title: Small RNA-seq analysis of circulating miRNAs in Cushing's syndrome

city: Valencia

first: Jose Santiago

last: Ibanez-Cabellos

state: VALENCIA

address: Av. de Blasco Ibanez, 15

institute: Universitat de Valencia

summary: Patients with Cushing’s syndrome (Cushing's syndrome) in remission show sustained fatigue, myopathy, and an increased prevalence of sarcopenia. The mechanisms that determine these persistent muscle problems are not well known. We aimed to identify circulating microRNAs (miRNAs) with differential expression that could be potential biomarkers for the diagnosis and/or prognosis in Cushing's syndrome. In this data set, we analized thirty-six women in sustained remission for 13 ± 7 years (mean ± SD) from Cushing's syndrome, with a median age (IQ range) of 51 (45.2–60) years and mean ± SD BMI of 27 ± 4 Kg/m2, and 36 matched healthy controls using small RNA-sequencing of small RNA purified from plasma samples of these patients and control. In 7 patients sarcopenia was present according to the European Working Group on Sarcopenia in Older People (EWGSOP) criteria. Validation was carried out using RT-qPCR. For the validation, Taqman probes were performed on QuantStudio 5 equipment (Applied Biosystems). In a first discovery group using RNA-sequencing, plasma samples of 18 Cushing's syndrome patients and 18 healthy subjects were investigated; circulating miR-28-5p, miR-495-3p and miR-654-5p were upregulated in Cushing's syndrome patients as compared with controls (p<0.05). In a validation study of the 3 upregulated miRNAs in 36 patients and 26 controls, no differences were observed by RT-qPCR; however, the expression of circulating miR-28-5p was upregulated in Cushing's syndrome patients with sarcopenia as compared with those without (AUC for fold-change in the ROC analysis, 0.798; p=0.0156). The optimized cut-off value for miR-28-5p to identify Cushing's syndrome patients with sarcopenia was 3.80, which yielded a sensitivity of 86% and a specificity of 69%.

overall_design: Plama miRNA profiles of 18 Cushing's syndrome and 18 healthy subjects were generated by deep sequencing using Illumina NextSeq 550.

first: Jose

last: Ibáñez-Cabellos

first: Marta

last: Seco-Cervera


title: FAM92A1 Regulates Neural Structure and Function through Membrane Remodeling

status: Public on Jul 05 2024

city: Helsinki

first: Liang

last: Wang

address: Viikinkaari 5

country: Finland

institute: University of Helsinki

type: Expression profiling by high throughput sequencing

summary: The Bin/amphiphysin/Rvs (BAR) domain protein FAM92A1 is a multifunctional protein engaged in the regulation of mitochondrial ultrastructure and ciliogenesis. However, the physiological role of FAM92A1 in the brain, particularly in neurons with specialized membrane architecture, remains unexplored. Here, we demonstrate that FAM92A1 is prominently expressed in the brain and neurons during early mouse embryonic development. FAM92A1 knockout mice display brain morphology alterations, age-associated memory decline, and cognitive deficits. Moreover, the absence of FAM92A1 triggers hippocampal neuron degeneration and disrupts neuronal complexity and synaptic plasticity. Crucially, FAM92A1 deficiency leads to anomalies in membrane remodeling and disturbances in neuronal endocytic processes. Through analysis of the crystal structure of the FAM92A1 BAR domain, we unveil its intrinsic capability to form dimers, contributing to a distinctive curved conformation critical for membrane remodeling. By combining structural and molecular simulation, we further gain a comprehensive understanding of how FAM92A1 interacts with membranes to induce lipid clustering and generate membrane curvature. In summary, our study provides valuable insights into the physiological function of FAM92A1 in the brain. It uncovers the molecular mechanisms through which FAM92A1 regulates synaptic plasticity and neural function by influencing membrane remodeling and endocytic processes.

overall_design: To investigate the changes in genes after the loss of FAM92A1, we dissected mouse hippocampus from three mice for each FAM92A1+/+, FAM92A1+/-, and FAM92A1-/- groups. All dissected mouse hippocampus was subjected to RNA-seq analysis. Comparative gene expression profiling analysis of RNA-seq data between wild type and FAM92A1-deficient group.

first: Xiaobo

last: Cen

first: Ziyun

last: Yang


title: Mutant androgen receptor induces neurite loss and senescence independently of ARE binding in a neuronal model of SBMA

city: Philadelphia

first: Diane

middle: E

last: Merry

state: PA

address: 1020 Locust St.

department: Biochemistry and Molecular Biology

country: USA

institute: Thomas Jefferson University

summary: Spinal and bulbar muscular atrophy (SBMA) is a slowly progressing neuromuscular disease caused by a polyglutamine (polyQ)-encoding CAG repeat expansion in the androgen receptor (AR) gene, leading to AR aggregation, lower motor neuron death, and muscle atrophy. AR is a ligand-activated transcription factor that regulates neuronal architecture and promotes axon regeneration; however, whether AR transcriptional functions contribute to disease pathogenesis is not fully understood. Using a differentiated PC12 cell model of SBMA, we identified dysfunction of polyQ-expanded AR in its regulation of neurite growth and maintenance. Specifically, we found that in the presence of androgens, polyQ-expanded AR inhibited neurite outgrowth, induced neurite retraction, and inhibited neurite regrowth. This dysfunction was independent of polyQ-expanded AR transcriptional activity at androgen response elements (AREs). We further showed that the formation of polyQ-expanded AR intranuclear inclusions promoted neurite retraction, which coincided with reduced expression of the neuronal differentiation marker -III-Tubulin. Finally, we revealed that cell death is not the primary outcome for cells undergoing neurite retraction; rather, these cells become senescent. Our findings reveal that mechanisms independent of AR canonical transcriptional activity underly neurite defects in a cell model of SBMA and identify senescence as a previously unappreciated pathway implicated in this pathology. These findings suggest that, in the absence of a role for AR canonical transcriptional activity in the SBMA pathologies described here, the development of SBMA therapeutics that preserve this activity may be desirable. This approach may be broadly applicable to other polyglutamine diseases such as Huntington’s disease and spinocerebellar ataxias.

overall_design: We used differentiated PC12 cells expressing polyglutamine expanded androgen receptor with baseline, reduced, and enhanced transcriptional activity treated with the androgen dihydrotestosterone (DHT) to identify common and differentially expressed genes. RNA-seq transcriptomic analysis was conducted on triplicate biological replicates for cell type. For comparison 1, control = 112Q and treatment = DBDH. For comparison 2, control = DBDL and treatment = SUMO

first: Jordyn

last: Karliner


title: Case Report: Persistent hypogammaglobulinemia and mixed chimerism after HLA class-II disparate-hematopoietic stem cell transplant

city: Leiden

first: Mirjam

last: van der Burg

address: Albinusdreef 2

department: Pediatrics

country: Netherlands

institute: Leiden University Medical Center

summary: Allogeneic hematopoietic stem cell transplantation (HSCT) is a curative treatment for various hematological, immunological and metabolic diseases, replacing the patient's hematopoietic system with donor-derived healthy hematopoietic stem cells. HSCT can be complicated by early and late events related to impaired immunological recovery such as prolonged hypogammaglobulinemia post-HSCT. We present a 16-year-old female patient with sickle-cell disease who underwent HSCT with stem cells from a human leukocyte antigen (HLA) class-II mismatched family donor. While cellular recovery was good post-HSCT, the patient developed mixed chimerism and suffered from cervical lymphadenopathy, recurrent airway infections and cutaneous SLE. She presented with hypogammaglobulinemia and was started on immunoglobulin substitution therapy and antibiotic prophylaxis. B-cell phenotyping showed that she had increased transitional and naïve mature B cells, reduced memory B cells, and diminished marginal zone/natural effector cells. In-depth immunophenotyping and B-cell receptor repertoire sequencing ruled out an intrinsic B-cell defect by expression of activation-induced cytidine deaminase (AID), presence of somatic hypermutations and differentiation into IgG- and IgA-producing plasma cells in vitro. Immunohistochemistry and flow cytometry of lymph node tissue showed a clear block in terminal B-cell differentiation. Chimerism analysis of sorted lymph node populations showed that exclusively patient-derived B cells populated germinal centers, while only a minor fraction of follicular helper T cells was patient-derived. Because of this discrepancy, we deduced that the HLA class-II disparity between patient and donor impedes facilitation of terminal B-cell differentiation in the lymph node. This case highlights disturbed cognate T-B interactions in the secondary lymphoid organs should be considered when deciphering prolonged hypogammaglobulinemia post-HSCT.

overall_design: Total PBMCs and B-cell subsets sorted from lymph node were used for B-cell receptor repertoire sequencing. Total RNA was isolated and amplified using BIOMED-2 primers, followed by 2x300bp sequencing on the Illumina MiSeq with the Nextera XT Library prep kit (Illumina). Raw reads were germline-allocated using IMGT and filtered for a Phred score of 25. Visualizations were created with Antigen Receptor Galaxy and Graphpad Prism and represent reads without introduced Ns and removal of duplicates based on ‘Top V, Top D, and Top J gene, identical complementarity determining region (CDR) 3-nucleotides’. IGHM reads of PBMCs were subdivided into naïve and MBCs based on having 0-to-2 or more than 2% somatic hypermutations (SHM).

first: Melanie

last: de Gier

first: Ingrid

last: Pico-Knijnenburg

first: Monique

middle: M

last: van Ostaijen-ten Dam

first: Dagmar

last: Berghuis

first: Frans

middle: J

last: Smiers

first: Adriaan

middle: A

last: van Beek

first: Hetty

last: Jolink

first: Patty

last: Jansen

first: Arjan

middle: C

last: Lankester


title: Single cell sequencing of stromal vascular fraction from facial infiltrating lipomatosis and normal facial adipose tissue

status: Public on Jul 06 2024

city: Shanghai

first: Hongrui

last: Chen

address: 639 Zhizaoju Road, Shanghai 200011, P.R.China.

department: Plastic and reconstruction surgery

country: China

institute: Shanghai ninth people's hospital

summary: Facial infiltrating lipomatosis is characterized by excessive growth of adipose tissue, The etiology is associated with somatic PIK3CA variant, but the specific mechanisms are not yet fully understood. In this study, we collected facial adipose tissue from both FIL patients and non-FIL individuals, isolated the stromal vascular fraction (SVF) and performed single-cell transcriptome sequencing on these samples. We mapped out the cellular landscape within the SVF and specifically focused on a deeper analysis of fibro-adipogenic precursor cells (FAPs).

overall_design: Adipose tissues from four FIL patients and three patients underwent facial cutaneous lesions were included in the single- cell sequencing experiments.

first: Bin

last: Sun

first: Chen

last: Hua

first: Xiaoxi

last: Lin


title: KMT2D regulates gene expression through regulating promoters/enhances upon glucose deprivation.

city: Los Angeles

first: Cunyu

last: Wang

state: CA

address: 10833 Le Conte Ave

department: School of Dentistry

institute: university of California, Los Angeles

type: Genome binding/occupancy profiling by high throughput sequencing

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

overall_design: Refer to individual Series


title: Effect of Snf-1 mutation on gene expression profiles in Y. lipolytica

city: Bangkok

first: Pichamon

last: Kiatwuthinon

address: 50 Ngamwongwan Rd. Ladyao Jatujak

department: Biochemistry

country: Thailand

institute: Kasetsart University

summary: To extensively investigate the effect of Snf-1 on n-alkane assimilation in Y. lipolytica, we generated ∆Snf-1 strain using a homologous recombination method. The gene expression profilling analysis between wild-type and ∆Snf-1 strains were compared.

overall_design: Comparative gene expression profiling analysis of RNAseq data from two strains of Y. lipolytica including wild type (CXAU-AI)  and mutant (∆Snf-1)

first: Napapol

last: Poopanitpan


title: Serum exosomes lncRNAs: TCONS_I2_00013502 and ENST00000363624 are new diagnostic markers for rheumatoid arthritis

status: Public on Jul 04 2024

city: Zhanjiang

first: Han

last: Wu

state: Guangdong

address: No.12 Minyou Road, xiashan district, Zhanjiang City

institute: The second Affiliated Hospital of Guangdong Medical University

summary: The lack of diagnostic markers limits the window of effectiveness for rheumatoid arthritis (RA) therapies. Here, we isolated exosomes of serum samples from four distinct groups RA patients, according to disease activity and with/without medication. Then, total RNA of exosomes was extracted for whole-transcriptome sequencing. Focusing on lncRNA sequencing, gene ontology (GO) and kyoto encyclopedia of genes and genomes (KEGG) pathway enrichment analyses were performed. We found that the number of upregulated lncRNAs were significantly higher than that of downregulated lncRNAs in each four RA groups. And most importantly, we identified two specific lncRNAs from differentially expressed lncRNAs, TCONS_I2_00013502 (up-regulated) and ENST00000363624 (down-regulated) in RA. Receiver Operating Characteristic (ROC) curve analysis showed that the two lncRNAs were promising biomarkers for RA diagnosis. These findings highlight lncRNAs of the serum exosome are important biomarkers and provide application potential for diagnosis of RA.

overall_design:
In the exploratory phase, the expression profiles of lncRNAs in exosomes derived from the serum of 13 patients with RA and 5 healthy individuals were investigated through sequencing.
In the validation phase, differentially expressed lncRNAs were selected, and quantitative polymerase chain reaction (qPCR) was performed to validate the findings using serum samples from 32 patients with RA and 32 healthy controls. The concentration of anti-CCP were simultaneously measured by cobas-602. The anti-CCP test result is considered positive when the value exceeds 5 RU/ml.

first: Qiuhua

first: Sijie

first: Chunlong

last: Yang


title: Epitranscriptomic maps of 5-methylcytosine reveal substrate diversity of NSUN methyltransferases and links to mRNA translation and turnover

first: Li

address: 131 Dong-An Road

department: Institutes of Biological Sciences

institute: Fudan University

summary: Multiple epitranscriptomic maps of the RNA modification 5-methylcytosine (m5C) have been prepared, often diverging markedly from each other in terms of site abundance and identity. Differences in detection methods, data depth and analysis pipelines, but also biological factors underly much of this disparity. To address this, we re-analysed available datasets from five human cell lines and seven tissues, generated by the prevailing bisulfite RNA sequencing method, with a coherent m5C site calling pipeline. We used the resulting union list of 6,393 m5C sites called across the broader transcriptome to study site distribution, enzymology, interaction with RNA-binding proteins and molecular function. We confirmed prior observations such as predominance of m5C sites within mRNA exons, enrichment around start codons, and that the tRNA:m5C methyltransferases (MTases) NSUN2 and 6 are the main m5C ‘writers’ for mRNAs. Each NSUN enzyme recognises mRNA features that strongly resemble their respective canonical substrates. Assessing proximity between mRNA m5C sites and footprints of RNA-binding proteins reconfirmed the mRNA export factor ALYREF as an m5C ‘reader’ and identified new candidates for functional interactions, including the RNA helicases DDX3X, involved in mRNA translation, and UPF1, an mRNA decay factor. Experiments in HeLa cells lacking NSUN2 showed that this affected both, steady-state level and UPF1-binding to target mRNAs. Our studies emphasise the emerging diversity of both, m5C writers and readers, affecting mRNA function.

overall_design: To investigate a functional association of UPF1 with m5C,  siRNA-mediated UPF1 knockdown was performed in both WT and NSUN2 KO HeLa cells.

first: Marco

last: Guarnacci

first: Pei-Hong

last: Zhang

first: Thomas

last: Preiss


title: Effect of early-life plane of nutrition on the transcriptional profile  of the anterior pituitary gland in heifer calves

city: Co. Meath

first: Kate

last: Keogh

address: Dunsany

department: Animal and Bioscience Department

country: Ireland

institute: Teagasc Grange

summary: The aim of this study was to examine changes in the transcriptional profile of the anterior pituitary gland of heifer calves offered varying planes of nutrition during the early life period. From approximately 3 weeks of age (19 ± 5 days of age) Angus X Holstein-Friesian heifer calves  were offered either a high (HP, n=13) or moderate (MP, n=15) plane of nutrition to achieve  target growth rates of 1.2 kg/d and 0.5 kg/d, respectively. After 18 weeks of differential feeding all calves were euthanized and anterior pituitary tissue samples harvested from each calf. RNA was extracted and subsequently subjected to RNAseq analysis. A total of 195 genes were identified as significantly differentially expressed between calves on the High plane of nutrition and those on the Moderate plane of nutrition. Biological pathway analysis revealed an enrichement from these genes for processes and pathways related to growth and reproductive system development.

overall_design: 28 anterior pituitary RNA samples were analyzed in total, 13 from High plane of nutrition group and 15 from the Moderate plane of nutrition group.  Calves on the High-plane of nutrition received 1.5 kg of milk replacer (10 liters) per day and ad libitum concentrates during the calf phase. During the subsequent post-weaning phase this group were then offered concentrates ad libitum and hay to appetite. During the calf phase, the Moderate plane of nutrition group received 0.5 kg of milk replacer (4 litres) per day and 1 kg of concentrates, subsequently receiving 1kg of concentrates and hay to appetite during the post-weaning phase. The calf phase lasted for 8 weeks and was followed by the post-weaning phase which lasted for 11 weeks. All samples were collected at the same time-point (21 weeks of age) all differentially expressed genes reported were up- or down-regulated in HP calves compared to MP calves.

first: Alan

middle: K

last: Kelly

first: David

last: Kenny


title: A cyclic peptide-grafted Fc with hepatocyte growth factor functionality ameliorates hepatic fibrosis in a non-alcoholic steatohepatitis mouse model.

city: Nagoya

first: Nishimura

last: Tatsunori

state: Aichi

address: 65 Tsurumai-cho, Showa-ku

department: Graduate School of Medicine

country: Japan

institute: Nagoya University

summary: Although growth factors have significant therapeutic potential because of their regenerative functions, their poor pharmacokinetics and low stability hinder their therapeutic application. Nonalcoholic steatohepatitis (NASH) is characterized by lipid accumulation, inflammation, and fibrosis in the liver, which can progress to life-threatening conditions such as cirrhosis and liver cancer. The hepatocyte growth factor (HGF)-Met receptor pathway has been proposed as a potential therapy for liver diseases including NASH due to its effects on liver regeneration, but the short half-life of recombinant HGF limits its clinical use. Here, we designed a Met agonist with a long half-life by grafting two Met-binding macrocyclic peptides into loops of the crystallizable region (Fc) of immunoglobulin. This surrogate Met agonist ameliorated liver fibrosis and inflammation in a mouse model of NASH, indicating its potential therapeutic use. This study provides a foundation to develop growth factor and cytokine mimetics and to expand their therapeutic applications.

overall_design: RNA-seq analysis of murine hepatic tissue of 2 experimental sets: 1) C57BL/6J inbred male mice (n=20) fed a CDAHF diet for 12 weeks. Then, 2 weeks of HFD and either 4.3 mg/kg Fc  (control n = 10), or 5 mg/kg Fc(mML1)B3 (n = 10) once a week . 2) C57BL/6J inbreed mice (n=16) were fed a CDAHFD for 12 weeks, Then 10 days of  standard diet and either 4.3 mg/kg Fc (control n = 9), or 5 mg/kg of Fc(mML1)B3 (n = 7) at days 1, 4, and 7.

first: Nichole

last: Rojas-Chaverra

first: Ryu

last: Imamura

first: Hiroki

last: Sato

first: Toby

last: Passioura

first: Emiko

last: Mihara

first: Tatsunori

last: Nishimura

first: Junichi

last: Takagi

first: Hiroaki

last: Suga

first: Kunio

last: Matsumoto

first: Katsuya

last: Sakai


title: A spatially resolved multiomic single-cell atlas of soybean development

city: ATHENS

first: Robert

middle: J.

last: Schmitz

state: GA

address: B416 Davison Life Sciences Department of Genetics University of Georgia 120 East Green Street

department: Genetics

institute: University of Georgia

summary: Cis-regulatory elements (CREs) precisely control the spatiotemporal gene expression in cells. Using a spatially resolved single-cell atlas of gene expression and chromatin accessibility across ten soybean tissues, we identified 103 distinct cell types and 303,199 accessible chromatin regions (ACRs). Nearly 40% of ACRs showed cell-type-specific patterns and were enriched for transcription factor (TF) binding motifs controlling cell-type specification and maintenance. We identified non-cell autonomous activity of NIN-LIKE PROTEIN 7 (NLP7), the Nodule Inception (NIN) gene regulatory network and de novo enriched TF motifs in the nodule infected cells. With comprehensive developmental trajectories for endosperm and embryo, we found that 13 sucrose transporters, sharing the DOF11 binding motif, were co-up-regulated in late peripheral endosperm and identified the key embryo cell-type specification regulators during embryogenesis, including a homeobox TF that promotes cotyledon parenchyma identity. This resource provides a valuable foundation for analyzing gene regulatory programs in soybean cell types across tissues and life stages.

overall_design: We generated a comprehensive accessible chromatin and transcriptome atlas across various cell types in soybean with scATAC-seq, snRNA-seq and spatial-RNA-seq(spRNA-seq), where we collected samples from ten major tissues at different stages of the soybean life cycle. These tissues included leaf, hypocotyl, root, nodule, young pod, and five stages of developing seeds: globular stage (GS), heart stage (HS), cotyledon stage (CS), early maturation stage (EMS), and middle maturation stage (MMS). For each tissue, we conducted scATAC-seq and snRNA-seq with at least two replicates.

first: Xuan

first: Ziliang

last: Luo

first: Alexandre

middle: P

last: Marand


title: Overexpression of miR-200s in murine ovarian cancer cells (ID8 and 28-2)

status: Public on Jul 03 2024

city: Guelph

first: Roger

last: Moorehead

address: 50 Stone Rd E

department: Biomedical Sciences

country: Canada

institute: University of Guelph

summary: Little is known about the role of miR-200s in the progression of ovarian cancer. In this study we overexpressed the miR-200c/141 cluster, the miR-200b/200a/429 cluster, or both clusters in two different murine ovarian cancer cell lines (ID8 and 28-2). As co-oeverexpression of both clusters provided the best miR-200 overexpression, we evaluated ID8 and 28-2 cells overexpressing both miR-200 clusters (ID8-200f and 28-2-200f) compared to empty vector control cells (ID8EV and 28-2EV). ID8-200f and 28-2-200f cells showed increased expression of several mesenchymal genes, decreased proliferation, decreased invasion and increased apoptosis compared to their respective controls. RNA sequencing of ID8EV, ID8-200f, 28-2EV and 28-2-200f cells revealed that overexpression of miR-200s primary regulated genes involved in epithelial mesenchymal transition EMT) and gene implicated in migration. Therefore, our work suggests that miR-200s can inhibit characteristics associated ovarian cancer progression (increased proliferation and invasion and promotion of EMT).

overall_design: Both the miR-200c/141 and the miR-200b/200a/429 clusters were overexpressed in ID8 and 28-2 cells creating ID8-200f and 28-2-200f cells. ID8EV and 28-2EV cells were also created by infecting with a emtpy lentiviral vector


title: CRISPR-Cas screen in ILC2 identifies Mef2d as a regulator of GATA3 and type-2 immunity [CRISPR screen]

city: Cambridge

first: Ana

last: Ferreira

address: Francis Crick Avenue, Cambridge Biomedical Campus

country: United Kingdom

institute: MRC - Laboratory of Molecular Biology

summary: Group 2 innate lymphoid cells (ILC2s) orchestrate tissue homeostasis, allergic disorders and anti-helminth protective immunity through their production of cytokines. However, their rarity has hampered high-throughput genetic screening approaches to uncover novel ILC2 regulators in an unbiased manner. We combined expansion of progenitors from multi-reporter mouse strains with optimised ILC differentiation cultures to perform CRISPR-Cas9 screens for regulators of ILC2 development and function. Mef2d emerged as a critical regulator of GATA3 and IL-13, and was essential for type-2 immunity as demonstrated by conditional Mef2d-deficient mice, including a Boolean-ILC2-Cre (BIC) mouse strain developed to enable ILC2-specific gene deletion. Mechanistically, Mef2d bound and transcriptionally repressed the Zc3h12a locus encoding the endonuclease Regnase-1, a negative regulator of IL-33 receptor (ST2) and GATA3 expression, thereby promoting IL-33-mediated ILC2 proliferation and cytokine production. Notably, in ILC2s Mef2d also acted downstream of leukotriene C4-induced calcium-mediated signalling to translocate NFAT1 to the nucleus to promote type-2 gene transcription. These Mef2d-mediated pathways converge and feedback to potentiate type-2 immunity.

overall_design: To identify regulators of GATA3 and IL-13 expression during ILC development and differentiation we performed CRISPR screens using transcription factor and cytokine reporter mice to provide CLPs which were in vitro expanded and transduced with a sgRNA library targeting 1131 transcription factors


title: Proteostasis perturbation of N-Myc by HSP70 inhibition improves treatment in neuroendocrine prostate cancer I

city: Sacramento

first: Clifford

middle: G.

last: Tepper

address: 4645 2nd Avenue, Room 2300A

department: Biochemistry and Molecular Medicine

institute: UC Davis School of Medicine

summary: N-Myc is a key driver of neuroendocrine tumors, such as neuroblastoma and neuroendocrine prostate cancer (NEPC). However, N-Myc is considered undruggable because it lacks clear binding sites for small molecules. One potential way to circumvent this challenge is to identify and target the protein homeostasis (proteostasis) system(s) that maintain N-Myc levels. Here we developed a novel spontaneously indefinite prostate cancer cell line UCDCaP and its paired castration-resistant line UCDCaP-CR, which exhibits neural lineage plasticity and high levels of N-Myc protein expression. Through rapid immunoprecipitation mass spectrometry assay, we revealed that heat shock protein 70 (HSP70) is a top partner for N-Myc. HSP70 is known to function in protein folding and it also coordinates with the E3 ubiquitin ligase, STIP1 homology and U-box containing protein 1 (STUB1), to regulate oncoprotein degradation. We find that HSP70 binds a conserved ‘SELILKR’ motif and prevents the access of STUB1 on N-Myc possibly through steric hindrance. When HSP70’s dwell time on N-Myc is increased by the conformational change of HSP70, STUB1 is in close proximity with N-Myc and becomes functional to promote N-Myc ubiquitination on the K416 and K419 sites of the bHLH-LZ domain and form the poly ubiquitination chains linked by the K11 and K63 sites. Inhibition of HSP70’s ATPase activity by an allosteric inhibitor, JG231, promoted degradation of N-Myc through enhancing STUB1 binding in the nucleus. Notably, JG231 significantly suppressed NEPC tumor growth and limited expression of neuroendocrine related pathways. Guided by a potential bypass route of N-Myc turnover in the cancer cell, we find that dual targeting of Aurora kinase A (AURKA) and HSP70 drastically suppressed N-Myc and tumor growth in NEPC.

overall_design: A total of 3 samples were analyzed in this study. The study focused on the NCI-H660 neuroendocrine prostate cancer cell line. Cell cultures were either treated with vehicle (DMSO) or treated with JG231. Total RNA was then isolated (Qiagen RNeasy MiniKit) and then for RNA-sequencing analysis.

first: Chengfei

last: Liu

first: Joy

last: Yang

middle: G


title: CRISPR-Cas screen in ILC2 identifies Mef2d as a regulator of GATA3 and type-2 immunity [ATAC-Seq]

overall_design: Primary mouse ATAC-seq data is generated using ex vivo expanded mouse ILC2s purified from IL-25 and IL-2/anti-IL-2-treated mice. Genomic accessibility between Il7rCre versus Mef2dIL7RKO mice were compared.


title: B3galt5 regulates obesity and insulin resistance by maintaining the intestinal integrity as a novel PXR target gene [ileum]

status: Public on Jul 02 2024

city: Chengdu

first: Yanping

last: Li

state: Sichuan

address: No.37 Guo Xue Xiang

institute: Sichuan University

summary: Pregnane X receptor (PXR), a xenobiotic receptor involved in drug metabolism, has been reported to regulate lipid and glucose metabolism. The intestine tract is the first inner barrier of the body, which dysfunction contributes to the development of metabolic disorders. However, the role of intestinal PXR in metabolic diseases remains largely unknown. In this study, we showed that activation of PXR by tributyl citrate (TBC), an intestinal-selective agonist of PXR, improved high fat diet (HFD)-induced obesity and insulin resistance. The metabolic benefit of intestinal PXR activation was associated with upregulation of β-1,3 galactosyltransferase 5 (B3galt5). Our results revealed that B3galt5 is mainly expressed in the intestine and is a direct transcriptional target of PXR. Whole-body and intestine-specific knockout of B3galt5 exacerbated HFD-induced obesity, insulin resistance and inflammation. Mechanistically, B3galt5 is essential to maintain the integrity of intestinal mucus barrier. Ablation of B3galt5 impaired the O-glycosylation of mucin2, destabilized the mucus layer, and increased the permeability of intestinal barrier. Furthermore, B3galt5 deficiency abolished the beneficial effect of intestinal PXR activation on metabolic disorders. Our results suggested the intestinal-selective activation of PXR regulated B3galt5 expression holds an important role in maintaining metabolic homeostasis, making it a potential therapeutic strategy in obesity.

overall_design:
To explore the novel target gene of PXR using RNA-seq to analyze the C57BL/6J mice ileum RNA expression
Mice were injected with 40 mg/kg prenenolone-16a-carbonitrile (PCN; in DMSO/corn oil 1:3, Cayman Chemical Company, cat 16343) intraperitoneally every 8 h for three times. Mice were sacrificed 8 h after the last administration and the Ileum tissues were collected for total RNA isolation

first: Jinhang

last: Zhang

first: Ya

last: Huang


title: Microglia-specific IL-10 gene delivery inhibits neuroinflammation and neurodegeneration in a mouse model of Parkinson’s disease

city: Milan

first: Edoardo

middle: Niccolò

last: Bellini

state: Lombardia

address: Via Olgettina 58

department: Neuroscience

country: Italy

institute: San Raffaele Hospital

summary: Neuroinflammation plays a key role in modulating dopaminergic neuronal (DAN) cell loss in Parkinson’s disease (PD). However, it remains unresolved how to effectively normalize this immune response given the complex interplay between the innate and adaptive immune responses occurring within a scarcely accessible organ like the brain. In this study, we uncovered a consistent correlation between neuroinflammation, brain parenchymal lymphocytes and DAN cell loss among several commonly used mouse models of PD generated by a variety of pathological triggers. We validated a viral therapeutic approach for the microglia-specific expression of Interleukin 10 (IL-10) to selectively mitigate the excessive inflammatory response. We found that this approach induced a local nigral IL-10 release that sustained a robust neuroprotection against α-Synuclein (αSYN)-induced neurodegeneration. Single-cell transcriptomics revealed the emergence of a molecularly alternative microglial cell state specifically induced by IL-10 enriched in markers of cell activation with enhanced expression of prophagocytic pathways. Indeed, Il-10 strongly promoted phagocytotic and clearance activities that effectively reduced the αSYN aggregate burden in the nigral area. Furthermore, IL-10 stimulated the differentiation of CD4+ T lymphocytes into active T regulatory cells. Finally, CD8+ T cells exhibited enhanced inhibitory characteristics in the presence of IL-10. Thus, IL-10 local transduction elicited a strong immunomodulation in the nigral tissue with enhanced suppression of lymphocyte toxicity, which rescued DAN cell loss. These results offer insights into the therapeutic benefits induced by IL-10, showcasing a promising gene delivery approach that minimizes undesired side effects and has a strong translational relevance.

overall_design: Nigrostriatal CD45+ cells from mice wt, infected with LV-IL10, LV-SNCA, LV-SNCA/IL10 were isolated by Fluorescence-activated cell sorting (FACS) and analyzed using scRNAseq.

first: Simone

last: Bido


title: Proteostasis perturbation of N-Myc by HSP70 inhibition improves treatment in neuroendocrine prostate cancer II

overall_design: A total of 4 samples were analyzed in this study. The study focused on the PS1172 prostate cancer cell line and the derived PS1172CR castration-resistant (CR) cell line. Duplicate cultures of each cell line were plated for 48 hours. Total RNA was then isolated (Qiagen RNeasy MiniKit) and then submitted for RNA-sequencing analysis.


title: Transcriptomic data from Malus domestica young leaves response to Acibenzolar-S-Methyl (ASM) treatment and nitrogen limitation or resupply.

city: Beaucouzé

first: Mathilde

last: ORSEL

address: 42, rue Georges Morel

country: France

institute: IRHS

summary:
Plant resistance inducers (PRIs) and nitrogen (N) nutrition are both factors known to act on plant defence but their interaction has been little studied and needs further research. The objective of the study was to further characterize this interaction on apple (Malus domestica) plantlets at the molecular level. The produced dataset allows the investigation of the molecular mechanisms underlying:
i) the biotic resistance induced by ASM treatments on apple seedlings,
ii) their response to different nitrogen (N) status (N limitation or subsequent N resupply),
iii) the combination of both N status and ASM treatment.

overall_design: Apple plantlets were grown under contrasted N conditions in order to create 3 different N status (High N reference, N-limited or N-resupplied for 24h after limitation). Plants were then treated with ASM or water (control). Gene expression profiling by RNAseq was performed on young leaf samples collected 2 days after treatments from 30 plants each. Data were obtained from 3 biological replicates for each of the 6 modalities for a total of 18 samples.

first: Terance

last: Mobarak

first: Marie-Noëlle

last: Brisset

first: Mickael

last: Delaire

last: Orsel


title: Cut&Run analysis for transcription factors in MOLT-4 cells

city: Singapore

first: Tze King

last: Tan

address: 14 Medical Drive

department: Cancer Science Institute

country: Singapore

institute: National University of Singapore

summary: Cut&Run analysis was performed in MOLT-4 cells to analyze the DNA bindings of BRD7, BRD9, DPF2 and GLTSCR1 on basal level.

overall_design: MOLT-4 cells were incubated with CUTANA pA/G-Mnase. Chromtin fragments were purified utilizing a Cutana DNA purification kit. Libraries were generated using a NEBNext Ultra II DNA library prep kit and analyzed on Illumina NovaSeq at GENEWIZ Azenta Life Sciences Co. Ltd.

first: Hyoju

last: Kim

first: Shi

middle: H

first: Tze

first: Takaomi

last: Sanda


title: GLUD1 determines murine muscle stem cell fate by controlling mitochondrial glutamate levels

city: Schwerzenbach

first: Adhideb

last: Ghosh

address: Schorenstrasse 16

department: Health Sciences and Technology

country: Switzerland

institute: ETH Zurich

summary: Muscle stem cells (MuSCs) enable muscle growth and regeneration after exercise or injury, but how metabolism controls their regenerative potential is poorly understood. We describe that primary metabolic changes can determine murine MuSC fate decisions. We found that glutamine anaplerosis into the TCA cycle decreases during MuSC differentiation and coincides with decreased expression of the mitochondrial glutamate deaminase GLUD1. Deletion of Glud1 in proliferating MuSCs resulted in precocious differentiation and fusion combined with loss of self-renewal in vitro and in vivo. Mechanistically, deleting Glud1 caused mitochondrial glutamate accumulation and inhibited the malate-aspartate shuttle (MAS). The resulting defect in transporting NADH reducing equivalents into the mitochondria induced compartment-specific NAD+/NADH ratio shifts. MAS activity restoration or directly altering NAD+/NADH ratios normalised myogenesis. In conclusion, GLUD1 prevents deleterious mitochondrial glutamate accumulation and inactivation of the MAS in proliferating MuSCs. It thereby acts as a compartment specific metabolic brake on MuSC differentiation.

overall_design: Sample type and origin: Primary myoblasts isolated from mouse hind-limb mucles were used. Experimental conditions: The data set includes 3 experimental conditions corresponding to 3 different time points during myogenic differentiation in vitro: myoblasts cultured in growth media for 24h (proliferating myoblasts), differentiated myotubes for 48h (end of differentiation step) and fused myotubes (end of fusion step) (see methods for details). Genotypes: myoblasts with 2 different genotypes were used: wild type (used as control) and Glud1 deficient myoblasts. Replicates: at least 3 experimental replicates per condition were included. Except for Glud1-deficient differentiated myoblasts where only 2 replicates were included.

first: Inés

last: Soro-Arnaiz

first: Katrien

last: De Bock


title: Chondrodysplasia-inducing COL2A1 p.Gly1170Ser causes an ER storage defect without associated unfolded protein response in chondronoids

first: Matthew

middle: D

last: Shoulders

state: MA

address: 77 Massachusetts Avenue

department: Chemistry

institute: Massachusetts Institute of Technology

summary: Collagenopathies are a group of clinically diverse disorders caused by defects in collagen folding and secretion. For example, mutations in the gene encoding collagen type-II (COL2A1), the primary collagen in cartilage, can lead to chondrodysplasias of various severities. One example is the Gly1170Ser substitution in procollagen-II, which causes precocious osteoarthritis and Legg-Calvé-Perthes disease. Here, we develop and characterize a novel induced pluripotent stem cell-based cartilage model of this disease, including both hetero- and homozygous genotypes. Biochemical characterization reveals that Gly1170Ser procollagen-II is notably slow to fold and secrete. Instead, procollagen-II accumulates intracellularly, consistent with an endoplasmic reticulum (ER) storage disorder. Intriguingly, though perhaps due to the pathologic substitution occurring within a triple-helical domain that lacks hydrophobic character, this intra-ER protein accumulation is not recognized by cellular stress responses, such as the unfolded protein response. Interactome studies showed that Gly1170Ser procollagen-II interacts to a greater extent with certain ER chaperones and modifying enzymes, consistent with its slow folding. These findings provide mechanistic elucidation into the etiology of this disease. Moreover, the expandable cartilage model developed here provides a valuable platform to rapidly screen and develop therapeutic strategies that can restore procollagen folding and secretion in this collagenopathy and others.

overall_design: To investigate the effect of the Gly1170Ser substitution (both heterozygous and homozygous) in COL2A1 on gene expression in iPSC-derived chondrocytes, heterozygous, homozygous, and their genetically matched wild-type control cells were grown and harvested at an early (day 34) or late (day 44) timepoint.

first: Kathryn

last: Yammine

first: Shireen

middle: R

last: Lamandé


title: Cut&Run analysis for transcription factors in MOLT-4 cells.

summary: Cut&Run analysis was performed in MOLT-4 cells to analyze the basal level of H3K4me3 and DNA binding of RUNX1 and SMARCA4.

first: H

first: T

first: S


title: Cut&Run analysis for transcription factors in DND-41 cells

summary: Cut&Run analysis was performed in DND-41 cells to analyze the basal level of H3K27ac and DNA bindings of BRG1 and RUNX1 after DMSO or ACBI-1 treatment.

overall_design: Treated DND-41 cells were incubated with CUTANA pA/G-Mnase. Chromtin fragments were purified utilizing a Cutana DNA purification kit. Libraries were generated using a NEBNext Ultra II DNA library prep kit and analyzed on Illumina NovaSeq at GENEWIZ Azenta Life Sciences Co. Ltd.


summary: Cut&Run analysis was performed in MOLT-4 cells to analyze the level of H3K27ac and DNA bindings of SMARCA4 and RUNX1 after DMSO or BRM014 treatment.

overall_design: Treated MOLT-4 cells were incubated with CUTANA pA/G-Mnase. Chromtin fragments were purified utilizing a Cutana DNA purification kit. Libraries were generated using a NEBNext Ultra II DNA library prep kit and analyzed on Illumina NovaSeq at GENEWIZ Azenta Life Sciences Co. Ltd.


title: CRISPR-Cas screen in ILC2 identifies Mef2d as a regulator of GATA3 and type-2 immunity [ChIP-Seq]

first: Ana

last: Ferreira

overall_design: Primary mouse ILC2 ChIP-seq data is generated using ex vivo expanded mouse ILC2s purified from IL-25 and IL-2/anti-IL-2-treated mice. Binding profiles of Mef2d, Gata3 and Nfat1 during unstimulated versus LTC4 stimulation were investigated.


title: RNA-seq analysis after 24hr of ACBI-1 treatment on DND-41 and MOLT-4 cells.

summary: RNA-seq analysis was performed in two T-ALL cell lines (DND-41 and MOLT-4) to analyze gene expression changes after 24 hours of 100nM ACBI-1 or DMSO treatment.

overall_design: DND-41 and MOLT-4 cells were treated with 100nM ACBI-1 or DMSO for 24 hours. Total RNA was harvested and RNA-seq analysis was performed.


title: H3K27ac Hi-ChIP analysis in MOLT-4 and DND-41 cells.

summary: H3K27ac Hi-ChIP analysis was performed in MOLT-4 and DND-41 cells to analyze active chromatin-chromatin interactions after DMSO or ACBI-1 treatment.

overall_design: The cells were crosslinked with formaldehyde for 10 min. DNA was enriched by chromatin-immunoprecipitation (ChIP), tagmented with Tn5, subjected for library construction, and sequenced by the DNBseq platform.  Hi-ChIP was performed using an antibody against H3K27ac (Abcam: ab4729).


title: B3galt5 regulates obesity and insulin resistance by maintaining the intestinal integrity as a novel PXR target gene

first: Yanping

last: Li

overall_design:
To explore the novel target gene of PXR using RNA-seq to analyze the C57BL/6J mice colon RNA expression
Mice were injected with 40 mg/kg prenenolone-16a-carbonitrile (PCN; in DMSO/corn oil 1:3, Cayman Chemical Company, cat 16343) intraperitoneally every 8 h for three times. Mice were sacrificed 8 h after the last administration and the colon tissues were collected for total RNA isolation

first: Jinhang

last: Zhang

first: Ya

last: Huang


title: ATAC-seq analysis for T-ALL cell lines

first: Tze King

summary: ATAC-seq analysis was performed in multiple T-ALL cell lines to analyze chromatin accessibility.

overall_design: The cells are incubated with Tn5 transposase. Tagmented DNA fragments are purified with Zymo DNA purification kit. ATAC libraries are purified and sequenced on Illumina NovaSeq at GENEWIZ Azenta Life Sciences Co. Ltd.


title: RNA-seq analysis after 3hr of ACBI-1 treatment on DND-41 and MOLT-4 cells.

summary: RNA-seq analysis was performed in two T-ALL cell lines (DND-41 and MOLT-4) to analyze gene expression changes after 3 hours of 100nM ACBI-1 or DMSO treatment.

overall_design: DND-41 and MOLT-4 cells were treated with 100nM ACBI-1 or DMSO for 3 hours. Total RNA was harvested and RNA-seq analysis was performed.


title: RNA-seq analysis after 8hr of ACBI-1 treatment on DND-41 and MOLT-4 cells.

summary: RNA-seq analysis was performed in two T-ALL cell lines (DND-41 and MOLT-4) to analyze gene expression changes after 8 hours of 100nM ACBI-1 or DMSO treatment.

overall_design: DND-41 and MOLT-4 cells were treated with 100nM ACBI-1 or DMSO for 8 hours. Total RNA was harvested and RNA-seq analysis was performed.


title: A CRISPR/Cas9 screen in embryonic stem cells reveals that Mdm2 regulates totipotency exit [CUT&Tag]

status: Public on Jul 01 2024

city: xiamen

first: Qingxiang

last: Gao

address: Xiang'an south way

institute: Xiamen University

summary: During early embryonic development, the transition from totipotency to pluripotency is a fundamental and critical process for proper development. However, the regulatory mechanisms governing this transition remain elusive. In this study, we conducted a comprehensive genome-wide CRISPR/Cas9 screen to investigate the 2-cell-like cells (2CLCs) phenotype in mouse embryonic stem cells (mESCs). This effort led to the identification of ten regulators that play a pivotal role in determining cell fate during this transition. Notably, our study revealed Mdm2 as a significant negative regulator of 2CLCs, as perturbation of Mdm2 resulted in a higher proportion of 2CLCs. Mdm2 appears to influence cell fate through its impact on cell cycle progression and H3K27me3 epigenetic modifications. In summary, the results of our CRISPR/Cas9 screen have uncovered several genes with distinct functions in regulating totipotency and pluripotency at various levels, offering a valuable resource for potential targets in future molecular studies.

overall_design: Identifiy the  transcriptome and H3K27me3 modification changes in different mouse embryonic stem cells

first: Chen

first: Xin


title: ATAC-seq analysis for DND-41 and MOLT4 cells with ACBI-1 treatment

summary: ATAC-seq analysis was performed in two T-ALL cell lines (DND-41 and MOLT-4) with either DMSO or ACBI-1 treatment at two different time points (3hr and 24hr).


title: DNA hydrogel based Wound Hemostatic Adjuvant

city: Hangzhou

first: Tianyi

last: Gu

address: No.268, Kaixuan Road

institute: Zhejiang University

summary: Severe traumatic bleeding may lead to extremely high mortality rates, and early intervention to stop bleeding plays as a critical role in saving lives. However, rapid hemostasis in deep non-compressible trauma using a highly water-absorbent hydrogel, combined with strong tissue adhesion and bionic procoagulant mechanism, remains a challenge. In this study, a DNA hydrogel (DNAgel) network composed of natural nucleic acids with rapid water absorption, high swelling and instant tissue adhesion is reported, like a “band-aid” to physically stop bleeding. The excellent swelling behavior and robust mechanical performance, meanwhile, enable the DNAgel band-aid to fill the defect cavity and exert pressure on the bleeding vessels, thereby achieving “compression hemostasis” for deep tissue bleeding sites. The DNAgel network also acts as an artificial DNA scaffold for erythrocytes to adhere and aggregate, and activates platelets, promoting coagulation cascade in a bionic way. The DNAgel achieves lower blood loss than commercial gelatin sponge (GS) in rat trauma models. In vivo evaluation in a full-thickness skin incision model also demonstrated the ability of DNAgel for promoting wound healing. Overall, the DNAgel band-aid with great hemostatic capacity is a promising candidate for rapid hemostasis and wound healing.

overall_design: The fresh whole blood of rats was collected and incubated with DNAgel and GS for 15 min, respectively. The high-throughput RNA sequencing (RNA-seq) and transcriptome analysis were performed by LC-BIO Bio-tech Ltd (Hangzhou, China). Bioinformatic analysis was performed using the OmicStudio tools at https://www.omicstudio.cn. The volcano plot, heatmap and GSEA plot were drawn based on the R version 4.1.3 (2022-03-10) on the OmicStudio platform.


title: A CRISPR/Cas9 screen in embryonic stem cells reveals that Mdm2 regulates totipotency exit [RNA-Seq]


title: Oncogenic roles of SWI/SNF chromatin remodeling factors: therapeutic targets in T-cell acute lymphoblastic leukemia

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

overall_design: Refer to individual Series


title: Flow-Mediated Modulation of the Endothelial Cell Lipidome

first: Soon-Gook

last: Hong

state: California

address: 630 Charles E Young Dr S

department: Medicine

institute: UCLA

summary: The luminal surface of the endothelium is exposed to dynamic blood flow patterns that are known to affect endothelial cell phenotype. In this study, human aortic endothelial cells (HAECs) were exposed to unidirectional laminar flow (20 dynes/cm2) or disturbed flow (±4 dynes/cm2, 2 Hz) for 48 hrs and then subjected to RNA sequencing.  The sequencing data was used to analyze changes in endothelial cell gene expression and pathways associated with lipid metabolism.

overall_design:
To investigate the effect of different flow patterns on endothelial cell lipidome, primary human aortic endothelial cells (HAECs) were seeded in ibidi µ-Slide 0.2 (ibidi #80166) or 0.6 (ibidi #80186) Luer ibiTreat. Unidirectional laminar flow (20 dynes/cm2) or disturbed flow (±4 dynes/cm2, 2 Hz) was applied to confluent monolayers using the ibidi pump system (ibidi #10902) for 48 hrs.
We then performed gene expression profiling analysis using data obtained from RNA sequencing of three replicates per treatment condition.
Comparative gene expression profiling analysis of RNA-seq data for UF-exposed HAECs and DF-exposed HAECs.

first: Julia

last: Mack


title: Hippo pathway-mediated YAP1/TAZ inhibition is essential for proper pancreatic endocrine specification and differentiation

city: bethesda

first: Efsun

last: Arda

state: Maryland

address: 41 medlars drive

institute: National Institutes of Health

summary: In order to investigate the effect of Hippo pathway in pancreatic endorince specification and differentiaion, single nucleus RNA seq was performed using mouse pancreatic tissues with Lats1&2 deletion in NGN3+ cells (NL mice). Wild type mice pancreatic tissues were also subject to same analysis as a control.

overall_design: Single nucleus RNA sequencing (snRNA-seq) analysis was performed using E17.5 pancreata from 1 control and 2 NL mice.

first: Wu

last: Yifan

first: Qin

last: Kunhua

first: Xu

last: Yi

first: Rajhans

last: Shreya

first: Lopez

last: Kevin

first: Liu

last: Jun

first: Nipper

last: Michael

first: Deng

last: Janice

first: Yin

last: Xue

first: Ramjit

last: Logan

first: Ye

last: Zhengqing

first: Luan

last: Yu

first: Pei

last: Wang


title: Aging Promotes Metabolic Dysfunction-associated Steatotic Liver Disease and Multi-Organ Dysfunction by Inducing Ferroptotic Stress

city: Durham

first: Liuyang

state: NC

address: 213 Research Drive

institute: Duke University

summary: Aging is an inevitable consequence of living that undermines cellular resiliency to cause tissue degeneration and eventual multi-organ dysfunction. Susceptibility to biological consequences of aging varies among organs and individuals. Stressors that challenge resiliency unmask these differences. Hence, aging increases the risk for failure of many metabolically-stressed organs and exacerbates liver degeneration related to obesity and diabetes. We discovered that aging promotes ferroptosis, a type of metabolism-regulated cell death, in hepatocytes. Hepatocytes that have launched the ferroptotic death program adapt their metabolism to survive but become damaged and dysfunctional. Metabolic stressors amplify this, increasing the burden of ferroptosis-adapted cells and liver damage. Blocking ferroptotic signaling in old livers reduces accumulation of adapted hepatocytes and reverts livers to a more youthful resilient state despite exogenous stressors. Ferroptosis is also induced in other organs that are damaged by chronic metabolic stress, identifying ferroptosis as a tractable conserved mechanism for aging-related multi-organ dysfunction.

overall_design: To determine the role of ferroptotic stress and senescence during diet-induced MASH, young and old mice were fed with a chow diet or CDA-HFD diet for 6 weeks, and injected with ferrostatin1. Bulk RNA-seq are performed.

first: Kuo

last: Du

first: Anna

last: Diehl


title: Microglial heterogeneity in the ischemic stroke mouse brain of both sexes

first: Wei

last: Yang

state: United States

address: 300 Duke Street

summary: The objective of the current study was to dissect the effect of permanent stroke on the diversity of microglia, the resident immune cells within the brain parenchyma.

overall_design: To characterize microglial diversity after permanent stroke in Tmem119-2A-CreERT2;Ai6 mice in which microglia were labeled with ZsGreen, we sorted ZsGreen positive cells on day 3 after stroke for scRNA-seq analysis. Four samples were analayzed: female_sham, female_stroke, male_sham, and male_stroke,  (F_Sham, F_Stroke, M_Sham, M_Stroke). Each sample were pooled from 4 mice.

first: Angela

last: del Aguila

first: Vaibhav

last: Jain


title: Gamma Delta T Cells Mediate Shrimp Allergy Through Regulatory Cytokine Pathways

city: Houston

first: Hongjie

last: Li

state: Texas

address: N910 Alkek Building

department: Huffington Center on Aging

institute: Baylor College of Medicine

summary: Shrimp allergy is the second most common food allergy in the United States. γδ T cells play a regulatory role in peanut immunotherapy, but their role in shrimp allergy remains unclear. We hypothesized γδ T cells play a regulatory role in shrimp allergic disease. We performed single cell RNA sequencing on peripheral cells from shrimp allergic (SA) and healthy control (HC) subjects after stimulation with shrimp tropomyosin. We found significant expansion of γδ T cells and three distinct clusters. One γδ T cell cluster predominated in SA, characterized as CD8+ with a cytotoxic expression profile. We found significant upregulation of TGF-β1 and downregulation of IL-7R in SA-stimulated vs. HC-stimulated γδ T cells, and IL-10 secretion in stimulated SA γδ T cells. γδ T cells play an important role in the pathogenesis of shrimp allergy through lymphocyte-mediated cytotoxin signaling and cytokine-mediated signaling pathways, including TGFβ-1, IL7/TSLP-IL7R, and IL10-IL10R pathways.

overall_design: Single-cell RNA sequencing of unstimulated (tropomyosin: 0 ug/mL) and stimulated (tropomyosin: 1 ug/mL) healthy and shrimp allergic patients

first: Tyler

last: Jackson

first: Carla

last: Davis

first: Bin

last: Su


title: Single cell gene expression analysis of hESCs undergoing neuronal differentiation

city: Oslo

first: Mari

last: Spildrejorde

state: OSLO

address: Sognsvannsveien 9

department: Research and Innovation, Neuroscience research unit

country: Norway

institute: Oslo University Hospital

summary: Single cell genome-wide gene expression profiling of human embryonic stem cells (hESCs) undergoing neuronal differentiation at four distinct time-points. hESCs are differentiated towards neuronal cells for 13 days, and sampled for downstream analysis at day 0, 6, 10 and 13 . Cells exposed to 50, 100, 200 and 400 nM citalopram from Day 1 were charachterised at Day 6, 10 and 13.

overall_design:
Genome wide gene expression profiling of (n= 16) samples from human hESCs at day 0, and day 6, 10 and 13 of neuronal differentiation
Submitters state "raw data is not provided due to data sensitivity. We have used embryonic stem cells and there is a possibility of tracing back data to the donor, thus we are not allowed to share raw data".

first: Magnus

last: Leithaug

first: Athina

last: Samara

first: Hans

last: Aass

first: Ankush

last: Sharma

first: Ganesh

last: Acharya

first: Hedvig

last: Nordeng

first: Kristina

last: Gervin

first: Robert

last: Lyle


title: Epigenome analysis of hESCs undergoing neuronal differentiation

type: Methylation profiling by genome tiling array

summary: Genome wide DNA methylation profiling of human embryonic stem cells of hESCs undergoing neuronal differentiation at four distinct time-points. hESCs are differentiated towards neuronal cells for 13 days, and sampled for downstream analysis at day 0, 6, 10 and 13 . Cells exposed to 50, 100, 200 and 400 nM citalopram from Day 1 were charachterised at Day 6, 10 and 13. The Illumina Infinium MethylationEPIC_v-1-0_B3 was used to obtain DNA methylation profiles accross approximately 850k CpGs in differentiating cells.

overall_design:
Bisulphite converted DNA from the 89 samples were hybridised to the Illumina Infinium MethylationEPIC BeadChip
Submitters state "raw data IDAT files are not provided due to data sensitivity. We have used embryonic stem cells and there is a possibility of tracing back data to the donor, thus we are not allowed to share raw data".


title: The immunosuppressive Tuberculosis-associated microenvironment inhibits viral replication and promotes HIV-1 latency in CD4+ T cells

city: Westmead

first: Gabriel

last: Duette

state: NSW

address: 176 Hawkesbury Road

department: Centre for Virus Research

country: Australia

institute: The Westmead Institute for Medical Research

summary: Mycobacterium tuberculosis (Mtb), the causative agent of tuberculosis (TB), is the most common coinfection among people living with HIV-1. This coinfection alters the efficacy of the immune response against both HIV-1 and Mtb, and is associated with accelerated HIV-1 disease progression and reduced survival. Enhanced HIV-1 replication in macrophages induced by Mtb coinfection may contribute to the worsened clinical outcomes observed in HIV-1/TB coinfected individuals. However, the impact of the HIV-1/TB coinfection on HIV-1 replication and latency in CD4+ T cells remains poorly studied.  In this study, we used the acellular fraction of tuberculous pleural effusion (TB-PE) as a proxy for the microenvironment generated by Mtb infection. Using this physiologically relevant fluid, we investigated whether viral replication and HIV-1 latency in CD4+ T cells are affected by a TB-associated microenvironment. Interestingly, our results revealed that TB-PE shaped the transcriptional profile of CD4+ T cells impairing T cell receptor-dependent cell activation and decreased HIV-1 replication. Moreover, this immunosuppressive TB microenvironment promoted viral latency and inhibited HIV-1 reactivation in CD4+ T cells from people living with HIV-1.  This study indicates that the immune response induced by TB may contribute to the persistence of the viral reservoir by silencing HIV-1 expression in individuals coinfected with both pathogens, allowing the virus to persist undetected by the immune system and increasing the size of the HIV-1 latent reservoir in cells at the site of the coinfection.

overall_design: To understand the effect of the immune response to TB on CD4+ T cells and its potential impact on HIV-1 replication, we assessed the transcriptional profile of primary CD4+ T cells treated with TB-PE. Bulk RNA sequencing (RNAseq) was performed on primary CD4+ T cells isolated from 3 healthy donors and treated with TB-PE. In addition, since T cell activation increases CD4+ T cells susceptibility to HIV-1 infection, we also investigated the impact of TB-PE on CD4 T cells activated by anti-CD3/CD28 antibodies.

first: Joey

last: Lai

first: Brian

last: Gloss

first: Samantha

last: Cronin

first: Sarah

last: Palmer


title: Andes virus mRNA vaccines: comparison of unmodified and modified mRNA platforms [VDJ-seq]

city: Nashville

first: Ivelin

last: Georgiev

state: TN

address: 11475 MRB IV 2213 Garland Ave

department: Pathology, Microbiology, and Immunology

institute: Vanderbilt University Medical Center

summary: Andes virus (ANDV) is a rodent-borne zoonotic orthohantavirus endemic in South America that causes hantavirus pulmonary syndrome in humans, with up to a 40% case fatality rate. We developed ANDV mRNA vaccines based on the M segment of the viral genome that codes for glycoproteins Gn and Gc in a single open reading frame of glycoprotein precursor (GPC). We generated RNAs either with regular uridine (U-mRNA) or N1-methylpseudouridine (m1Ψ-mRNA). Mice immunized by either ANDV U-mRNA or m1Ψ-mRNA developed similar germinal center responses in lymph nodes. Single cell RNA and BCR sequencing of germinal center B cells from vaccinated mice demonstrated similar levels of activation, except an additional cluster of cells exhibiting strong interferon response that was present in animals vaccinated with U-mRNA but not m1Ψ-mRNA. Furthermore, similar immunoglobulin class-switching and somatic hypermutations were observed for the two vaccines. Golden Syrian hamsters were immunized intramuscularly with 2 doses of the vaccines on days 0 and 21. The titers of Gn/Gc-binding antibodies were moderately greater for U-mRNA construct than for m1Ψ-mRNA construct, however, the titers of ANDV-neutralizing antibodies were equivalent. Vaccinated animals were challenged with a lethal dose of ANDV at 21 days after the boost, along with the naïve control group. All control animals succumbed to infection whereas all vaccinated animals survived without any detectable disease or viral load. The data demonstrate the development of effective vaccines against ANDV and the lack of a significant effect of m1Ψ mRNA modification on immunogenicity and protection in the hamster model.

overall_design: Ten-week-old BALB/c mice, four per group, were immunized intramuscularly with one dose of 25 µg of either U-mRNA or m1Ψ-mRNA formulated with LNP. Twelve days after immunization animals were euthanized, and draining inguinal and popliteal lymph nodes harvested and pooled for each mouse for cell sorting. Germinal center B cells were sorted and sequenced to generate single cell 5' gene expression and V(D)J libraries.

first: Ivan

middle: V

last: Kuzmin

first: Ruben

last: Soto Acosta

first: Perry

last: Wasdin

first: Chad

last: Mire

first: Taylor

middle: B

last: Engdahl

first: Woohyun

last: Moon

first: Vsevolod

last: Popov

first: James

last: Crowe

middle: S

first: Mariano

last: Garcia-Blanco

last: Abbott

first: Alexander

last: Bukreyev


title: Gene expression and Epigenome analysis of hESCs undergoing neuronal differentiation

first: Mari

last: Spildrejorde


summary:
Comparison of unmodified and modified mRNA platforms for Andes virus mRNA vaccine in rodent models
Kuzmin IV, Soto-Acosta R, Pruitt L, Wasdin PT, Kedarinath K, Hernandez KR, Gonzales KA, Hill K, Weidner NG, Mire C, Engdahl TB, Moon WJ, Popov V, Crowe JE, Georgiev IS, Garcia-Blanco MA, Abbott RK, Bukreyev A.

This SuperSeries is composed of the SubSeries listed below.


title: Andes virus mRNA vaccines: comparison of unmodified and modified mRNA platforms [scRNA-seq]

middle: E

first: Robert


title: Gene expression analysis of hESCs undergoing neuronal differentiation exposed to citalopram

summary: Genome-wide gene expression profiling of human embryonic stem cells (hESCs) undergoing neuronal differentiation at four distinct time-points. hESCs are differentiated towards neuronal cells for 13 days, and sampled for downstream analysis at day 0, 6, 10 and 13 . Cells exposed to 50, 100, 200 and 400 nM citalopram from Day 1 were charachterised at Day 6, 10 and 13.

overall_design:
Genome wide gene expression profiling of (n= 89) samples from human embryonic stem cells (hESCs) at day 0, 6, 10 and 13 of neuronal differentiation. Cells were exposed to citalopram from Day 1 and onwards.
Submitters state "raw data is not provided due to data sensitivity. We have used embryonic stem cells and there is a possibility of tracing back data to the donor, thus we are not allowed to share raw data".

first: Magnus

last: Leithaug

first: Athina

last: Samara

first: Hans

last: Aass

first: Ankush

last: Sharma

first: Ganesh

last: Acharya

first: Hedvig

middle: M

last: Nordeng

first: Kristina

last: Gervin

last: Lyle


title: KMT2D regulates Fanconi anemia pathway genes through regulating promoters/enhances upon glucose deprivation (CHIPseq)

city: Los Angeles

first: Cunyu

last: Wang

address: 10833 Le Conte Ave

department: School of Dentistry

institute: university of California, Los Angeles

summary: Histone H3 lysine (H3K4) methyltransferase KMT2D is a key regulator of gene expression, mainly through promoting H3K4 methylation and activating enhancers, and plays critical roles in development, differentiation, metabolism, and tumor suppression. Here, our study showed that KMT2D was recruited to the enhancer regions of several Fanconi anemia (FA) pathway genes, such as ATR, FANCM, REV3L, and TOP3A, in response to glucose deprivation. Notably, KMT2D loss led to significant impairment of promoter/enhancer signals, as evidenced by decreased H3K4me1, H3K4me3, and H3K27ac signals, on several FA pathway genes. Our ChIP-Seq results demonstrated that the inactivation of promoters/enhancers due to KMT2D loss is a key contributor to the downregulation of FA pathway genes in glucose-deprived KMT2D-deficient HNSCC.

overall_design: Chromatin binding profiling of KMT2D in wild-type SCC23 HNSCC cells with glucose deprivation treatment for 6 hours (-Glc). Chromatin binding profiling of histone modification H3K4me1, H3K27ac, H3K4me3, H3K9me3, and H3K27me3 in WT and KMT2D knockout (KMT2D-KO) SCC23 HNSCC cells under glucose deprivation treatment for 6 hours (-Glc).

first: Jing

first: Wei

last: Liu


title: An integrated ’omics approach highlights the role of epigenetic events to explain and predict response to neoadjuvant chemotherapy and bevacizumab in breast cancer

first: Thomas

last: Fleischer

address: Ullernchausséen 70

department: Department of Cancer Genetics, Institute for Cancer Research

summary: DNA methylation profiling (Illumina Infinium 450K) was performed on fresh frozen tumor biopsies taken before, during and after treatment from patients receiving neoadjuvant chemotherapy with or without the anti-angiogenic drug bevacizumab

overall_design: Bisulphite converted DNA from 290 samples from 132 patients were hybridised to the Illumina Infinium 150k Human Methylation Beadchip

first: Mads

last: Haugen

first: Jørgen

last: Ankill

first: Laxmi

last: Silwal-Pandit

first: Anne-Lise

last: Børresen-Dale

first: Ingrid

last: Hedenfalk

last: Hatschek

first: Jörg

last: Tost

first: Olav

last: Engebraaten

first: Vessela

middle: N

last: Kristensen


title: Whole exome sequencing of HNSCC PDX with KMT2D-inactivating mutation

summary: Histone H3 lysine (H3K4) methyltransferase KMT2D is a key regulator of gene expression, mainly through promoting H3K4 methylation and activating enhancers, and plays critical roles in development, differentiation, metabolism, and tumor suppression. Our study found that KMT2D-deficient HNSCC is sensitive to 2-DG plus DNA damage agents or 2-DG plus PAPR inhibitors in HNSCC mouse models and human cell lines. To further investigate the therapeutic potential of these treatments, we aimed to use patient-derived xenograft (PDX) with MT2D-inactivating utations. We sequenced one PDX from the University of Colorado School of Medicine by whole exome sequencing which has the nonsense mutation of KMT2D.

overall_design: High-quality genomic DNA was extracted from PDX and whole exome sequencing was performed.


title: Identification of VDR target genes in intestinal epithelial cells

city: Herlev

first: Lauge

last: Kellermann

state: --- Select ---

address: Borgmester Ib Juuls Vej 1

department: Department of Gastroenterology

country: Denmark

institute: Herlev Hospital


title: RNA-Seq: The transcriptional response to 1,25(OH)2D3 in human colonic organoids

summary: Colonic organoids generated from one healthy individual were treated with 1,25(OH)2D3 or vehicle for 6 and 24h. Differential expressed genes are compared across treatments.

overall_design: Colonic organoids were generated from one healthy individual and treated with 1µM 1,25(OH)2D3 or vehicle (ethanol) for 6h and 24h in triplicates for all treatments and time points.

first: Marianne

middle: T

last: Pedersen

first: Jens

last: Johansen


title: Vitamin D receptor (VDR) ChIP-Seq on human colonic organoids stimulated with 1,25(OH)2D3

summary: Genome-wide VDR chromatin binding sites in colonic organoids stimulated with 1,25(OH)2D3 for 2h.

overall_design: Colonic organoids derived from a healthy control individual were stimulated with 1,25(OH)2D3 or vehicle (ethanol) for 2h. Two independent ChIP seq experiments (ChIP A and B) were preformed with organoids from the same donor.


title: Gene regulation profiling by KMT2D under different glucose conditions in HNSCC (RNAseq)

summary: Histone H3 lysine (H3K4) methyltransferase KMT2D is a key regulator of gene expression, mainly through promoting H3K4 methylation and activating enhancers, and plays critical roles in development, differentiation, metabolism, and tumor suppression. To investigate the mechanisms by which KMT2D loss promotes HNSCC and to identify potential therapeutic targets, we generated KMT2D wild-type (KMT2D-WT) and KMT2D knock-out (KMT2D-KO) SCC23 HNSCC cells and performed RNA-seq under different glucose conditions.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for KMT2D-WT and KMT2D-KO SCC23 HNSCC cells under high glucose conditions (+Glc) or with glucose deprivation treatment for 6 hours (-Glc).


title: Astrocytic autophagy plasticity modulates Aβ clearance and cognitive function in Alzheimer’s disease

status: Public on Jun 30 2024

city: Seoul

first: Hoon

last: Ryu

state: Seoul

address: Seoul

department: Brain Science Institute

country: South Korea

institute: Korea Institute of Science and Technology

summary:
Astrocytes, one of the most resilient cells in the brain, transform into reactive astrocytes in response to toxic proteins such as amyloid beta (Aβ) in Alzheimer’s disease (AD). However, reactive astrocyte-mediated non-cell autonomous neuropathological mechanism is not fully understood yet. We aimed our study to find out whether Aβ-induced proteotoxic stress affects the expression of autophagy genes and the modulation of autophagic flux in astrocytes, and if yes, how Aβ-induced autophagy-associated genes are involved Aβ clearance in astrocytes of animal model of AD.
Whole RNA sequencing (RNA-seq) was performed to detect gene expression patterns in Aβ-treated human astrocytes in a time-dependent manner. To verify the role of astrocytic autophagy in an AD mouse model, we developed AAVs expressing shRNAs for MAP1LC3B/LC3B (LC3B) and Sequestosome1 (SQSTM1) based on AAV-R-CREon vector, which is a Cre recombinase-dependent gene-silencing system. Also, the effect of astrocyte-specific overexpression of LC3B on the neuropathology in AD (APP/PS1) mice was determined. Neuropathological alterations of AD mice with astrocytic autophagy dysfunction were observed by confocal microscopy and transmission electron microscope (TEM). Behavioral changes of mice were examined through novel object recognition test (NOR) and novel object place recognition test (NOPR).
Here, we show that astrocytes, unlike neurons, undergo plastic changes in autophagic processes to remove Aβ. Aβ transiently induces expression of LC3B gene and turns on a prolonged transcription of SQSTM1 gene. The Aβ-induced astrocytic autophagy accelerates urea cycle and putrescine degradation pathway. Pharmacological inhibition of autophagy exacerbates mitochondrial dysfunction and oxidative stress in astrocytes. Astrocyte-specific knockdown of LC3B and SQSTM1 significantly increases Aβ plaque formation and hypertrophic reactive astrocytes in APP/PS1 mice, along with a significant reduction of neuronal marker and cognitive function. In contrast, astrocyte-specific overexpression of LC3B reduced Ab aggregates in the brain of APP/PS1 mice An increase of LC3B and SQSTM1 protein is found in astrocytes of the hippocampus in AD patients.
Taken together, our data indicates that Aβ-induced astrocytic autophagic plasticity is an important cellular event to modulate Aβ clearance and maintain cognitive function in AD mice.

overall_design: Whole RNA sequencing to detect gene expression patterns in Aβ-treated human astrocytes in a time-dependent manner

first: Justin

last: Lee

first: Junghee


title: Annelid methylomes reveal ancestral developmental and ageing-associated epigenetic erosion across Bilateria

city: London

first: Jose

last: Martin-Duran

address: Mile End road

institute: Queen Mary University of London

type: Methylation profiling by high throughput sequencing

summary:
Background
DNA methylation in the form of 5-methylcytosine (5mC) is the most abundant base modification in animals. However, 5mC levels vary widely across taxa. Whilst vertebrate genomes are hypermethylated, in most invertebrates, 5mC concentrates on constantly and highly transcribed genes (gene body methylation; GbM) and, in some species, on transposable elements (TEs), a pattern known as ‘mosaic’. Yet, the role and developmental dynamics of 5mC and how these explain interspecific differences in DNA methylation patterns remain poorly understood, especially in Spiralia, a large clade of invertebrates comprising nearly half of the animal phyla.
Results
Here, we generate base-resolution methylomes for three species with distinct genomic features and phylogenetic positions in Annelida, a major spiralian phylum. All possible 5mC patterns occur in annelids, from typical invertebrate intermediate levels in a mosaic distribution to hypermethylation and methylation loss. GbM is common to annelids with 5mC, and methylation differences across species are explained by taxon-specific transcriptional dynamics or the presence of intronic TEs. Notably, the link between GbM and transcription decays during development, alongside a gradual and global, age-dependent demethylation in adult stages. Additionally, reducing 5mC levels with cytidine analogues during early development impairs normal embryogenesis and reactivates TEs in the annelid Owenia fusiformis.
Conclusions
Our study indicates that global epigenetic erosion during development and ageing is an ancestral feature of bilateral animals. However, the tight link between transcription and gene body methylation is likely more important in early embryonic stages, and 5mC-mediated TE silencing probably emerged convergently across animal lineages.

sample_organism:
Owenia fusiformis
Capitella teleta
Dimorphilus gyrociliatus

overall_design: Methylomes of three annelid species (O. fusiformis, C. teleta and D. gyrociliatus) at different developmental timepoints to investigate the 5mC dynamics during embryogenesis and the life cycle of this animals using WGBS, EM-seq and Nanopore sequencing. To investigate the impact of DNA methylation during development, we combined cytidine analogs, RNA-seq and EM-seq.

first: Kero

last: Guynes

first: Luke

middle: A

last: Sarre

first: Allan

last: Carrillo-Baltodano

first: Billie

last: Davies

first: Lan

last: Xu

first: Yan

last: Liang

first: Francisco

last: Martín-Zamora

first: Paul

middle: J

last: Hurd

first: Alex

last: de Mendoza

first: José

last: Martín-Durán


title: Vascular and renal response to neonatal hyperoxia in 1-year old rats

city: Miami

first: Augusto

middle: F

last: Schmidt

state: FL

address: 1611 NW 12th ave

institute: University of Miami Miller School of Medicine

summary: We evaluated the transcriptional changes in the aorta and kidney of 1 year old rats that were exposed to hyperoxia or normoxia in the neonatal period.

overall_design: Newborn rats were randomly assigned to normoxia or hyperoxia (85%) from postnatal day 1 to postnatal day 14

first: Merline

last: Benny

first: Marissa

last: deFreitas


title: Total RNA-seq using mouse tissues from Cyp2e1 +/+ mice upon pair-fed, Cyp2e1 +/+ mice upon EtOH-fed, Cyp2e1 -/- mice upon pair-fed,  and Cyp2e1 -/- mice upon EtOH fed.

city: Charleston

first: Je-Hyun

last: Yoon

state: SC

address: 173 Ashley Avenue BSB 512B, MSC 509

institute: Medical University of South Carolina

summary:
To investigate the effect of Cyp2e1 and EtOH on mRNA expression in mouse tissues, we profiled total RNA expression using mouse tissues from Cyp2e1 -/- mice and Cyp2e1 +/+ mice upon EtOH fed mice compared to control.
We compare the expression level of RNA and profile circular RNA expression using circular RNA junction.

overall_design: We prepared Cyp2e1 -/- and Cyp2e1 +/+ mice for making binge alcohol mouse model and control. After we prepared binge alcohol mouse model and control, brain, intestine, and liver tissues from the mice were isolated for total RNA-seq.

first: Young-Kook

last: Kim


title: Single nucleus transcriptome of the brains response to ischemic stroke

status: Public on Jun 29 2024

city: Vienna

first: Daniel

last: Bormann

state: Vienna

address: Waehringer Guertel 18-20

department: Department of Thoracic Surgery

country: Austria

institute: Medical University of Vienna

summary: The mammalian brain consists of heterogeneous cell populations with distinct responses to injury. Here we used single nucleus RNA sequencing (snRNAseq) to analyze cell type specific  transcriptional responses of the rodent brain to ischemic stroke, 48h after the induction of middle cerebral artery occlusion.

overall_design: Cerebral ischemia was induced by permanent intraluminal occlusion of the right middle cerebral artery (MCAO). 48h after filament based occlusion of the middle cerebral artery infarction, or the absence of infarction in Sham operated animals was confirmed via T2-weighted MRI. Brain samples were harvested, snap frozen in liquid nitrogen, nuclei were extracted from frozen brain tissue using nuclei extraction buffer (Miltenyi) and standard discontinuous iodixanol gradient separation, followed by library preparation using standard 10x Genomics workflows and snRNAseq (NovaSeq 6000, Illumina). In total brains from n=4 Sham operated rats (=Sham), n=3 rats with moderate MCAO induced infarctions (mMCAO) and n=4 rats with severe MCAO induced infarctions (sMCAO) were sequenced. Left and right hemispheres were sequenced separately. Hence, we obtained datasets from the left and right hemispheres of Sham operated rats (Sham L and Sham R, respectively), as well as left (=contralateral to ischemic lesion) and right (= ipsilateral to ischemic lesions) hemispheres of mMCAO and sMCAO infarcted rats. The following datasets were generated: Sham L = left brain hemispheres of Sham operated controls ; Sham R = right hemispheres of Sham operated controls;   mMCAO contra:  left brain hemispheres of MCAO group animals, with moderate infarction severity, hemispheres contralateral to infarction;  mMCAO ipsi: right brain hemispheres of MCAO group animals, with moderate infarction severity, hemispheres ipsilateral to infarction, sMCAO contra: left brain hemispheres of MCAO group animals, with severe infarction severity, hemispheres contralateral to infarction;  sMCAO ipsi: right brain hemispheres of MCAO group animals, with severe infarction severity, hemispheres ipsilateral to infarction.

first: Michael

last: Knoflach

first: Emilia

last: Poreba

first: Christian

last: Riedl

first: Giulia

last: Testa

first: Cyrille

last: Orset

first: Anthony

last: Levilly

first: Andreá

last: Cottereau

first: Phillip

last: Jauk

first: Simon

last: Hametner

first: Bahar

last: Golabi

first: Dragan

last: Copic

first: Katharina

last: Klas

first: Martin

last: Direder

first: Hannes

last: Kühtreiber

first: Melanie

last: Salek

first: Stephanie

last: zur Nedden

first: Gabriele

last: Baier-Bitterlich

first: Stefan

last: Kiechl

first: Carmen

last: Haider

first: Verena

last: Endmayr

first: Romana

last: Höftberger

first: Hendrik

last: Ankersmit

last: Mildner

first: Nadine

last: Stranzl


title: Investigation of melanoma cell transcriptomic states in subpopulations with high and low levels of ALDH1A3

city: Edinburgh

last: Patton

address: Western General Hospital, Crewe Road South

department: Institute of Genetics and Cancer

institute: MRC Human Genetics Unit

summary: Cancer progression and therapy resistance arise from metabolic and transcriptional adaptations, but how these are interconnected is less well understood. We propose that in melanoma, the pan-cancer stem cell marker aldehyde dehydrogenase 1A3 (ALDH1A3) couples TFAP2B-neural crest stem cell gene transcription with a distinct acetyl-histone H3 landscape via its substrate acetaldehyde. Mechanistically, nuclear acetyl-CoA synthetase 2 (ACSS2) colocalises with ALDH1A3 to bind glucose metabolic and neural crest stem cell gene loci and promote local acetyl-histone H3 modification dependent on ALDH1A3. Consistent with clinical observations that dedifferentiation into stem cell states confer poor prognosis in melanoma patients and mediate drug resistance, we found in a zebrafish melanoma model that targeting the ALDH1A3-high subpopulation with an ALDH1 suicide inhibitor delayed BRAF inhibitor drug-resistant relapse. Our work identifies ALDH1A3 as a master coordinator of acetaldehyde metabolites that directly regulates chromatin-based gene regulation, and which can be pharmacologically targeted to improve therapeutic outcomes.

overall_design: A375 melanoma cells with the highest and lowest ALDH activity were sorted based on Aldefluor signal and profiled for differential gene expression.

first: Yuting

last: Lu

first: Andrea

last: Coates

first: Graeme

last: Grimes

first: Richard

last: Clark


title: Investigation of melanoma cell chromatin acetylation states with high and low levels of ALDH1A3

overall_design: ChIP-seq profiling the acetyl-histone H3 landscape in A375 control and ALDH1A3 KO cells using antibody targeting pan-acetyl-histone H3 (K9, K14, K18, K23, K27); CUTandTAG profiling the acetyl-histone H3K23 and acetyl-histone H3K27 landscapes in MPD002 cells with or without ALDH1A3 knockdown.

first: Robert

last: Illingworth


title: Changes in tryptophan de novo NAD synthesis associated with chronic intestinal inflammation

status: Public on Jun 28 2024

city: Boston

first: Rhian

last: Stavely

address: 185 Cambridge Street, CPZN, 6100

department: Pediatric Surgery

country: USA

institute: Massachusetts General Hospital

summary: Tryptophan metabolism and de novo NAD biosynthesis is associated with behavioural changes in IBD patients. The microbiota and tryptophan de novo syntheses of the distal colon, distal colon and brain of C57BL/6 control mice and Winnie mice littermates with chronic intestinal inflammation were compared using RNA-Seq. Differentially expressed genes were analysed using Micromon software. Changes in tryptophan and nicotinamide metabolism-associated gene expressions, metabolites, and abundance of gut microbiota associated with chronic intestinal inflammation and dysregulated tryptophan metabolism in the Winnie mouse distal colon and brain were apparent. Our findings shed light on the physiological alterations in tryptophan metabolism, specifically its diversion from the serotonergic pathway towards the kynurenine pathway and the consequential effects on de novo NAD+ synthesis, in the context of chronic intestinal inflammation

overall_design: To investigate Winnie mouse brains versus C57BL/6 mouse brains, 5 males and 5 females from each group were euthanised with pentobarb, their brains were extracted and immediately frozen in -80 degrees. Then RNA was extracted using Trizol and Qiagen RNA tissue extractor.

first: Jeannie

last: Devereaux

first: Kulmira

last: Nurgali

first: Ainsley

last: Robinson


title: Pig Synthetic Blastocyst-like Structures from Embryonic Stem Cells

first: Chengchen

last: Zhao

state: Zhejiang

address: Dunyu Road No.600, Xihu District

institute: Westlake University

summary: We developed an efficient method to generate porcine blastocyst-like structures (termed blastoids) from ESCs.

overall_design: Blastoids were generated from pig ESCs in iBlastoid medium and collected on day 6. Single-cell RNA-seq was performed using 10x Genomics Chromium.

first: Jinzhu

last: Xiang

first: Hanning

last: Wang

first: Duanqing

last: Pei


title: Respiratory SARS-CoV-2 Infection Induces Skeletal Muscle Atrophy and Long-Lasting Energy Metabolism Suppression

first: Boston University

last: Microarray and Sequencing Resource

address: 72 East Concord Street, E631

department: Microarray and Sequencing Resource

institute: Boston University

summary: Coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) impacts human health beyond acute infection. Myalgia and fatigue represent two of the most prevalent symptoms in post-COVID-19 syndrome. To determine the mechanisms underlying prolonged muscle symptoms, we characterized longitudinal histopathological and transcriptional changes of skeletal muscle after acute respiratory SARS-CoV-2 infection in a COVID-19 hamster model and compared them with respiratory influenza A virus (IAV) infection. Histopathological and bulk RNA sequencing analyses at 3, 30, and 60 days post infection (dpi) showed no evidence of direct viral invasion, inflammatory cell infiltration, or microthrombi in skeletal muscle, but myofiber atrophy was observed in the SARS-CoV-2 group at 60 dpi, accompanied by downregulation of myofiber genes, atrogenes, and cytoplasmic ribosomal protein genes, and upregulation of autophagy genes. There was persistent downregulation of genes involved in mitochondrial oxidative phosphorylation, fatty acid beta-oxidation, and tricarboxylic acid cycle in the SARS-CoV-2 but not the IAV group. Moreover, TNF-alpha/NF-kB and TGF-beta signaling pathways were differentially regulated in the SARS-CoV-2 group. Our findings suggest that persistent muscle symptoms after COVID-19 may be caused by muscle atrophy and energy metabolism suppression, and that the systemic inflammatory cytokine response may contribute, in part, to the skeletal muscle abnormalities.

overall_design: Golden hamsters were intranasally administered PBS (mock-infected) or intranasally infected with 100,000 pfu of H1N1 influenza A virus (A/California/04/2009, IAV) or 1000 pfu SARS-CoV-2 (USA-WA1/2020), and RNA was extracted from the quadriceps muscle at 3 days (all infection groups) or at 30 or 60 days (IAV or SARS-CoV-2 infection groups only) and sequenced.

first: Sachiko

last: Homma

first: Lan

last: Zhou


title: CRISPR-Cas screen in ILC2 identifies Mef2d as a regulator of GATA3 and type-2 immunity

city: Cambridge

first: Ana

last: Ferreira

address: Francis Crick Avenue, Cambridge Biomedical Campus

institute: MRC - Laboratory of Molecular Biology

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

overall_design: Refer to individual Series


title: CRISPR screens and ILC2-targeted Boolean circuitry reveal Mef2d-potentiation of type-2 immunity [RNA-Seq]

summary: Group 2 innate lymphoid cells (ILC2s) orchestrate tissue homeostasis, allergic disorders and anti-helminth protective immunity through their production of cytokines. However, their rarity has hampered high-throughput genetic screening approaches to uncover novel ILC2 regulators in an unbiased manner. We combined expansion of progenitors from multi-reporter mouse strains with optimised ILC differentiation cultures to perform CRISPR-Cas9 screens for regulators of ILC2 development and function. Mef2d emerged as a critical regulator of GATA3 and IL-13, and was essential for type-2 immunity as demonstrated by conditional Mef2d-deficient mice, including a Boolean-ILC2-Cre (BIC) mouse strain developed to enable ILC2-specific gene deletion. Mechanistically, Mef2d bound and transcriptionally repressed the Zc3h12a locus encoding the endonuclease Regnase-1, a negative regulator of IL-33 receptor (ST2) and GATA3 expression, thereby promoting IL-33-mediated ILC2 proliferation and cytokine production. Notably, in ILC2s Mef2d also acted downstream of leukotriene C4-induced calcium-mediated signalling to translocate NFAT1 to the nucleus to promote type-2 gene transcription. These Mef2d-mediated pathways converge and feedback to potentiate type-2 immunity.

overall_design: Il7rCre control or Mef2dIL7RKO mice were treated with 3 doses of IL-33 intranasally and lung ILC2s were flow purified for RNA-seq analysis.


title: O-GlcNAcase Regulates Pluripotency States of Human Embryonic Stem Cells [ChIP-seq]

status: Public on Jun 27 2024

city: Hangzhou

first: Qianyu

last: Liu

address: YuhangTang Road

institute: Zhejiang University

summary: To study the occupancy by EP300 throughout the genome.

overall_design: Enrichment for occupancy by EP300 was probed with a specific antibody against EP300 in naïve hESCs

first: Wen

last: Yi


title: Maturing Neutrophils of Lower Density Associate with Thrombocytopenia in Puumala Orthohantavirus-Caused Hemorrhagic Fever with Renal Syndrome - Human samples

city: Helsinki

first: Tomas

last: Strandin

address: Haartmaninkatu 3

department: Virology

country: Finland

institute: University of Helsinki

summary: Puumala orthohantavirus-caused hemorrhagic fever with renal syndrome (PUUV-HFRS) is characterized by significant neutrophil activation. Neutrophils, the most abundant immune cells in circulation, are equipped to respond rapidly to infections and exhibit a notable heterogeneity. This study aims to identify and characterize different neutrophil subsets in the circulation of PUUV-HFRS patients, focusing on low-density granulocytes (LDGs) and normal density polymorphonuclear cells (PMNs). The study finds that PMNs show activation of antiviral pathways, while circulating LDGs increase following acute PUUV-HFRS. Additionally, PUUV-associated LDGs, primarily immature, likely reflect increased bone marrow neutrophil production. Notably, the frequency of LDGs and a "left shift" in blood are associated with the extent of thrombocytopenia, a hallmark of severe HFRS, suggesting a role for maturing neutrophils in disease pathogenesis. Unlike COVID-19 associated LDGs, PUUV LDGs did not exhibit significant immunosuppressive ability, indicating inherent biological differences in LDG responses based on the causative virus or infection kinetics. RNA sequencing was performed on neutrophils isolated from four cohorts: healthy control PMNs, PUUV-infected PMNs, PUUV-infected CD16- LDGs, and PUUV-infected CD16+ LDGs. The RNA sequencing and subsequent bioinformatics analysis were conducted by the Biomedicum Functional Genomics Unit at the Helsinki Institute of Life Science and Biocenter Finland at the University of Helsinki.

overall_design: In this study, we investigated the gene expression profiles of neutrophil subsets in patients with Puumala orthohantavirus-caused hemorrhagic fever with renal syndrome (PUUV-HFRS). Neutrophils were isolated from four distinct cohorts: healthy control polymorphonuclear neutrophils (HC PMNs), PUUV-infected polymorphonuclear neutrophils (PUUV PMNs), PUUV-infected CD16- low-density granulocytes (LDGs), and PUUV-infected CD16+ LDGs. Each cohort comprised four samples, resulting in a total of sixteen samples. The neutrophils were isolated and lysed in Trizol, followed by RNA extraction using the RNeasy micro kit. RNA sequencing was performed using the NEBNext Ultra II directional RNA library preparation method, and sequencing was conducted on the Illumina NextSeq platform. The raw and processed RNA-seq data were analyzed to quantify gene expression, enabling us to identify and characterize the different neutrophil subsets and their activation pathways during PUUV-HFRS. The findings reveal insights into the role of neutrophil heterogeneity and density in the immune response and pathogenesis of PUUV-HFRS.


title: Inverse genetics tracing the differentiation pathway of human chondrocytes

city: Okayama

first: Ziyi

state: Okayama

address: 2-5-1 Shikata-cho, Kita-ku

department: Molecular Biology and Biochemistry

institute: Okayama University

summary: Objective: Mammalian somatic cells can be reprogrammed into induced pluripotent stem cells (iPSCs) via the forced expression of Yamanaka reprogramming factors. However, only a limited population of the cells that pass through a particular pathway can metamorphose into iPSCs, while the others do not. This study aimed to clarify the pathways that chondrocytes follow during the reprogramming process. Design: The fate of human articular chondrocytes under reprogramming was investigated through a time-coursed single-cell transcriptomic analysis, which we termed an inverse genetic approach. The iPS interference technique was also employed to verify that chondrocytes inversely return to pluripotency following the proper differentiation pathway . Results: We confirmed that human chondrocytes could be converted into cells with an iPSC phenotype. Moreover, it was clarified that a limited population that underwent the silencing of SOX9, a master gene for chondrogenesis, at a specific point during the proper transcriptome transition pathway, could eventually become iPS cells. Interestingly, the other cells, which failed to be reprogrammed, followed a distinct pathway toward cells with a surface zone chondrocyte phenotype. The critical involvement of cellular communication network factors (CCNs) in this process was indicated. The idea that chondrocytes, when reprogrammed into iPSCs, follow the differentiation pathway backward was supported by the successful iPS interference using SOX9. Conclusions: This inverse genetic strategy is expected to be utilized to identify the master genes for the differentiation of various somatic cells. The utility of CCNs in articular cartilage regeneration is also supported.

overall_design: Human normal articular chondrocytes were commercially available (Lonza, Basel, Switzerland). The chondrocytes in the original stock were collected via centrifugation and resuspended in the basal medium (Lonza) containing human recombinant fibroblast growth factor 2 (rhFGF-B: Lonza), R3-insulin-like growth factor-I (R3-IGF-1: Lonza), fetal bovine serum (FBS), gentamicin/amphotericin-B (GA), and insulin and transferrin (chondrocyte growth medium: CGMTM). Cells were plated, expanded in the monolayer culture at 37 °C in a 5% CO2 and detached for subcultures using 0.05% trypsin/0.02% EDTA (Biowest, Nuaillé, France). The chondrocytes were infected with SRV iPSC-3 recombinant virus, incubated for 2 hours at room temperature and further incubated in a CO2 incubator overnight at 37 °C. The following day, cells were collected and seeded on an iMatrix-511-coated (Takara Bio, Kusatsu, Japan) plate with a Stemfit Basic 02 medium (Ajinomoto, Tokyo, Japan). The medium was removed and replenished every 2 days until day 8. Thereafter, the medium was changed to the basal medium containing rhFGF-2, and cells were cultured until day 10.

first: Hang

last: Do

first: Mitsuaki

last: Ono

first: Wakana

last: Kitagawa

first: Anh

last: Dang

first: Tomoko

last: Yonezawa

first: Takuo

last: Kuboki

first: Toshitaka

last: Oohashi

first: Satoshi

last: Kubota


title: O-GlcNAcase Regulates Pluripotency States of Human Embryonic Stem Cells [CUT&Tag]

country: China

summary: In order to study the occupancy by OGA throughout the genome.

overall_design: Enrichment for occupancy by OGA was probed with a specific antibody against OGA in WT or OGA-depleted naïve hESCs


title: Histopathological-based analysis of human kidney spatial transcriptomics data: toward precision pathology

city: St. Louis

first: Dian

last: Li

state: MO

address: 660 S. Euclid Ave., CB 8118

department: Nephrology / Medicine

institute: Washington University in St. Louis School of Medicine

summary: In this study, we conducted a histopathological-based analysis of 10x Genomics Visium spatial transcriptomics data on a human kidney sample corresponding as closely as possible to the reality of the interpretation of a kidney biopsy in a healthcare or research context.

overall_design:
A kidney specimen from a 33-year-old deidentified patient with chronic kidney disease (CKD) as a consequence of diabetic and hypertensive nephropathy was used for analysis. Utilizing the H&E- and PAS-stained bright-field mosaics as references, a kidney pathologist (PI) conducted morphological-based clustering for each spatially-resolved spot on the Visium slide. This involved manually outlining each kidney morphological structure and/or lesion for the purpose of cluster identification.

A GitHub repository containing the R, Julia, and Python scripts we wrote for the main and supplemental figures can be found at:

first: Pierre

last: Isnard

first: Haojia

last: Wu

first: Benjamin

last: Humphreys


title: Small molecule probe for IBD risk variant GPR65 I231L alters cytokine signaling networks through positive allosteric modulation

first: Ramnik

last: Xavier

state: MA

address: 415 Main Street

institute: Broad Institute of MIT and Harvard

summary: BRD5075 is a positive allosteric modulator of GPR65. Activity has been demonstrated by cAMP production in HeLa cells overexpessing GPR65 and target engagment has been confirmed. BRD5075 also decreases TNF-a secretion by GPR65 WT BMDCs. Goal is to identify differentially expressed genes in response to this GPR65 PAM

overall_design: Total 90 samples. GPR65 WT, variant I231L, and KO BMDCs stimulated with different pH and GPR65 PAM in triplicate on the same plate for each timepoint (2 hours and 6 hours). Positive control is PGE2, negative is DMSO.

first: Ilona

last: Neale

first: Daniel

last: Graham

first: Joshua

last: Park

first: Bihua