediators of vascular pathology during infection II

city: Cambridge

first: Rudolf

last: Jaenisch

address: 455 Main St.

institute: Whitehead Institute for Biomedical Research

summary: Nearly 20% of hospitalized patients diagnosed with severe SARS-CoV-2 infection are at risk for thromboembolic events. We developed a model of SARS-CoV-2 infection using human-induced pluripotent stem cell-derived endothelial cells, pericytes, and smooth muscle cells to recapitulate the vascular pathology associated with SARS-CoV-2 exposure. Our results demonstrate that perivascular cells, particularly smooth muscle cells (SMCs), are a specifically susceptible vascular target for SARS-CoV-2 infection. Utilizing RNA sequencing, we characterized the transcriptomic changes accompanying SARS-CoV-2 infection of SMCs, and endothelial cells (ECs). We observed that infected human SMCs shift to a pro-inflammatory state and increase the expression of key mediators of the coagulation cascade. Further, we showed human ECs exposed to the secretome of infected SMCs produce hemostatic factors that can contribute to vascular dysfunction, despite not being susceptible to direct infection. The findings here recapitulate observations from patient sera in human COVID-19 patients and provide mechanistic insight into the unique vascular implications of SARS-CoV-2 infection at a cellular level.

overall_design: Human endothelial cells (ECs), pericytes (PCs), and smooth muscle cells (SMCs) were differentiated from human pluripotent stem cells (hPSCs). hPSC-derived ECs, PCs, and SMCs were treated with either live SARS-CoV-2, heat-inactivated (HI) SARS-CoV-2 or mock-infected (no virus). We compared the transcriptional changes induced by each of these conditions in both ECs and SMCs.  We also compared the transcriptional changes induced by the treatment of ECs with media from SMCs pr PCs that were infected with SARS-CoV-2 or HI SARS-CoV-2.  For the media transfer experiments, ECs were exposed to media from SMCs that had been exposed to live or HI SARS-CoV-2, and then RNA was isolated from the ECs 48 hours after exposure.  No treatment samples were used for comparison of cell types with the primary cell counterpart.

first: Richards

last: Alexsia


title: Early precursor-derived pituitary gland tissue-resident macrophages play a pivotal role in modulating hormonal balance [bulk RNA-Seq]

city: Turku

first: Pia

last: Rantakari

address: Tykistökatu 6B

department: Turku Bioscience Centre

country: Finland

institute: University of Turku

summary: Macrophages were identified in the pituitary gland long ago, but until recently, we lacked the resolution to comprehensively profile the pituitary gland macrophage compartment. To address this, we examined the developmental kinetics and distinctive characteristics of pituitary macrophages within the developing and homeostatic gland. Whole pituitary glands were collected from newborn (NB) to adult (8 weeks old) wild-type (WT) mice.  Our analysis revealed diverse macrophage subsets with marked differences in gene expression profiles and spatial localization, undergoing dynamic phenotypic changes from neonatal stages to adulthood. We observed distinct transcriptional profiles between macrophage subpopulations in the anterior and posterior lobes, underscoring significant molecular divergences.  Fate-mapping analyses showed that pituitary macrophages originate from early yolk sac precursors and persist throughout life primarily through local proliferation, even under pathophysiological conditions.

overall_design: To investigate the transcriptome changes in the pituitary gland after macrophage depletion. Macrophages were ablated with function-blocking αCSF1R antibody. The first injection was given during embryo development at E6.5. After birth, an αCSF1R antibody was given every second week (4 times in total), and tissues were collected one week after the last injection (at the age of 5 weeks).

first: Henna

last: Lehtonen

first: Heli

last: Jokela

first: Julian

last: Hofmann

first: Lauriina

last: Tola

first: Arfa

last: Mehmood

first: Florent

last: Ginhoux

first: Burkhard

last: Becher

first: Melanie

last: Greter

first: Gennady

middle: G

last: Yegutkin

first: Marko

last: Salmi

first: Heidi

last: Gerke


title: Early precursor-derived pituitary gland tissue-resident macrophages play a pivotal role in modulating hormonal balance [scRNA-Seq]

overall_design: 10x Chromium single-cell RNA sequencing on CD45 positive cells isolated from the pituitary glands of male C57BL/6N mice using fluorescence-activated cell sorting.


title: Early precursor-derived pituitary gland tissue-resident macrophages play a pivotal role in modulating hormonal balance

summary: This SuperSeries is composed of the scRNA-Seq and bulk RNA-Seq SubSeries listed below.

overall_design: Refer to individual Series


title: Transcriptome Analysis with Human Vascular smooth muscle cells under 2D and 3D cell culture conditions

city: Guangzhou

first: Jiang

last: Liu

address: 106 Zhongshan 2nd Road, Guangzhou City, Guangdong Province

institute: Guangdong Provincial People's Hospital ,Guangdong Academic of Medical Sciences

summary: The current objectives are to use of transcriptome sequencing analysis technology comprehensively understand the potential molecular mechanisms between vascular smooth muscle cells grown in 2D environments (2D-VSMCs) and vascular smooth muscle cells grown in 3D PGA environments (3D-VSMCs).These findings help in gaining a better grasp of the situation of how VSMCs survive on PGA scaffolds and have guiding significance for TEBV culture.

overall_design: Human vascular smooth muscle cells  were cultured in 2D and PGA based 3D culture for 4 days. Total RNA was extracted using Trizol. RNA-SEQ was carried out to profile the gene expression in both culture conditions.

first: Peng

first: Lijun

last: Fang

first: Xichun

last: Wang

first: Shilong

last: Zhong

first: Yueheng

last: Wu

first: Zibei

last: Feng

first: Zhanyi

last: Lin


title: Inhibition of DEK contributes to the tolerance of replication stress and restores HSC function in Fanconi anemia [12-ATAC]

city: Chongqing

first: Zhe

last: Chen

address: 1 Yixueyuan Road, Yuzhong District

institute: Chongqing Medical University

summary: Replication stress drives functional decline in HSCs and is a major driver of BM failure in Fanconi anemia (FA). At present, how HSCs respond and counteract replication stress remains largely unknown. Using integrated multi-omics, we demonstrate that global chromatin relaxation is a prerequisite for the activation of stress-responsive genes and replication fork stabilization/progression in HSCs under replication stress. The reduced DEK (a chromatin architectural protein) contributes to the chromatin relaxation in HSCs, whereas DEK-overexpressed HSCs are confronted with a strong replication challenge, resulting in impaired HSC maintenance and hematopoiesis. Fancd2 deletion induces DEK expression and causes replication stress in HSCs, while haploinsufficiency of DEK promotes chromatin opening in Fancd2-deficient HSCs and substantially recovers HSC function. Notably, DEK expression is abnormally up-regulated in bone marrow (BM) CD34+ cells from FA patients. Inhibition of DEK significantly restores the proliferation capacity in vitro and engraftment in vivo of BM CD34+ cells from FA patients. At the molecular level, we identify that the transcriptional factor ATF2 directly promotes DEK transcription, mainly relying on the phosphorylated ATF2 (Thr69/71). Wild-type HSCs reduces DEK expression to counteract replication stress through the ATR kinase to phosphorylate ATF2 at Ser490/498. However, Fancd2 deficiency induces hyper-phosphorylation of p38 that further phosphorylates ATF2 at Thr69/71, causing DEK accumulation in HSCs. Collectively, our findings provide the first evidence of a functional link between chromatin relaxation and replication stress tolerance in HSCs, and highlight DEK as a potential molecular target for FA.

overall_design: Bone marrow CD34+ cells from five healthy donors and five patients with FA were isolated, followed by ATAC-seq


title: Inhibition of DEK contributes to the tolerance of replication stress and restores HSC function in Fanconi anemia [13-CUT-Tag]

overall_design: HSCs (CD48-CD150+LSK) from Wild-type, DekTg, and Dek-cKO mice were sorted, followed by DEK CUT&Tag-seq. Besides, HSCs (CD48-CD150+LSK) from DekTg mice were sorted and treated control aptamer and DTA-64 (2 µM), followed by DEK CUT&Tag-seq


title: Endogenous fine-mapping of functional regulatory elements in complex genetic loci in HAP1 [CUT&Tag]

first: Ke

last: Zhao

overall_design: Nine scRNA-seq datasets under varying multiplicity-of-infection (MOI) conditions, three single-cell multiome datasets (ATAC + Gene Expression) from modified HAP1 clones(dCas9-KRAB-blast, dCAS9-VP64-blast and Cas9-blast stably expressed HAP1 clones), four ATAC-seq datasets from wild-type and CRISPR-edited HAP1 cells(64bp deletion, 17bp deletion and CRISPRi around rs73156934-tagged cis-regulate element(CRE)), six CUT&TAG histone modification datasets and 1 Micro-C dataset from wild-type HAP1 cells, and three 4C-seq datasets for validating CRE-gene regulatory relationships in HAP1 cells.

first: Yao

last: Zhou

first: Mulin

last: JunLi


title: Suppression of NRF2-dependent cancer growth by a covalent allosteric molecular glue

city: San Diego

first: Brent

last: Kuenzi

address: 5820 Nancy Ridge Dr

institute: Vividion Therapeutics

summary: The NRF2 transcription factor is constitutively active in cancer where it functions to maintain oxidative homeostasis and reprogram cellular metabolism. NRF2-active tumors exhibit NRF2-dependency and resistance to chemo/radiotherapy. Here we developed VVD-065, a first-in-class NRF2 inhibitor that acts via an unprecedented allosteric molecular glue mechanism. In the absence of stress or mutation, NRF2 is rapidly degraded by the KEAP1-CUL3 ubiquitin-ligase complex. VVD-065 specifically and covalently adducts C151 on KEAP1, which in turn promotes KEAP1-CUL3 complex formation, leading to dramatic enhancement of NRF2 degradation. Previously reported C151-directed compounds decrease KEAP1-CUL3 interactions and stabilize NRF2, thus establishing KEAP1_C151 as a tunable regulator of the KEAP1-CUL3 complex and NRF2 stability. VVD-065 inhibited NRF2-dependent tumor growth and sensitized cancers to chemo/radiotherapy, supporting an open Phase I clinical trial (NCT05954312).

overall_design: To understand the downstream effects of KEAP1 activation, we measured the transcriptional changes following oral dosing of VVD-065 in immunocompromised mice bearing KYSE70 tumor xenografts

first: Matthew

last: Patricelli

first: Nilotpal

last: Roy

first: Tine

last: Wyseure


title: Pde4b-regulated cAMP signaling pathway in the AUDGABA-S1TrSst circuit underlies acute stress-induced anxiety-like behavior

city: qingdao

first: xufeng

last: xu

address: No. 16 Jiangsu Road

institute: qingdao university

summary: Acute stress-induced anxiety is an important way for animals to avoid danger. However, neural and molecular mechanisms that underlie control of anxiety behavior are largely elusive. Here, we find acute physical stress activates a large number of neurons in the primary somatosensory cortex, trunk region (S1Tr). Single-cell sequencing reveals the S1Tr c-fos positive neurons activated by acute stress are largely GABAergic somatostatin (Sst) neurons. These S1TrSst neurons activated by acute stress showed desensitization during subsequent anxiety-like behavior tests. Selective inhibition or apoptosis of S1TrSst neurons mimics acute stress effects to induce anxiety. In contrast, selective activation of S1TrSst neurons reduced acute stress-induced anxiety. Furthermore, we demonstrate that S1TrSst cells receive inputs from the secondary auditory cortex, dorsal area (AUD) GABAergic neurons to modulate acute stress-induced anxiety. Finally, from the results of spatial transcriptome sequencing and precise projection-specificity Pde4b protein knockdown strategy, we show that acute stress reduces Pde4b-regulated cyclic adenosine monophosphate (cAMP) signaling pathway activity in the AUDGABA-S1TrSst projections and resulting in a hypoactivity of S1TrSst neurons during subsequent behavioral tests. Our study unveils a neural and molecular mechanism for acute stress-elicited anxiety and affords a theoretical basis for clinical treatment of anxiety disorders.

overall_design: Investigation of neuronal gene expression profiles in the S1Tr brain region of mice following acute stress exposure

first: zhixin

last: xiao

first: xiaoya

last: wang

last: zhou

first: xuetong

last: yi

first: xiaoqi

last: zhang

first: qilin

last: wu

first: zhuo

last: li


title: Elucidating ADNP Syndrome Gene Dysregulation through a Novel Approach in iPSC-Derived GABAergic Neuron Development [ADNP_scRNA-seq]

city: New York

last: Yan

email: linda.lee2@mssm.edu, nan.yang1@mssm.edu

address: 1425 Madison Ave

department: Neuroscience

institute: Mount Sinai

summary: GABAergic interneurons are essential for maintaining neural circuit function, and their dysfunction is linked to a range of neurodevelopmental and neurodegenerative disorders. Traditional approaches for differentiating human pluripotent stem cells (PSCs) into neuronal cells are often hindered by challenges such as incomplete neural differentiation, prolonged culture times, and variability across PSC lines. To overcome these limitations, we developed an innovative strategy by combining the overexpression of transcription factors Ascl1 and Dlx2 with dual-SMAD and WNT inhibition, effectively driving the differentiation of PSCs into diverse, region-specific GABAergic neuron subtypes. Through single-cell sequencing, we thoroughly characterized the heterogeneity of these induced neurons (iNs) and uncovered the regulatory mechanisms underlying their fate specification. Our findings demonstrate the successful generation of multiple disease-relevant GABAergic neuron types. Furthermore, we investigated the impact of the ADNP syndrome-associated p.Tyr719* variant on GABAergic neuron differentiation, revealing that this mutation significantly alters neuronal subtype composition and synaptic transmission. This study advances our understanding of the diversity of PSC-derived GABAergic neurons and underscores their potential as models for exploring neurological disorders.

overall_design: We performed single cell RNA-seq to measure transcriptomic profiles in human induced GABAergic (GABA) neurons derived from  neural progenitor cells (NPCs) with Ascl1 and Dlx2 overexpression and mutations in ADNP gene

first: Ruiqi

last: Hu

first: Linda

last: Boshans

first: Sai

last: Ma

first: Bohan

last: Zhu

first: Xuran

last: Wang

last: Yang


title: Peripheral nerve injury induces dystonia-like movements and dysregulation in the energy metabolism: A multi-omics descriptive study in Thap1+/− mice

city: Würzburg

first: Chi Wang

last: Ip

address: Josef-Schneider-Strasse 11

department: Neurology

institute: Neurologische Klinik und poliklinik

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


title: Prdm16os is a brain and astrocyte-specific lncRNA implicated with Alzheimer’s disease

city: Gottingen

first: Andre

last: Fischer

state: NS

address: Von Sieboldstr 3a

department: Epigenetic Mechanisms in Dementia

institute: German Center for Neurodegenerative Diseases (DZNE)

summary: Astrocytes provide crucial support for neurons, contributing to synaptogenesis, synaptic maintenance, and neurotransmitter recycling. Under pathological conditions, deregulation of astrocytes contributes to neurodegenerative diseases such as Alzheimer’s disease (AD), highlighting the growing interest in targeting astrocyte function to address early phases of AD pathogenesis. While most research in this field has focused on protein-coding genes, non-coding RNAs, particularly long non-coding RNAs (lncRNAs), have emerged as significant regulatory molecules. In this study, we identified the lncRNA PRDM16-DT as highly enriched in the human brain, where it is almost exclusively expressed in astrocytes. PRDM16-DT and its murine homolog, Prdm16os, are downregulated in the brains of AD patients and in AD models. In line with this, knockdown of PRDM16-DT and Prdm16os revealed its critical role in maintaining astrocyte homeostasis and supporting neuronal function by regulating genes essential for glutamate uptake, lactate release, and neuronal spine density through interactions with the RE1-Silencing Transcription factor (Rest) and Polycomb Repressive Complex 2 (PRC2). Notably, CRISPR-mediated overexpression of Prdm16os mitigated functional deficits in astrocytes induced by stimuli linked to AD pathogenesis. These findings underscore the importance of PRDM16-DT in astrocyte function and its potential as a novel therapeutic target for neurodegenerative disorders characterized by astrocyte dysfunction.

overall_design: To investigate the function of Prdm16os, we performed a knockdown (KD) using Gapmers (Qiagen) in primary mouse astrocytes on DIV12. We harvested the cells on DIV14, isolated RNA, generated libraries and performed total RNA sequencing

first: Sophie

last: Schroeder


summary:
To elicit a dystonia-like phenotype in a genetically predisposed DYT-THAP1 mouse model (Thap1+/-) by performing a right sciatic nerve crush injury.
To identify novel pathophysiological pathways and possible biomarker, we performed a multi-omic analysis of three dystonia-relevant brain regions

overall_design: Comparative gene expression profiling analysis of RNA-seq data for the cortex, striatum and cerebellum of naive and nerve-injured wildtype and Thap1+/- mice

first: Colette

last: Reinhold

first: Rhonda

last: McFleder

first: Panagiota

last: Arampatzi

first: Tom

last: Gräfenhan

first: Thorsten

last: Bishler


title: RNA-seq of UHRF1 KO A549 cells vs. control A549 cells.

city: Beijing

first: Zhou

last: Xindi

state: Beijing

address: 30 Shuangqing Road

department: School of Medicine

institute: Tsinghua University

summary: To investigate the gene expression signatures of ICAM-1 candidate negative regulator UHRF1, we generate the UHRF1 KO cells and profiled the transcriptome changes by paired-end RNA seq.

overall_design: Comparative gene expression profiling analysis of RNA-seq data of UHRF1 KO A549 cells versus control A549 cells.

first: Xindi

last: Zhou

first: Deng

last: Pan


title: scRNA-seq of TIL CD45+ cells with BiAb treatment

summary: To investigate the effects of fusion protein on tumor microenvironment, we performed scRNAseq of tumor infiltrating immune cells of antibody treated MC38 tumor.

overall_design: Comaprative the CD45+ single cell effects of Cetuximab and Cet×ICAM1-D1 treated MC38 tumors.

first: Tian

last: Xu


title: RNA-seq, Bisulfite-seq in A549 cells and scRNA-seq of TIL

type: Methylation profiling by high throughput sequencing

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus


title: Targeting FTO suppresses hepatocellular carcinoma by inhibiting ERBB3 and TUBB4A expression [MeRIP]

first: Zhe

last: Chen

summary: Fat mass and obesity-associated protein (FTO) is an N6-methyladenosine (m6A) demethylase and plays a critical oncogenic role in various cancers. Here we show that FTO is an effective target in hepatocellular carcinoma (HCC). FTO is highly expressed in clinical patients of HCC and presents as a poor prognostic factor. Genetic depletion of FTO dramatically attenuated HCC progression in vivo. Pharmacological inhibition of FTO by FB23/FB23-2 remarkably suppressed the proliferation and migration of HCC cell lines in vitro, as well as inhibiting the HCC tumorigenicity in xeno-transplanted mice. Mechanistically, FB23-2 suppressed the expression of Erb-b2 receptor tyrosine kinase 3 (ERBB3) and human tubulin beta class Iva (TUBB4A) by increasing the m6A level in these mRNA transcripts. The decreased ERBB3 resulted in inhibited Akt-mTOR signaling that subsequently impaired the proliferation and survival of HCC cells. Moreover, FB23-2 abolished the organization of the tubulin cytoskeleton, while enforced expression of TUBB4A rescued the migration of HCC cells. Collectively, our study demonstrated that FTO plays a critical role in HCC by maintaining the proliferation and migration of cells, and highlights the broad potential of FTO inhibitors for targeting HCC.

overall_design: To find the targets of FB23-2 via inhibiting FTO, MHCC97H cells with FB23-2 (2 μM) treatment for 48 h were selected for RNA sequencing (RNA-seq) and methylated RNA immunoprecipitation sequencing (MeRIP-seq), DMSO-treated MHCC97H cells as vehicle.


title: Peripheral nerve injury induces dystonia-like movements and dysregulation in the energy metabolism: A multi-omics descriptive study in Thap1+/- mice

summary: To elicit a dystonia-like phenotype in a genetically predisposed DYT-THAP1 mouse model (Thap1+/-) by performing a right sciatic nerve crush injury.

overall_design:
To identify novel pathophysiological pathways and possible biomarkers, we performed a multi-omic analysis of three dystonia-relevant brain regions
Comparative gene expression profiling analysis of miRNA-seq data for the cortex, striatum, and cerebellum of naive and nerve-injured wildtype and Thap1+/- mice


title: Inhibition of DEK contributes to the tolerance of replication stress and restores HSC function in Fanconi anemia (8-ATAC)

overall_design: HSCs (CD48-CD150+LSK) from wild-type C57BL/6J mice were transduced by pMSCV or MSCV-H1C retrovirus, followed by ATAC-seq


title: Inhibition of DEK contributes to the tolerance of replication stress and restores HSC function in Fanconi anemia (9-ATAC)

overall_design: HSCs (CD48-CD150+LSK) from Fancd2+/+Dekfl/+, Fancd2-/-Dekfl/+, Fancd2+/+Dek-HET, and Fancd2-/-Dek-HET mice were sorted, followed by ATAC-seq


title: Inhibition of DEK contributes to the tolerance of replication stress and restores HSC function in Fanconi anemia (7-RNA)

overall_design: Total RNA of HSCs (CD48-CD150+LSK) from DekCon and DekTg mice were extracted, followed by RNA-seq.


title: Inhibition of DEK contributes to the tolerance of replication stress and restores HSC function in Fanconi anemia (6-ATAC)

overall_design: HSCs (CD48-CD150+LSK) from DekCon and DekTg mice were sorted and cultured with DMSO or Hydroxyurea (HU, 100 µM) for 48 h, followed by ATAC-seq


title: Elucidating ADNP Syndrome Gene Dysregulation through a Novel Approach in iPSC-Derived GABAergic Neuron Development [scRNA-seq]

last: Yan

overall_design: We performed single cell RNA-seq to measure transcriptomic profiles in human induced GABAergic (GABA) neurons derived from human pluripotent stem cells (iPSCs) and neural progenitor cells (NPCs) with Ascl1 and Dlx2 overexpression for various durations to identify disease-relevant models

first: Ruiqi

last: Hu

first: Linda

last: Boshans

last: Ma


title: Inhibition of DEK contributes to the tolerance of replication stress and restores HSC function in Fanconi anemia (4-CUT-Tag)

overall_design: HSCs (CD48-CD150+LSK) from wild-type C57BL/6J mice were sorted and cultured with DMSO, Gemcitabine (Gem, 0.5 µM), or Hydroxyurea (HU, 100 µM) for 48 h, followed by H3K27ac CUT&Tag-seq


title: Inhibition of DEK contributes to the tolerance of replication stress and restores HSC function in Fanconi anemia (3-CUT-Tag)

overall_design: HSCs (CD48-CD150+LSK) from wild-type C57BL/6J mice were sorted and cultured with DMSO, Gemcitabine (Gem, 0.5 µM), or Hydroxyurea (HU, 100 µM) for 48 h, followed by H3K4me3 CUT&Tag-seq