title: SMPDL3b in Podocytes: Decoupling Proteinuria from CKD Progression in Experimental Alport Syndrome

city: Miami

first: Alla

last: Mitrofanova

state: Florida

address: 1580 NW 10th Ave, Batchelor Bldg, 6th floor

institute: University of Miami

summary: Alport Syndrome (AS) is a rare genetic disease with impaired production of collagen type IV alpha 3, 4 and 5 chains in the glomerular basement membranes (GBM), which results amongst others in progressive loss of kidney function. In AS, abnormalities in the GBM and associated podocyte detachment may potentially result from the dysregulation of sphingolipid metabolism. Here we investigated whether renal sphingomyelin phosphodiesterase acid-like 3b (SMPDL3b) overexpression modulates the generation of sphingosine-1-phosphate (S1P) and contributes to renal failure in Col4a3 knockout mice, a mouse model of AS. We found a 3-fold increase in SMPDL3b expression in glomeruli and murine podocytes isolated from Col4a3 knockout mice. Increased SMPDL3b expression occurred in association with increased glomerular S1P levels, while podocyte specific Smpdl3b deletion in Col4a3 knockout mice was sufficient to restore S1P levels and to reduce proteinuria and podocyte foot process but not sufficient to protect from renal failure. Thus, podocyte S1P may be a key modulator of proteinuria and podocyte integrity in AS. Our study in experimental AS suggests that SMPDL3b-derived S1P may dissociate proteinuria from renal failure, and suggests that improvement of glomerular structure and function may not always translate in protection from CKD progression in AS.

overall_design: RNA serquence data in whole cell extracts of differentiated murine podocytes. RNA from immortalized wildtype (IMWT) and Alport Syndrome (IMAS) podocytes was isolated and analyzed for differences in the RNA expression profile. RNA extraction, Preparation and sequencing RNA libraries, Illumina Sequencing RNA Data Analysis.

first: Rachel

last: Njeim

first: Alessia

last: Fornoni


title: Network-based bioinformatics highlights broad importance of human milk hyaluronan

status: Public on Dec 19 2024

city: Oklahoma City

first: Kathryn

last: Burge

state: OK

address: 800 Research Parkway

department: Pediatrics

country: USA

institute: University of Oklahoma Health Sciences Center

type: Expression profiling by high throughput sequencing

summary: Hyaluronic acid 35 kDa (HA35) is protective against murine necrotizing enterocolitis and is present in high levels in colostrum and early human milk. We utilized pathway and network analysis of bulk RNA-seq transcriptomics of the distal ileum of mouse pups to determine molecular mechanisms through which HA35 may influence small intestinal maturation and protection against inflammatory disease.

overall_design: Bulk transcriptomics was performed on the distal ileum from CD-1 mouse pups treated once daily with 30mg/kg body weight 35 kDa hyaluronic acid (n = 4) or vehicle control (equal volume sterile water, n = 4) from postnatal day of life 7-14.

middle: Y

first: Hala

last: Chaaban

first: Adam

middle: P

last: Wilson


title: MircorRNA in Hippocampus in human samples

city: San Sebastián

first: David

last: Otaegui

state: Guipuzcoa

address: Paseo Dr begiristain s/n

department: Neurosciences

country: Spain

institute: Biodonostia Research institute

first: Muñoz-Culla

last: Maider

first: Otaegui

last: David

first: Alberro

last: Ainhoa

first: Matheu

last: Ander


title: Noncanonical role of ALAS1 as a heme-independent inhibitor of small RNA–mediated silencing [hepatocyte RNA-Seq]

city: New York

first: Seungjae

last: Lee

state: NY

address: 430 East 67th St.

department: Department of Developmental Biology

institute: Sloan-Kettering Institute

summary: RNA interference (RNAi) is a fundamental strategy for gene silencing via small interfering RNAs (siRNAs) that program Argonaute (Ago) effectors, and is related to endogenous silencing via microRNAs (miRNAs). Although most efforts focus on factors that are required for miRNA/RNAi function, biological systems are typically subject to repression. Using molecular genetic approaches, we uncover ALAS1, the first enzyme in the heme biosynthesis pathway, as a repressor of miRNA accumulation in cells and in mice. This was non-intuitive, since heme is known to play a positive role as a cofactor for the nuclear miRNA processing machinery. Instead, we show that ALAS1 (but not other core heme biosynthesis factors) limits RISC assembly and activity. Notably, ALAS1 is the target of one the few current FDA-approved siRNA drugs (Givosiran). Our data have implications for side effects of Givosiran, but also for RNAi therapies in general. We provide evidence that depletion of ALAS1 enhances siRNA-mediated knockdown, suggesting that Givosiran is a viable RNAi adjuvant.

overall_design: To investigate the cooperative function of Alas in mouse hepatocytes, we engineered a conditional mouse allele that permits Cre-inducible deletion of Alas1 exons 4-5 and further depleted Alas2 by injecting GalNAc-conjugated siRNAs.

first: Sangmi

first: Robert

last: Desnick

first: Makiko

last: Yasuda

first: Eric

middle: C

last: Lai


title: B7-H4 regulates β cell mass and insulin secretion by modulating cholesterol metabolism

city: Shanghai

first: Fangzhen

last: Xia

address: Zhizaoju Road NO.639

country: China

institute: Department of endocrinology, Shanghai Ninth People's Hospital

summary: Chronic islet inflammation is a hallmark of type 2 diabetes (T2D) and involves in the dysfunction of β cells. However, how β cells participate in this process remains unclear. Here, we report that the immune checkpoint molecule B7-H4(B7S1, B7x, VTCN1) expressed in β cells is critical to maintain β cell mass and insulin secretion. Lesion of B7-H4 in β cells results in glucose intolerance due to less β cell mass  and deficient insulin secretion with upregulated cytokines and activated signal transducer and activator of transcription 5 (Stat5) signaling, while overexpression of B7-H4 in β cells ameliorates glucose intolerance in high-fat diet (HFD)-treated mice. Mechanistically, B7-H4 deficiency actives the Stat5 signaling, which inhibits the expression of Apolipoprotein F (ApoF), leading to reduced cholesterol efflux and accumulated cholesterol in β cells, thereby impairing the insulin processing and secretion. Inhibiting Stat5 activity or overexpression of ApoF in β cells can rescue the glucose intolerance and insulin secretion deficiency in β-cell-specific B7-H4 knockout (B7-H4 cKO) mice. Our study demonstrates that β cell expressed immune checkpoint molecule B7-H4 is essential for islet immune homeostasis and β cell function maintenance, and for the first time unravels the mechanism by which B7-H4 regulates insulin secretion through regulating cholesterol metabolism via Stat5 signaling, which may shed new light on the development of novel strategies for T2D treatment.

overall_design: B7-H4 fl/fl mice, in which exon 2 of the Vtcn1 gene is flanked by two loxP sites, were generated by Cyagen Biosciences (Suzhou, China) with CRISPER/Cas9 system. B6.FVB-Tg(Pdx1-cre)6Tuv/J , commonly called Pdx-Cre mice in which Cre recombinase is under the transcriptional control of the mouse Pdx1 promoter, and B6.Cg-Tg(Ins2-cre)25Mgn/J, commonly called RIP-Cre mice in which Cre recombinase is under the control of the rat insulin II promoter were purchased from the Jackson Laboratory. Mice homozygous for the floxed B7-H4 allele (F/F) were crossed with Pdx-Cre or Rip-Cre transgenic mice to create β-cell–specific B7-H4 knockout mice (Pdx-Cre; B7-H4 fl/fl ; PB7H4KO or RIP-Cre; B7-H4 fl/fl ; βB7H4KO) and their control littermates. To elucidate the therapeutic effect of B7-H4 on obesity associated metabolic dysfunction, 6 weeks old WT mice fed with 8 weeks of NCD or HFD were then injected with adeno-associated virus serotype 8 vector (AAV8)–rat Insulin2 promoter (RIP)–specific B7-H4-ZSGreen or with control virus (Hanbio Biotechnology Co.Ltd., Shanghai, China). To clarify the role of ApoF in β cell function affected by β-cell–specific B7-H4 knockout,  6 weeks old flox and βB7H4KO mice fed with 8 weeks of HFD were injected  with AAV8-Apof-ZSgreen or control virus (Genomeditech, Shanghai, China).The viruses (1012 genome copy particles/mL) were infused at a rate of 6 μL/min by pancreatic intraductal viral infusion as described previously.


title: Small RNAs in plasma extracellular vesicles define biomarkers of premanifest changes in Huntington's disease

city: Barcelona

first: Ana

middle: Gámez

last: Valero

state: Barcelona

address: Universitat de Barcelona, Casanova 143, 4th Floor-Nord

institute: University of Barcelona (Neuroscience Institute)

type: Non-coding RNA profiling by high throughput sequencing

summary: Despite the advances in the understanding of Huntington’s disease (HD), there is the need for molecular biomarkers to categorize mutation-carriers during the preclinical stage of the disease preceding functional decline. Small RNAs (sRNAs) are a promising source of biomarkers since their expression levels are highly sensitive to pathobiological processes. Here, using an optimized method for plasma extracellular vesicles (EVs) purification and an exhaustive analysis pipeline of sRNA sequencing data, we show that EV-sRNAs are downregulated early in mutation-carriers and that this deregulation is associated with premanifest cognitive performance. Seven candidate sRNAs (tRF-Glu-CTC, tRF-Gly-GCC, miR-451a, miR-21-5p, miR-26a-5p, miR-27a-3p, and let7a-5p) were validated in additional subjects, showing a significant diagnostic accuracy at premanifest stages. Of these, miR-21-5p was significantly decreased over time in a longitudinal study; and miR-21-5p and miR-26a-5p levels correlated with cognitive changes in the premanifest cohort. In summary, the present results suggest that deregulated plasma EV-sRNAs define an early biosignature in mutation carriers with specific species highlighting the progression and cognitive changes occurring at the premanifest stage.

overall_design:
To evaluate the biomarker potential of plasma extracellular vesicles (EVs) and their sRNAs in Huntington's disease, we first analyzed the the sRNA distribution in the different plasma subfractions obtained through Size-Exclusion Chromatography (SEC) using samples from healthy individuals.
Plasma samples from healthy non-mutation carriers and Huntington's disease patients, both at the premanifest and manifest stages, were subjected to SEC and ultrafiltration to isolate EVs. RNA from the vesicles was obtained and 10 samples per group were used for smallRNAseq.
Small RNAseq data were processed and analyzed using two bioinformatic tools: ExceRpt and SeqCluster. Comparative univariate and multivariate analyses were performed to identify differentially expressed small RNAs between Huntington's disease patients and healthy subjects. Results from smallRNAseq were further validated using qRT-PCR in 20 samples per group.

first: Marina

last: Herrero-Lorenzo

first: Jesús

last: Pérez-Pérez

first: Georgia

last: Escaramís

first: Saül

last: Martínez-Horta

first: Rocío

last: Pérez-González

first: Elisa

last: Rivas-Asensio

first: Jaime

last: Kulisevsky

last: Gámez-Valero

first: Eulàlia

last: Martí


title: Single-cell transcriptomics reveals a pivotal role of DOCK2 in Sjögren’s disease

city: Gainesville

first: Cuong

last: Nguyen

address: 2015 SW 16th Ave

institute: University of Florida

summary: Sjögren's disease (SjD) is an autoimmune condition characterized by the dysfunction of the salivary and lacrimal glands. The study aimed to decipher the pathogenic cell populations and their immunological pathways in the salivary glands.  We further determined the therapeutic effect of inhibiting DOCK2 shared by novel clusters of CD8Cd8+ T cells in a SjD mouse model.

overall_design: Salivary glands of C57BL/6.NOD-Aec1/2 (51 weeks old, n=2 female, 2 male) and B6 (64 weeks old, n=2 female, 2 male) mice were explanted and digested in a buffer containing 1 mg/ml DNase (Sigma-Aldrich, St. Louis, MO) and 1 mg/ml Collagenase Type 4 (Worthington, Lakewood, NJ, USA) in RPMI (Lonza, Allendale, NJ) complete media (10% FBS, 2 mM L-glutamine, 0.05 mM β-mercaptoethanol).  Tissues were placed in a MACS C tube (Miltenyi Biotec, San Diego, CA) for desiccation on GentleMACS V1.02 for a pulse of 38 seconds. After a 10-minute incubation at 37°C, the digest buffer was removed and placed into 4°C RPMI complete media.  The process was repeated twice. Single-cell suspensions were centrifuged (2500 rpm, 10 min, 4°C) and resuspended in PBS for filtration through a 70-μm sterile cell strainer (Fisher, Pittsburgh, PA). After a wash with PBS, cells were resuspended again in PBS for lymphocyte isolation with Lympholyte-M cell separation media (Cedar Lane, Burlington, Canada) per the manufacturer's instructions. Single-cell suspensions were stained for DAPI, and live cells were sorted with a sorter (SH800S, Sony, San Jose, CA) into RPMI containing 10% FBS on ice for single-cell sequencing library preparation.

first: Yiran

last: Shen

first: Alexandria

last: Voigt

first: Indraneel

last: Bhattacharyya

middle: Q


title: Improved health by combining dietary restriction and promoting muscle growth in DNA repair-deficient progeroid mice

city: Utrecht

first: Wilbert

last: Vermeij

address: Heidelberglaan 25

country: Netherlands

institute: Princess Máxima Center

summary: Ageing is a complex multifactorial process, impacting all organs and tissues, with DNA damage accumulation serving as a common underlying cause. To decelerate ageing, various strategies have been applied to model organisms and evaluated for health and lifespan benefits. Dietary restriction (DR, also known as caloric restriction) is a well-established long-term intervention recognised for its universal anti-ageing effects. DR temporarily suppresses growth, and when applied to progeroid DNA repair-deficient mice doubles lifespan with systemic health benefits. Counterintuitively, attenuation of myostatin/activin signalling by soluble activin receptor (sActRIIB), boosts the growth of muscle, and in these animals prevents muscle wasting, improves kidney functioning, and compresses morbidity. Here we investigated a combined approach, applying an anabolic regime (sActRIIB) at the same time as DR to Ercc1d/- progeroid mice. Following both single treatments and combined, we monitored global effects on body weight, lifespan and behaviour, and local effects on muscle and tissue weight, muscle morphology and function, and ultrastructural and transcriptomic changes in muscle and kidney. Lifespan was mostly influenced by DR (extended from approx. 20 to 40-weeks; p<0.001), with sActRIIB clearly increasing muscle mass (35-65%) and tetanic force (p<0.001). The combined regime yielded a stable uniform body weight, but increased compared to DR alone, synergistically improved motor coordination and further delayed the onset and development of balance problems. sActRIIB significantly increased muscle fibre size (p<0.05) in mice subjected to DR and lowered all signs of muscle damage. In the neuromuscular junction of Ercc1d/- mice the junctional folding of the basement membrane at the postsynaptic side was either rudimentary or completely missing. Single interventions by sActRIIB treatment or DR only partially rescued this phenotype, while in the double intervention group the regularly shaped junctional foldings were maintained. In kidney of Ercc1d/- mice we observed a mild but significant foot process effacement, which was restored by either intervention. Transcriptome analysis also pointed towards reduced levels of DNA damage in muscle and kidney by DR, but not sActRIIB, while these levels retained lower in the double intervention. In muscle we found synergistic effects of combining sActRIIB with DR, but not in kidney, with an overall better health in the double intervention group. Crucially, the benefits of each single intervention are not lost when administered in combination, but rather strengthened, even when sActRIIB was applied late in life, opening opportunities for translation to human.

overall_design: RNA sequencing was performed on mouse quadriceps muscle and kidney specimens of 16-week-old male Ercc1d/- progeroid mice subjected to sActRIIB, dietary restriction, or double intervention, with mock ad libitum Ercc1d/- and Ercc1+/+ wildtype controls.

first: Rutger

middle: A

last: Ozinga

first: Ivar

last: van Galen

first: Oliver

last: Kretz

first: Ketan

last: Patel


title: TOR balances plant growth and cold tolerance by orchestrating amino acid-derived metabolism in tomato

first: Liwen

last: Fu

address: 800 Dongchuan RD. Minhang District

institute: Shanghai Jiao Tong University

summary: The target of rapamycin (TOR) kinase is a central signaling hub that plays a crucial role in precisely orchestrating plant growth and development. It is increasingly evident that TOR also has a significant impact on plant stress responses. This suggests that TOR is intricately involved in maintaining the balance between plant growth and stress responses. Nevertheless, despite the observed effects, the specific mechanisms through which TOR operates in these processes remain obscure. In this study, we investigated how the tomato (Solanum lycopersicum) TOR (SlTOR) affects plant growth and cold responses. We demonstrated that the inhibition of SlTOR transcriptionally primes cold stress responses, consequently enhancing tomato cold tolerance. Furthermore, a widely targeted metabolomics analysis revealed the disruption of amino acid metabolism homeostasis under cold stress upon SlTOR inhibition. This disruption led to the accumulation of two important defense metabolites: salicylic acid (SA) and putrescine (Put). Subsequent gain- and loss-of-function analyses validated the crucial roles of salicylic acid and putrescine in enhancing tomato cold tolerance. Our study provides a mechanistic framework that elucidates how SlTOR modulates amino acid-related metabolism to enhance tomato cold tolerance, which sheds light on the complex interplay between growth and stress responses orchestrated by TOR.

overall_design: To investigate the biological function of tomato TOR in the regulation of transcriptional reprogramming. Four biological replicates of 10-day-old tomato seedlings were treated with or without 5 μM Torin2. We then proformed gene expression profiling analysis using data obtained from RNA-seq. Comparative gene expression profiling analysis  was further carried out between mock-treated (m) and Torin2-treated (T) tomato seedlings.

first: Zihao

last: Li

first: Lin

last: Yang

first: Yanni

last: Wu

first: Ran

last: Zhang

first: Sen

last: Yu


title: Mechanosensitive Piezo1 is crucial for periosteal stem cell-mediated fracture healing

city: Shijiazhuang

first: Siming

last: Jia

address: 河北省石家庄市河北医科大学第三医院

institute: 河北医科大学

overall_design: At 10 days (n = 5) and 20 (n = 5) days post fracture (dpf), the callus was isolated from each mouse

first: Weijian

last: Liu

first: Wei

last: Chen

first: Yingze


title: Interaction of methyl-CpG-binding protein 2 (MeCP2) with distinct enhancers in the mouse cortex.

city: Dallas

first: Hume

last: Stroud

state: TX

address: 5323 Harry Hines Blvd

institute: UT Southwestern

type: Genome binding/occupancy profiling by high throughput sequencing

summary:
Mutations in methyl-CpG-binding protein 2 (MeCP2) cause Rett
syndrome. MeCP2 is thought to regulate gene transcription by binding to
methylated DNA broadly across the genome. Here, using cleavage under
target and release under nuclease assays in the adult mouse cortex, we
show that MeCP2 strongly binds to specific gene enhancers that we call
MeCP2-binding hotspots (MBHs). Unexpectedly, we find that MeCP2
binding to MBHs occurs in a DNA methylation-independent manner at
MBHs. Multiple MBH sites surrounding genes mediate the transcriptional
repression of genes enriched for neuronal functions. We show that MBHs
regulate genes irrespective of genic methylation levels, suggesting that
MeCP2 controls transcription via an intragenic methylation-independent
mechanism. Hence, disruption of intragenic methylation-independent gene
regulation by MeCP2 may in part underlie Rett syndrome.

overall_design: CUT&RUN (MECP2, IgG and H3), CUT&TAG (MECP2) and ChIPseq (H3 and H3K9me3) samples in various condition

first: Gyan

last: Mishra

first: Eric

middle: X

last: Sun

first: Tiffany

last: Chin

first: Mandy

last: Eckhardt

first: Michael

middle: E

last: Greenberg


title: Urolithin A and Nicotinamide Riboside differentially regulate innate immune defenses and metabolism.

city: Baltimore

first: Supriyo

last: De

state: Maryland

address: 251 Bayview Blvd

department: Laboratory of Genetics and Genomics

institute: NIA-IRP, NIH

summary: During aging, general cellular processes such as autophagic clearance, DNA repair, mitochondrial health, metabolism, nicotinamide adenine dinucleotide (NAD+) levels, and immunological responses become compromised. Urolithin A (UA) and NAD+ supplementation stimulate mitophagy, reduce pathological plaque load, and inflammation while improving learning in models of Alzheimer’s disease (Hou et al 2021; Fang et al 2019). Here we have performed a During aging, general cellular processes such as autophagic clearance, DNA repair, mitochondrial health, metabolism, nicotinamide adenine dinucleotide (NAD+) levels, and immunological responses become compromised. Urolithin A (UA) and NAD+ supplementation stimulate mitophagy, reduce pathological plaque load, and inflammation while improving learning in models of Alzheimer’s disease (Hou et al 2021; Fang et al 2019). Here we have performed a pairwise analysis of UA and nicotinamide riboside (NR) to investigate how these compounds modulate inflammation. The role of microglia in driving neuroinflammation is becoming more recognized in several neurodegenerative diseases, yet the effects of these drugs on microglia cells have not been thoroughly investigated. As both UA and NR are recognized as safe dietary supplements, their function in normal cells is equally important to understand as in a disease state. Here we investigated the effects of UA and NR in the human microglia cell line, HMC3.

overall_design: HMC3 cells were treated with 10 µM UA or 3 mM NR for 1 week. For gene expression analysis, we used the human neuroinflammation panel (XT-CSO-MNROI1-12).

first: Vilhelm

last: Bohr


title: Multi-organ immunity on a 3D-printed multi-tissue chip with tubing-free impeller pump

city: Charlottesville

first: Anwaruddin

last: Mohammad

state: VA

address: Pinn Hall

department: Bioinformatics Core

institute: University of Virginia

summary: Multi-organ-on-chip systems (MOOCs) have the potential to mimic communication between organ systems and reveal mechanisms of health and disease. However, many existing MOOCs are challenging for non-experts to implement, due to complex tubing, electronics, or pump mechanisms. In addition, few MOOCs have incorporated immune organs such as the lymph node (LN), limiting their applicability to critical events such as vaccination. Here we developed a 3D-printed, user-friendly device and companion tubing-free impeller pump to co-culture two or more tissue samples, including a LN, under a recirculating common media. Native tissue structure and immune function were incorporated by maintaining slices of murine LN tissue ex vivo in 3Dprinted mesh supports for at least 24 hr. In a two-compartment model of a LN and an upstream injection site, vaccination of the multi-tissue chip was similar to in vivo vaccination in terms of locations of antigen accumulation and acute changes in activation markers and gene expression in the LN. We anticipate that in the future, this flexible platform will enable models of multi-organ immune responses throughout the body.

overall_design: We compared the response to vaccination in murine lymph nodes, where the vaccination was performed in a multi-tisse chip, in vivo, or in standard well plate (wells). Our model vaccine was comprised of R848 as the adjuvant mixed with rhodamine-labeled ovalbumin (Rho-OVA) as the antigen. PBS was used as a negative control for vaccination. Following 6 hrs, RNA was isolated from whole lymph nodes after in vivo vaccination or sliced lymph nodes after on-chip and well plate conditions. Each sample from in vivo vaccination was comprised of two whole lymph nodes from two mice pooled together. Each sample from both on-chip and well plate vaccination was comprised of 6 total lymph node slices from two mice pooled together. This experiment was replicated three times. RNA was analyzed by bulk RNA sequencing.

first: Sophie

middle: R

last: Cook

first: Alexander

middle: G

last: Ball

first: Rebecca

last: Pompano


title: The effect of senescent cells depletion in obese hearts

city: Los Angeles

first: Yao Wei

last: Lu

state: CA

address: 2011 Zonal Ave

department: Department of Medicine

institute: University of Southern California

summary: Obesity is a major contributor to metabolic and cardiovascular diseases. Senescence is a highly dynamic process activated by diverse stimuli and increased cellular senescence has been associated with age-related disorders. Here, we investigated the impact of cellular senescence in obesogenic diet-related metabolic and cardiac dysfunctions. An obesogenic diet induced an increase on body weight gain and adiposity, glucose intolerance, insulin resistance, dyslipidemia, and hepatic disorders in mice; however, these alterations were prevented by a senolytic cocktail (dasatinib and quercetin), which leads to removal of senescent cells. In addition, the elimination of senescent cells counteracted the activation of the senescent program and DNA damage in the white adipose tissue (WAT) induced by an obesogenic diet. Obese mice had an increase of the senescence-associated secretory phenotype (SASP) and DNA damage in the heart, cardiac hypertrophy, and diastolic dysfunction; however, the use of a senolytic combination abolished these myocardial alterations caused by an obesogenic diet. Transcriptomic analysis of the hearts revealed that obese mice exhibited a downregulation of genes associated with fatty acid metabolism, oxidative phosphorylation, PI3K AKT MTOR signaling, P53 pathway, and DNA repair; however, the treatment with senolytic cocktail induced an increase of these pathways in the heart. Collectively, these data suggest that obesogenic diet elicits WAT and cardiac senescence program in mice, and that targeting senescent cells may be a novel therapeutic strategy for attenuating obesity-related metabolic and cardiac disorders.

overall_design: C57BL/6NTac female mice at 8 weeks of age were fed a control diet (69.1% kcal carbohydrate, 10.4% kcal fat, 20.5% kcal protein, Envigo®) or an obesogenic diet, which consisted of a combination of a high-fat diet (21.3% kcal carbohydrate, 60.3% kcal fat, 18.4% kcal protein, 5.1 kcal/g, Envigo®) and sweetened condensed milk (55% sugar, 13% fat, 7% protein, 3.25 kcal/g, Nestlé®) for 24 weeks (de Oliveira Silva et al., 2020). To assess the impact of senescent cells removal in obesity-induced metabolic and cardiac dysfunctions, we used a senolytic cocktail. After 16 weeks of obesogenic diet feeding, some obese female mice were treated with a senolytic cocktail containing dasatinib (5mg/kg) and quercetin (50mg/kg) diluted in 10% PEG400, which was administered orally (via gavage) for three consecutive days weekly (Xu et al., 2018) until the 24th week of treatment. Other obese female mice were treated with vehicle (10% PEG400) via oral gavage. Total RNA was isolated from the hearts using the TRIzol protocol (Invitrogen). RNA integrity and quality was assessed using the Qubit RNA. One μg of total RNA per sample was used for the library preparations. The libraries were prepared and sequenced by GENEWIZ (Azenta Life Sciences) using Illumina (2x150bp, at least 60M reads per sample).

first: Tábatha

last: de Oliveira Silva

first: Guilherme

last: Lunardon

first: Yao

middle: W

first: Gabriela

last: Diniz


title: ARMC5 selectively degrades SCAP-free SREBF1 and is essential for fatty acid desaturation in adipocytes

city: Suita

first: Yosuke

last: Okuno

address: 2-2, Yamadaoka

country: Japan

institute: Osaka University Graduate School of Medicine

summary:
Background: Armadillo repeat containing 5 (ARMC5)-CUL3 complex was recently identified as an E3 ubiquitin ligase of full-length SREBF. Although ARMC5 was involved in SREBF2 pathway in the adrenocortical cells and might participate in the development of primary bilateral macronodular adrenal hyperplasia (PBMAH), the role of ARMC5 in adipocytes has not been investigated.
Methods and Results: Adipocyte-specific Armc5 knockout mice were generated. In the white adipose tissue (WAT) of these mice, all the stearoyl-CoA desaturase (Scd) were drastically downregulated under refed condition and high fat-high sucrose diet. Consistently, unsaturated fatty acids were decreased, while saturated fatty acids were increased in the WAT of these mice. Assay for transposase-accessible chromatin with high-throughput sequencing (ATAC-Seq) peaks at the SREBF1-binding sites were markedly diminished around the Scd1 locus in the WAT of adipocyte-specific Armc5 knockout mice. Armc5-deficient 3T3-L1 adipocytes also exhibited downregulation of Scd. Mechanistically, disruption of Armc5 restored decreased full-length SREBF1 in the CHO cells deficient for Scap, an escort protein required for the transport of SREBF from the ER to the Golgi. In HEK293T cells, overexpression of Scap inhibited ARMC5-mediated degradation of full-length SREBF1, and overexpression of Armc5 increased nuclear SREBF1/full-length SREBF1 ratio in the presence of exogenous Scap in HEK293T cells.
Conclusions: ARMC5 exerts selective removal of SCAP-free SREBF1 and stimulates SCAP-mediated SREBF1 processing, hence is essential for fatty acid desaturation in vivo.

overall_design: RNA-Seq of epididymal WAT from control flox and adipocyte-specific Armc5 KO.


title: Noncanonical role of ALAS1 as a heme-independent inhibitor of small RNA–mediated silencing [hepatocyte_sRNA-seq]

first: Seungjae

last: Lee

overall_design: To investigate the role of ALAS in small RNA regulation, we established ALAS1/2 mutant hepatocyte and validated small RNA expressions

first: Sangmi

first: Robert

last: Desnick

first: Makiko

last: Yasuda

last: Lai


title: Noncanonical role of ALAS1 as a heme-independent inhibitor of small RNA–mediated silencing [HEK293T_sRNA-seq]

overall_design: To investigate the role of ALAS1 in small RNA regulation, we established ALAS1 knock-out HEK293T cells and validated small RNA expressions

first: Eric


title: Noncanonical role of ALAS1 as a heme-independent inhibitor of small RNA–mediated silencing [Ago2-CLEAR-CLIP]

overall_design: To investigate functional effects on miRNA-mediated targeting upon loss of ALAS1, we performed Ago2 CLEAR-CLIP


title: Excess shed mesothelin disrupts pancreatic cancer cell clustering to impair peritoneal colonization

city: Lebanon

first: Christine

last: Alewine

state: NH

address: One Medical Center Drive, Williamson 651,

institute: Dartmouth Cancer Center

summary: Peritoneum is the second most common site of metastasis in patients with pancreatic ductal adenocarcinoma (PDAC). Peritoneal colonization is impaired in PDAC cells with knock-out (KO) of the cancer surface antigen mesothelin (MSLN) or by introducing Y318A mutation in MSLN to prevent binding to mucin-16 (MUC-16). MSLN has a membrane-bound form but is also shed to release soluble MSLN (sMSLN). Their individual roles in peritoneal metastasis are unknown. Here, a C-terminal truncated MSLN mutant (∆591) incapable of cell membrane insertion but proficient in secretion was engineered. Expression of ∆591 MSLN failed to rescue peritoneal metastasis in MSLN KO cells and inhibited peritoneal colonization when overexpressed in WT PDAC cells. Exposing PDAC cells to conditioned medium (CM) containing excess sMSLN impaired cancer cell clustering in vitro and in peritoneal fluid in vivo, while CM containing only Y318A sMSLN did not. These data demonstrate that interaction of membrane-bound MSLN with MUC-16 promotes cell clustering that is critical for efficient peritoneal metastasis. However, peritoneal colonization by MSLN KO cells was rescued by expression of ∆591 mutant MSLN bearing Y318A mutation, suggesting that sMSLN also has a MUC-16-independent role in peritoneal spread. Alterations in inflammatory signaling pathways occurred following KO cell exposure to CM containing sMSLN, and CM from cancer cells with intact peritoneal metastasis provoked increased KO cell secretion of IL-1α. While excess sMSLN inhibits cell clustering and peritoneal colonization, sMSLN may also promote PDAC peritoneal metastasis independent of MUC-16.

overall_design: Human pancreatic cancer cell lines KLM1 and T3M4 with MSLN knockout (KO) were cultured with the supernanat from MSLN KO KLM1 and T3M4 cell lines (Mock),  engineering of KLM1 KO cells over-expressing full-length WT (WT) and Y318A (Y318A) mutant MSLN supernatant, as well as from the wild type KLM1 and T3M4 (WT) for 4 hours.  total RNA was extracted for bulk RNAseq analysis.

first: Theressa

last: Ewa

first: Neel

last: Panchwagh

first: Chin-Hsien

last: Tai

first: Leela

last: Avula

first: Sarah

last: Joseph

first: Michael

last: Rudloff

first: Xianyu

last: Zhang


title: Inhibition of Hippo signaling pathway preserves renal mitochondrial quality control homeostasis and protects against diabetic kidney diseases

city: Guangzhou

first: Bin

last: LI

state: Guangdong

address: 58th Zhongshan Road II

department: Department of Nephrology, The First Affiliated Hospital

institute: Sun Yat-sen University

summary: Diabetic kidney disease (DKD) has become the leading cause of end-stage renal disease worldwide. Therefore, efforts to understand DKD pathophysiology and prevent its development are highly warranted. Here, we analyzed renal cortex from healthy mice, diabetic mice, and diabetic mice treated with the Hippo signaling pathway inhibitor XMU-MP-1. Our study first confirmed that XMU-MP-1 improved DKD in type 2 diabetic mice induced by high-fat diet feeding and intraperitoneal injection of streptozotocin. Through RNA-sequencing of isolated renal cortex, we found that activation of the Hippo signaling pathway, chemokine signaling pathway and cell activation involved in immune response were positively associated with the progression of DKD, while mitochondrion organization was negatively associated with the development of DKD. By contrast, administration of XMU-MP-1 partly suppressed the irritation of chemokine signaling pathway, and organ or tissue specific immune response, but preserved mitochondrion organization within renal cortex of DKD. In vitro, cell culture of immortalized renal tubule cells revealed that overexpression of yes-associated protein 1, the downstream core mediator of the Hippo signaling pathway, reduced high glucose and palmitic acid-induced tubule cell impairment by improving the maintenance of mitobiogenesis and mitophagy. Overall, our data emphasize that the renal protective effects of suppressing Hippo signaling pathway are likely through preserving the mitochondrial quality control homeostasis in tubule cells.

overall_design: The mice were categorized into four groups: (1) Mice on a normal diet (Normal diet + Citrate buffer + Vehicle) group (n=6); (2) Mice on a normal diet with XMU-MP-1 administration (Normal diet + Citrate buffer + XMU-MP-1) group (n=6); (3) Mice on a high-fat diet with streptozotocin (STZ) administration (High-fat diet + STZ + Vehicle) group (n=6); and (4) Mice on a high-fat diet with STZ and XMU-MP-1 administration (High-fat diet + STZ + XMU-MP-1) group (n=6). XMU-MP-1 (Selleck, Shanghai, China), dissolved in solvent (2% DMSO [MP Biomedicals, Solon, OH, USA], 30% PEG300 [Selleck], 2% Tween80 [Selleck], and ddH2O), was administered intraperitoneally at a dosage of 1 mg/kg body weight every two days to wild-type diabetic mice induced by a combination of high-fat diet feeding and intraperitoneal streptozotocin injection. The solvent of XMU-MP-1 was used as vehicle control in this experiment. XMU-MP-1 treatment commenced two weeks post intraperitoneal streptozotocin injection and continued for an additional 12 weeks. At the conclusion of the experiments, mice were anesthetized and humanely euthanized by cervical dislocation, and mice samples were collected for further analysis.

first: Y

last: Siyang

first: Z

last: Meng

first: L

last: Bin

first: C

last: Wei


title: Analysis of the effect of PSF on the gene expression profile in neuronal cells

city: Itabashi-ku

first: Ken-ichi

last: Takayama

address: Sakaecho

institute: Tokyo Metropolitan Institute of Gerontology

summary: The multi-domain RNA-binding protein (RBP), polypyrimidine tract-binding protein (PTB)-associated splicing factor (PSF) or splicing factor, proline and glutamine-rich (SFPQ), regulate alternative splicing and transcriptional activation/repression by binding to the target RNA molecules. However, the gene regulatory mechanism of PSF in human neuronal cells is still unclear. We investigated the effect of PSF knockdown on the gene expression profile by RNA-seq using human neuronal cells, SH-SY5Y.

overall_design: We treated SH-SY5Y  cells with siPSF or siControl. After 72 h incubation, RNA-seq analysis was performed.


title: Transcriptome of E17 embryo with mutation in ADCK2 and rescue of muscle alteration by prenatal CoQ10 treatment.

city: Sevilla

first: Daniel Jose

last: Moreno Fernandez-Ayala

address: Carretera de Utrera, Km. 1

institute: Universidad Pablo de Olavide

summary:
A transcriptomic study was performed on muscle, liver and brain a group of 3 embryos with a mutation in ADCK2 gene (controls being wild-type embryos from the same litter).
Haploinsufficiency of ADCK2 leads to reduced mitochondrial coenzyme Q (CoQ) levels in skeletal muscle, resulting in mitochondrial myopathy, alterations in fatty acid beta-oxidation and physical incapacity in both humans and mice at adult stage (PMID: 31480808).
Adck2 heterozygous mice exhibit defects from the embryonic development stage, particularly in skeletal muscle. Prenatal administration of CoQ10 can mitigate the defects observed during embryonic development.
Ablation of Adck2 affects postnatal myogenesis, with more severe consequences in older mice. Consequently, heterozygous Adck2 mice display accelerated muscle wasting associated with ageing. Prenatal administration of CoQ10 can prevent these defects.

overall_design: We used microarrays to detail the global program of gene expression underlying specific mutations in the ADCK2 gene in muscle of E17 embryo. We also analysed liver because its role in fatty acid metabolism, and brain (control, non-afected organ). We identified distinct classes of regulated genes during pathogenesis.

first: Daniel

middle: J

last: Fernández-Ayala

first: Juan

middle: D

last: Hernández-Camacho

first: Plácido

last: Navas


title: Effect of organic fertilizer substitution on nitrogen metabolism of highland barley

city: XiNing

first: JiaHui

last: Yan

state: QingHai

address: Twenty miles of Pu Town, Chengbei District

department: Institute of Plant Protection

institute: College of Agriculture and Forestry Sciences, Qinghai University

summary: To investigate the effects of organic fertilizer replacing chemical fertilizer on the growth and development of barley (Kunlun-14), a pot experiment was conducted. The study examined the impacts of different ratios of organic fertilizer replacing chemical fertilizer nitrogen (0%, 40%, 100%, denoted as OFR0, OFR40, OFR100, respectively) on the growth characteristics, leaf carbon-nitrogen balance, and nitrogen metabolism enzyme activities of barley.

overall_design: Design three different proportions of organic fertilizers replacing chemical fertilizer nitrogen (0%, 40%, 100%), recorded as ORF0, ORF40, and ORF100 respectively. The total application amounts of N, P, and K in each treatment should be kept consistent. In the pot experiment, each flower pot is calculated based on 3.5kg of soil. ORF0 uses urea: 357mg, superphosphate 1545mg, and potassium sulfate 140mg; ORF40 uses urea: 215mg, superphosphate 1443mg, potassium sulfate 122mg, and organic fertilizer 1117mg; ORF100 uses superphosphate. 1289 mg, potassium sulfate 96 mg, and organic fertilizer 2793 mg; the above-ground and underground parts of highland barley were collected during the flowering period for transcriptome sequencing.


title: Cell death at the Solanum lycopersicum and Capsicum graft junction is due to genetic incompatibility

city: Norwich

first: Hannah

middle: Rae

last: Thomas

address: Norwich Research Park, Colney Lane

department: Cell and Developmental Biology

country: United Kingdom

institute: John Innes Centre

summary: Graft compatibility is the ability of two plants to form cohesive vascular connections. Tomato and pepper grafts are incompatible but the underlying causes of this phenomenon remain unknown.  We utilzied a broad array of techniques to profile graft compatibility including viability, biophysical stability, and growth. Cell death in the junction was quantified using trypan blue and TUNNEL assays. Transcriptomic analysis of cell death in the junction was preformed using RNA-sequncing. Finally a meta-transcriptomic analysis was conducted with published datasets to further explore the genetic signature of graft incompatibility.We found that all varieties of pepper tested across two species were incompatible with tomato. Tomato and pepper graft incompatibility is characterized by stem instability, reduced growth, and persistent cell death in the graft junction. We showed that tomato and pepper heterografts have prolonger transcriptional activity, with defense processes highly enrched. We identfied a large subset of NLRs and genes involved in programmed cell death which were upregulated in incompatible tissue. We also identified a set of genes with orthologs in both tomato and pepper which are upregulated in incompatible grafts including biosythesis of steroidal glycoalkaloids. Finally we utilized various biological stressors to explore the genetic signature of grafting. We found a significant overlap in the genetic profile of grafting and plant parsitism. We also identified over 1000 genes uniquely upregulated in incompatible grafting including genes in involved in DNA-damage repair. Based on the broad upregulation of NLRs and genes involved with programmed cell death, prolonged cell death in the junction, and DNA damage, we have determined that tomato and pepper graft incompatibility is likely caused by a form of genetic incompatibility which triggers an autoimmune-like response.

sample_organism:
Capsicum annuum
Solanum lycopersicum

platform_organism:
Capsicum annuum
Solanum lycopersicum

overall_design: The scion and stock of tomato:tomato, pepper:pepper, tomato:pepper, pepper:tomato,  ungrafted tomato, and ungrafted pepper tissue was collected at 7, 14, and 21 DAG. Total RNA was extracted and sequenced.

first: Alice

last: Gevorgyan

first: Alexandra

last: Hermanson

first: Samantha

last: Yanders

first: Matthew

last: Norman

first: Lindsay

last: Erndwein

first: Erin

last: Sparks

first: Maragret

middle: H

last: Frank


title: Antiandrogen treatment endows ZMYDN8 dependent epigenetic reprogramming to orchestrate neuroendocrine tumor transdifferentiation [scRNA-seq]

first: Hanling

last: Wang

address: Shanghai

institute: Shanghai Institute of Nutrition and Health, Chinese Academy of Science

summary: The transdifferentiation from adenocarcinoma following androgen deprivation therapy (ADT) is thought to be the primary process leading to the development of neuroendocrine prostate cancer (NEPC). However, it remains unclear how lineage factors interact with ADT to endow the lineage transition. Through an integrated analysis of NEPC-based CRISPR-Cas9 screening and scRNA-seq tracking of tumor transitions, we unveil that antiandrogen-induced ZMYND8-dependent epigenetic programming orchestrates the transdifferentiation of NEPC. We demonstrate that the ablation of Zmynd8 restricts NEPC development in Pten/Trp53/Rb1 knockout mouse models. Conversely, ASCL1-induced upregulation of ZMYND8 shapes the neuroendocrine (NE) lineage to confer resistance to AR-targeted therapy. Mechanistically, FOXM1, a key regulator in castration-resistant prostate cancer (CRPC), stabilizes ZMYND8 binding to chromatin regions harboring H3K4me1-K14ac modification and FOXM1 targeting. Antiandrogen treatment frees the SWI/SNF chromatin remodeling complex from AR, enabling its interaction with ZMYND8/FOXM1 to upregulate key NE lineage regulators (e.g., FOXA2, SOX2, and POU3F2), thus inducing transdifferentiation. Having identified ZMYND8's link to adverse disease outcomes in CRPC patients, we develop a small molecule, ZMYND8i-34, designed to selectively inhibit its histone recognition. In pre-clinical models, ZMYND8i-34 treatment effectively blocks NE transdifferentiation and curtails CRPC development. Together, our results highlight the importance of antiandrogen treatment endowing ZMYND8-dependent epigenetic reprogramming to orchestrate lineage fate and suggest that targeting ZMYND8 may hold the potential to restrict NEPC development and overcome treatment resistance.

overall_design: To understand the cell lines changes in ZMYND8-OE condition, we treated cell lines with ZMYND8-OE in low MOI of virus and cultured the mixture of cells with CSS (Charcoal-Stripped FBS). Then, we collected the partially ZMYND8 overexpressed cells at three time points in CSS condition, day0, day3 and day30 and used three time points cells (day0, day3 and day30) for scRNA-seq separately.

first: Jun

last: Qin


title: Antiandrogen treatment endows ZMYDN8 dependent epigenetic reprogramming to orchestrate neuroendocrine tumor transdifferentiation [RNA-seq]

overall_design: To understand the cell lines changes in ZMYND8-OE condition, we treated cell lines with or without ZMYND8-OE and cultured with CSS (Charcoal-Stripped FBS) in three time points (day0, day3 and day30), which mimic the ADT progress. SiFOXM1 and inhibitor iZMYND8-34 treatment were used to rescue ZMYND8 function, while the control groups were EV and DMSO. The RNA-seq results of all the groups mentioned above to uncover the function of ZMYDN8 in ADT progress and the way to rescue its function or cure the disease caused by ZMYND8 overexpressed.


title: Gene Expression Analysis of Hypertrophic and Dilated Cardiomyopathy Associated TPM1 mutations in Engineered Heart Tissues

city: New Haven

first: Stuart

last: Campbell

state: CT

address: 55 Prospect Street, Malone 301

department: Biomedical Engineering

institute: Yale University

summary: Hypertrophic and dilated cardiomyopathies (HCM and DCM, respectively) are inherited disorders that may be caused by mutations to the same sarcomeric protein but have completely different clinical phenotypes.  The precise mechanisms by which point mutations within the same gene bring about phenotypic diversity remain unclear. Our objective has been to develop a mechanistic explanation of diverging phenotypes in two TPM1 mutations, E62Q (HCM) and E54K (DCM). Drawing on data from the literature and novel experiments with stem cell-derived cardiomyocytes expressing the TPM1 mutations of interest, we constructed computational simulations that provide plausible explanations of the distinct muscle contractility caused by each variant. In E62Q, increased calcium sensitivity and hypercontractility was explained most accurately by a reduction in effective molecular stiffness of tropomyosin and alterations in its interactions with the actin thin filament that favor the ‘closed’ regulatory state. By contrast, the E54K mutation appears to act via long-range allosteric interactions to increase the association rate of the C-terminal troponin I mobile domain to tropomyosin/actin. These mutation-linked molecular events produce diverging alterations in gene expression that can be observed in human engineered heart tissues. Modulators of myosin activity confirm our proposed mechanisms by rescuing normal contractile behavior in accordance with predictions.

overall_design: Isogenic iPSC cell lines were created with either homozygous E62Q or homozygous E54K TPM1 mutations. These cells were differentiated into cardiomyocytes and seeded onto porcine decellularized myocardium

first: Saiti

last: Halder

first: Michael

last: Rynkiewicz

first: Lynne

last: Kim

first: Lorenzo

last: Sewanan

first: Jeffrey

last: Moore

first: William

last: Lehman


title: Antiandrogen treatment endows ZMYDN8 dependent epigenetic reprogramming to orchestrate neuroendocrine tumor transdifferentiation

city: Shanghai

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

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Refer to individual Series


title: Transcriptional regulation of KCNA2 coding Kv1.2 by EWS::FLI1: involvement in controlling the YAP/Hippo signalling pathway and cell proliferation

city: Nantes

first: Anaïs

last: Postec

address: 1 Rue Gaston Veil

country: France

institute: Nantes University

summary: Ewing sarcoma (ES), the second main pediatric bone sarcoma, is characterised by a chromosomal translocation leading to the formation of fusion proteins like EWS::FLI1. While several studies have shown that potassium channels drive the development of many tumours, limited data exist on ES. This work therefore aimed to study the transcriptional regulation of KCNA2 and define the involvement of the Kv1.2 channel encoded by KCNA2 in a key function of ES development, cell proliferation.

overall_design: Estimated transcripts expression between several Ewing Sarcoma cell line A-673 with a siRNA control and another siRNA targeting the gene KCNA2.

first: Maryne

last: Dupuy

first: Mathilde

last: Mullard

first: Aurélie

last: Chantôme

first: Philippe

last: Hulin

first: Régis

last: Brion

first: Maxime

last: Gueguinou

first: Laura

last: Regnier

first: Marie

last: Potier-Cartereau

first: Bénédicte

last: Brounais-Le Royer

first: Marc

last: Baud’huin

first: Steven

last: Georges

first: François

last: Lamoureux

first: Benjamin

last: Ory

first: Françoise

last: Redini

first: Christophe

last: Vandier

first: Franck

last: Verrecchia


title: Antiandrogen treatment endows ZMYDN8 dependent epigenetic reprogramming to orchestrate neuroendocrine tumor transdifferentiation [ATAC-seq]

first: Hanling

last: Wang

overall_design: ATAC-seq data for LNCap cells with or without ZMYND8-OE uncovered the chromatin openness changes during the ADT progress on ZMYND8-OE condition.

first: Jun

last: Qin


title: RBPMS promotes contractile smooth muscle splicing and alters phenotypic behaviour of human embryonic stem cell derived vascular smooth muscle cells.

city: Cambridge

first: Irina

last: Mohorianu

address: Puddicombe Way

department: Wellcome-MRC Cambridge Stem Cell Institute

institute: University of Cambridge

summary: Differentiated Vascular Smooth Muscle Cells (VSMCs) express a unique network of splice isoforms (smooth muscle specific alternative splicing - SM-AS) in functionally critical genes including those comprising the contractile machinery. We previously described RNA Binding Protein Multiple Splicing (RBPMS) as a potent driver of contractile, aortic tissue like SM-AS in VSMCs using rodent models. What is unknown is how RBPMS affects VSMC phenotype and behaviour. Here, we use human embryonic stem cell-derived VSMCs (hES-VSMCs) to dissect the role of RBPMS in SM-AS in human cells and determine the impact on VSMC phenotypic properties. hES-VSMCs are inherently immature and display only partially differentiated SM-AS patterns while RBPMS levels are undetectable endogenously. Hence, we used an over-expression system and found that RBPMS induces SM-AS patterns in hES-VSMCs akin to the contractile tissue VSMC splicing patterns in multiple events. We present in silico and experimental findings that support RBPMS’ splicing activity as mediated through direct binding and via functional cooperativity with splicing factor RBFOX2 on a significant subset of targets. Finally, we demonstrate that RBPMS can alter the motility and the proliferative properties of hES-VSMCs to mimic a more differentiated state. Overall, this study emphasizes a critical splicing regulatory role for RBPMS in human VSMCs and provides evidence of phenotypic modulation by RBPMS.

overall_design: Control or anti-RBFOX2 siRNAs were used

first: Aishwarya

last: Jacob

first: Ilias

last: Moutsopoulos

first: Alex

last: Petchey

first: Rafael

last: Kollyfas

first: Sanjay

last: Sinha

first: Cristopher

last: Smith


title: Study on the mechanisms of Piezo1-lars2 signal path mediating the mechanical loading on endochondral ossification process of fracture healing

first: Siming

last: Jia

address: 河北省石家庄市河北医科大学第三医院

summary: In China, the incidence of fracture non-unions is relatively high. Impaired endochondral ossification may lead to nonunion due to improper mechanical loading. As a mechanosensitive ion channel protein, Piezo1mediates mechanical transduction and induces calcium inward flow. Our early study showed that the osteogenic and angiogenic capacity of chondrocytes was reduced, if Piezo1 gene was knocked out on chondrocytes. Moreover, the expression of mitochondrion translation related gene LARS2 was signaling increased. Therefore, we hypothesize that the mechanical loading may cause endochondral ossification through the Piezo1- LARS2 signaling pathway. We will use conditioned gene knockout mice and Peizo1 knockdown stable cell line to support the hypothesis. Our goals include investigating the changes of Piezo1 expression during endochondral ossification of femoral fracture healing in mice under mechanical loading; investigating the response of Piezo1 channels on chondrocytes to mechanical stimulation and its role and mechanism in regulating chondrocyte osteogenesis and angiogenesis, maintaining the normal mitochondrial function in vitro; analysing the key molecular and signal network downstream of Piezo1 through the multi-omics techniques; and exploring the response mechanism of Piezo1 under vibrating stimulus. This project aims to study the molecular mechanism of Piezo1-mediated force-biological information transition and its role in endochondral ossification. We hope it provides an effective intervention target for preventing and treating fracture nonunion.

overall_design: three samples from the callus tissue of  Col2CreERT; Piezo1-/- and wt mice

first: Weijian

last: Liu

first: Wei

last: Chen

first: Yingze

last: Zhang


title: Antiandrogen treatment endows ZMYDN8 dependent epigenetic reprogramming to orchestrate neuroendocrine tumor transdifferentiation [ChIP-seq]

overall_design: ChIP-seq data for LNCap cells with or without ZMYND8-OE uncovered ZMYND8 binding places change during the ADT progress on ZMYND8-OE condition.


title: A post-transcriptional regulatory checkpoint restrains acquisition of cytotoxic activity in tumor-infiltrating CD4+ T cells

city: London

first: Maria

last: Vila de Mucha

address: 72 Huntley Street

institute: UCL Cancer Institute

summary: Tumor-infiltrating cytotoxic CD4+ T cells (TCTX) are a promising therapeutic target, but how their activity is regulated is poorly understood. Here, we show that a post-transcriptional checkpoint restrains TCTX activity in suppressive tumor microenvironments. In untreated tumors, poised TCTX express high levels of Gzmb mRNA but produce little Granzyme B (GzmB) protein. Differentiation into poised TCTX is regulated by the Blimp-1-Bcl6 axis and requires type I interferon signaling. GzmB protein is produced by tumor-infiltrating TCTX upon treatment with TREG-depleting aCTLA-4 antibodies and following blockade of LAG-3 and PD-1. These treatments downregulate Zfp36l1, encoding an RNA binding protein. Deletion of Zfp36l1 and its paralog Zfp36 increases GzmB protein in CD4+ T cells in untreated tumors and reduces tumor mass. These results demonstrate that TCTX activity is restrained in tumors through LAG-3 and PD-1 and the ZFP36 family of RNA binding proteins. We propose that these pathways represent targets for therapeutic intervention.

overall_design: We wished to analyse the transcriptome of tumor infiltraing CD4+ T cells in the murine tumor model MCA205, in both untreated and anti-CTLA-4 treated conditions. To do so, we carried out single cell RNA sequencing of FACS sorted CD3+ T cells taken from MCA205 tumors treated with  anti-CTLA-4 on days 6, 9 and 11 post tumor inoculation or left untreated.

first: María

first: Richard

last: Jenner

first: Sergio

last: Quezada


title: Effect of depletion of TAS1R3 on testicular gene expression in mice fed long-term western diet

city: Seoul

first: Hobin

last: Seong

state: KOREA, REPUBLIC OF

address: Gwanak-ro 1, Seoul National University

country: South Korea

institute: Seoul National University

summary: Recent studies suggest that the western diet negatively affects male reproduction, but few underlying molecular mechanisms have been established. Also, several studies have reported TAS1R3 is expressed in testis, epididymis and sperm, but their role in male reproduction remains unknown. To investigate the function of taste receptor TAS1R3 in the male reproductive function under western diet condition, we performed RNA-sequencing analysis on testes from C57BL/6 mice fed long-term (18 weeks) western diet.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for testes from Tas1r3+/+ and Tas1r3-/- mice fed long-term western diet

first: Woo-Jeong

last: Shon

first: Dong-Mi

last: Shin


title: HSD17B13 liquid-liquid phase separation activates autocrine platelet-activating factor signaling in NASH

city: Nan jing

first: jing

last: ye

address: Jiangsu Province Nanjing,Qixia District, Xianlin Road No.163

institute: Nanjing University

summary: Loss-of-function variants in 17-beta hydroxysteroid dehydrogenase 13 (HSD17B13) are associated with decreased inflammation in human chronic liver disease. The underlying mechanism by which HSD17B13 promotes liver inflammation remains largely elusive. Here we find that HSD17B13 undergoes liquid-liquid phase separation (LLPS) around lipid droplets (LDs) in the liver of NASH patients. Dimerization of HSD17B13 drives LLPS formation and regulates its enzyme activity. HSD17B13 LLPS promotes leukocyte adhesion and fibrinogen expression in hepatocytes via activating autocrine platelet activating factor (PAF) signaling. Importantly, adeno-associated viral (AAV)-mediated xeno-expression of human HSD17B13 promotes liver inflammation in Hsd17b13-/- mice induced by high fat diet/carbon tetrachloride treatment. In conclusion, HSD17B13 LLPS triggers liver inflammation via promoting PAF-mediated leukocyte adhesion. Targeting HSD17B13 phase transition would be a promising approach to treat liver inflammation in chronic liver disease.

overall_design: For HepaRG cell lines, cells were divided into two groups, including HepaRG-NC and HepaRG-HSD17B13 overexpression group. Cells were calculated and RNA samples were extracted from 1 × 10^6 HepaRG cells using trizol. CDNA libraries were prepared using TruSeq PE Cluster Kit v3-cBot-HS (Illumina) according to the manufacturer’s instructions. Murine hepatocytes were divided into NC and human HSD17B13 overexpression groups. Cells were calculated and RNA samples were extracted for RNA seq.


title: Hypoxia-responsive ZEB2 regulates a network of calcium handling genes in the injured heart

city: Utrecht

first: Cornelis

middle: J.

last: Boogerd

address: Uppsalalaan 8

department: Developmental Biology and Stem Cell Research

institute: Hubrecht Institute

summary: Intracellular calcium (Ca2+) overload is known to play a critical role in the development of cardiac dysfunction. Despite the remarkable improvement in managing the progression of heart disease, developing effective therapies for heart failure (HF) remains a challenge. A better understanding of molecular mechanisms that maintain proper Ca2+ levels and contractility in the injured heart could be of therapeutic value.  Here, we report that the transcription factor Zinc finger E-box-binding homeobox2 (ZEB2) is induced by Hypoxia-inducible factor 1-alpha (HIF1a) in hypoxic cardiomyocytes and regulates a network of genes involved in Ca2+-handling and contractility during ischemic heart disease. Gain- and loss-of-function studies in genetic mouse models revealed ZEB2 in cardiomyocytes to be necessary and sufficient to protect the heart against ischemia-induced diastolic dysfunction and structural remodeling. Moreover, RNA sequencing (RNA-seq) of ZEB2-overexpressing (Zeb2 cTg) hearts post-injury implicated ZEB2 in regulating numerous Ca2+-handling and contractility-related genes. Mechanistically, ZEB2 overexpression increased the phosphorylation of phospholamban (PLN) at both serine-16 and threonine-17, implying enhanced activity of sarcoplasmic reticulum Ca2+-ATPase (SERCA2a), thereby augmenting SR Ca2+ uptake and contractility. Furthermore, we observed a decrease in the activity of Ca2+-dependent calcineurin/NFAT signaling in Zeb2 cTg hearts, which is the main driver of pathological cardiac remodeling. On a post-transcriptional level, we showed that ZEB2 expression can be regulated by the cardiomyocyte-specific microRNA-208a (miR-208a). Blocking the function of miR-208a with antimiR-208a increased ZEB2 expression in the heart and effectively protected from the development of pathological cardiac hypertrophy. Together, we present ZEB2 as a central regulator of contractility and Ca2+-handling components in the mammalian heart. Further mechanistic understanding of the role of ZEB2 in regulating Ca2+ homeostasis in cardiomyocytes is an essential step towards the development of improved therapies for HF.

overall_design:
To investigate the effect of ischemic injury on cardiac gene expression, we performed Ischemia/Reperfusion (IR) injury and isolated infarct area plus border zone tissue from hearts 14 days after injury. RNA-seq was performed on tissue from three animals per group, from Sham treated WT C57Bl/6J and 14 days post IR injury.
To investigate the effect of ZEB2 overexpression, we performed IR injury on ZEB2 overexpressing hearts and wildtype littermate controls and harvested infarct area plus border zone 14 days after injury. RNA-seq was performed on tissue from three animals per group.

first: Monika

last: Gladka


title: The intrinsically disordered region of SPIN1 regulates its function by phase separation [RNA-seq]

city: baoding

first: yukun

last: wang

state: hebei

address: hebei

institute: hebei university

summary: To further assess the role for IDR in target gene regulation, we performed transcriptome analysis by RNA sequencing (RNA-seq).

overall_design: Comparative gene expression  analysis of RNA-seq data for overexpression-SPIN1-WT and overexpression-SPIN1-△IDR

first: Yukun

last: Wang

first: Chen

last: Wu


title: The intrinsically disordered region of SPIN1 regulates its function by phase separation.


title: The intrinsically disordered region of SPIN1 regulates its function by phase separation [ChIP-seq]

summary: As a transcriptional coactivator, SPIN1 has been reported to bind to chromatin.To assess the effect of IDR on chromatin binding, we investigated genome-wide chromatin association of SPIN1-WT or SPIN1-△IDR stably overexpressed cell lines by chromatin immunoprecipitation followed by high-throughput sequencing.

overall_design: Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for  the histone modifications H3K4me3 and FLAG in overexpression-Vector ,overexpression-SPIN1-WT or overexpression-SPIN1-△IDR SGC7901 cells.


title: Single cell transcriptomic profiling of corneal epithelium of diabetic non-human primates reveals that FOXO3 regulates ROS by reprogramming glucose metabolism

city: Stockholm

first: Jiawei

last: Zhong

address: Blickagången 16, Flemingsberg

department: Department of Medicine, Huddinge

country: Sweden

institute: Karolinska Institutet

summary: Hyperglycemia leads to functional decline of various corneal cells, which induces corneal diseases. However, cellular and molecular mechanisms underlying how hyperglycemia affects the corneal epithelium have not been determined. Here, by undertaking single-cell transcriptomics of corneal epithelial cells obtained from healthy and diabetic non-human primates, we identified cell type-specific transcriptional alterations and shared changes across cell types, such as upregulation of FOXO3. Targeted depletion of FOXO3 in human corneal epithelial cells regulated metabolic remodeling by inhibiting glycolysis and oxidative phosphorylation. Furthermore, FOXO3 can not only sense reactive oxygen species (ROS) in corneal epithelial cells and scavenge ROS, but also mediate the proliferation and migration of corneal epithelial cells, demonstrating FOXO3 as a guardian factor in maintaining primate corneal epithelial homeostasis. Our study provides novel insight into the role of FOXO3 in metabolic remodeling and maintenance of ROS homeostasis in corneal epithelium at single-cell level, laying a foundation for decoding the molecular mechanism of the diagnosis and treatment of diabetic eye disease.

sample_organism:
Macaca fascicularis
Homo sapiens

platform_organism:
Macaca fascicularis
Homo sapiens

overall_design:
scRNA-seq of corneal epithelium of healthy and diabetic crab-eating monkey;
RNA-seq of FOXO3 knowndown and control human corneal epithelial cells (HCEC)


title: miRNAs have a role on Prunus persica response to topic application of PpPep2 endogenous peptide

city: Girona

middle: Pla

last: de Solà-Morales

state: Catalonia

address: Campus Montilivi (EPS-1)

department: Institute for Agricultural and Food Technology

institute: Universitat de Girona

summary: We charactarize the regulated miRNAs of Prunus persica to preventive treatment of PpPep2 1 µM endogenous peptide. The peptide treatment was applied to leaves and we sequenced the miRNAs after 1 and 24 h using untreated plants as control. miRNA profiles were generated by deep sequencing, in triplicate, using Illumina HiSeq 2500. Differential expression analysis of annotated pre_miRNA gene biotypes was performed with R package DESeq2 v1.26.0, considering significant epression values below FDR <0.05. We obtained 117,267,647 raw reads and we identified a total of 33 differential expressed miRNAs (DEM) belonging to 18 miRNA families upon PpPep2 application, accounting for 15% of the P. persica annotated miRNAs. miRNA regulation occurs principally one day after peptide treatment. The predicted mRNA targets and function of orthologous miRNAs are compatible with a regulation of PTI-related processes. Together with the transcriptomic response of P. persica to PpPep2 described at our previous RNA-Seq (Foix et al 2021), this suggests that miRNAs would have an active role in PTI regulation at these stages.

overall_design: miRNA profiles of  P. persica pretreated with PpPep2 after 1 and 24 h (t1, t24 h), untreated plants as control (t0).

last: Pla

first: Laura

middle: F

last: Pericot

first: Anna

last: Esteve-Codina


title: Transcriptomes of WT and cmnac-nor mutant lines during melon fruit development and ripening

city: Beijing

first: Jinfang

address: No.50 Zhanghua Road, Haidian District

institute: Beijing Academy of Agricultural and Forestry Sciences

summary: Analysis of gene expression level. The hypothesis tested in the present study that CmNAC-NOR positively regulated melon fruit development and ripening.

overall_design: mRNA profiles of  wild type melon(WT, Cucumis melo L.) and cmnac-nor mutant fruits at15 DAP,25 DAP, 35 DAP and 45 DAP were generated by deep sequencing, in triplicate, using Illumina Novaseq 6000.

first: Yong

last: Xu


title: Transcription start site profiling of HCMV infected cells using dRNA-seq and STRIPE-seq with temporal kinetics using SLAM-seq

city: Würzburg

first: Christopher

middle: Sebastian

last: Jürges

state: BY

address: Versbacher Straße 7

department: Institut für Virologie und Immunbiologie

country: Germany

institute: Julius-Maximilians-Universität Würzburg

summary: Primary human foreskin fibroblasts (HFF) were infected with wild-type HCMV strain TB40 or BAC2 at a multiplicity of infection (MOI) of 10. Cells were treated with 4sU 1h before harvesting. Sequencing libraries were treated with IAA and further prepared using the dRNA-seq and STRIPE-seq protocol from the indicated timeoints of infection

overall_design: HFF were infected with HCMV strain TB40 or BAC2. Cells were treated with 4sU as extracted RNA with IAA using the SLAM-seq protocol. Transcription start sites were enriched using the dRNA-seq and STRIPE-seq protocol on total cellular RNA. Two independent biological replicates were analysed.

middle: S

first: Manivel

last: Lodha

first: Mirko

last: Trilling

first: Pranjali

last: Bhandare

first: Elmar

last: Wolf

first: Albert

last: Zimmermann

first: Khanh

last: Le-Trilling

first: Lars

last: Dölken

first: Florian

last: Erhard


title: Ahulkenoids restore multiple phenotypes in vitro and in vivo in Myotonic Dystrophy type 1

city: San Sebastián

first: MIKEL

last: GARCIA PUGA

address: Paseo Dr. Begiristain sn

institute: Biodonostia HRI

summary: Myotonic dystrophy type 1 is an autosomal dominant disorder without available treatment to slow or stop disease progression nowadays. It is clinically characterized by a progressive muscular weakness and multisystem degeneration. In this study, we characterized the impact of a new family of FKBP12 “reshapers”   (known as ahulkenoids). Treatment with different ahulkenoids reversed the accumulation of oxidative stress, the impaired cell viability and proliferation as well as mitochondrial activity and metabolism defects in DM1 derived primary fibroblasts. Moreover, RNAseq analysis confirmed the restoration of molecular pathways related to cell cycle and cell metabolism. Importantly, treatment with ahulkenoids significantly maintained locomotor activity and extended lifespan of a Drosophila model of the DM1 disease. In summary, our results revealed the benefits of a new family of compounds, entitled ahulkenoids, in the pre-clinical setting of DM1

overall_design: DM1-derived fibroblasts treated with 0.1 µM AHK7 for 72 h.

first: Mikel

last: García-Puga

first: Gorka

last: Guereñu

first: Ariadna

last: Bargiela

first: Jorge

last: Espinosa-Espinosa

first: Jesús

middle: M

last: Aizpurua

first: Ainara

last: Vallejo-Illarramendi

first: Rubén

last: Artero

first: Adolfo

last: López de Munain

first: Ander

last: Matheu


title: Heterogeneity of Active Mast Cells, Endothelial Cells, and Fibroblasts: Unique Transcriptomic Features of Synovial Single-Cell Sequencing in Hemophilic Arthritis

status: Public on Dec 18 2024

city: Guangzhou

first: Kun

last: Lin

address: NO.261, Longxi Road, Liwan District

institute: Guangzhou University of Chinese Medicine

summary: Purposes Recurrent synovial bleeding is the major cause of Hemophilic Arthritis (HA). Up to now, the transcriptomic profiles of bleeding syvonium in HA patient have largely remained unknown. Exploring the transcriptomic characteristics of synovium obtained from HA patients undergoing total knee arthroplasty (TKA), and uncovering potential pathological mechanisms of HA synovium through bioinformatics analysis and differential analysis. Methods Single-cell RNA sequencing (scRNA-seq) technology was utilized to identify distinct cellular subsets within HA synovium. Comparative analysis was conducted with scRNA-seq data from osteoarthritis (OA) and rheumatoid arthritis (RA) synovium to assess transcriptional differences. Histological evaluation, immunofluorescence (IF), immunohistochemistry (IHC) and in vitro cell testing were performed for validation. Results We observed that the single-cell transcriptomic characteristics of HA synovium differ significantly from those of OA and RA patients. Mast cells, identified as unique immune cells in HA synovium, are actively involved and may be the initiating factor for changes in vascular permeability or bleeding in HA synovium. The transcriptomic features of endothelial cells and fibroblasts, which are non-immune stromal cells, reveal the synovial microenvironment: repeated bleeding leads to iron deposition, which causes substantial ferroptosis within the synovium. Based on the pseudotime analysis of endothelial cells, along with IHC staining and in vitro cell assays, it was demonstrated that ferroptosis in endothelial cells induces vascular damage and triggers a significant migration of endothelial cells. The stress of ferroptosis leads to the differentiation of HMOX1+ endothelial cells. Cell interaction analysis shows that HMOX1+ endothelial cells can induce the SCARA5+ fibroblast subtypes via FTL/SCARA5. This may be a critical target for alleviating cellular ferroptosis in HA synovium. Conclusions Through scRNA-seq, we elucidated the heterogeneity of synovial tissues in three diseases. Activated mast cells around blood vessels may be potential target cells for bleeding and rebleeding, with TPSB2 emerging as a potential target. We also identified endothelial cell subpopulations under ferroptosis stress and their interactions with fibroblasts. Notably, we discovered a SCARA5+ fibroblast subpopulation that regulates ferroptosis, which could serve as a potential target gene for preventing and treating recurrent bleeding in HA.

overall_design: Adjust the cell concentration to the ideal range of 700-1200 cells/μL. Gel beads containing barcode information are mixed with cells and enzymes. Then encapsulated in oil surfactant droplets within a microfluidic system, forming Gel Beads-In-Emulsions (GEMs). GEMs flow into a collection reservoir where gel beads dissolve, releasing barcode sequences, reverse transcribing cDNA fragments, and labeling the samples. Gel beads are ruptured and oil droplets shattered to PCR amplify cDNA using it as a template. The products from all GEMs are mixed to construct a standard sequencing library. Initially, cDNA is enzymatically fragmented into segments around 200-300 bp, followed by traditional second-generation sequencing library construction processes like adding sequencing adapters and primers, and finally PCR amplification to obtain a DNA library. High-throughput sequencing of the prepared library is performed using Illumina sequencing platform's paired-end sequencing mode. After obtaining raw data, the official analysis software Cell Ranger from 10x Genomics (https://support.10xgenomics.com/single-cell-gene-expression/software/overview/welcome) is used for data filtering, alignment, quantification, cell identification, and obtaining gene expression matrices for each cell.


title: TFEB controls expression of human syncytins during cell-cell fusion [RNA-Seq]

city: Berkeley

first: Meagan

middle: Nicole

last: Esbin

state: California

address: 475 Li Ka Shing Center

department: Molecular and Cell Biology

institute: University of California Berkeley

summary: During human development, a subset of differentiating fetal cells form a temporary organ, the placenta, which invades the uterine wall to support nutrient, oxygen, and waste exchange between the mother and fetus until birth. Most of the human placenta is formed by a syncytial villous structure which arises via cell-cell fusion of underlying fetal trophoblast stem cells. Genetic and functional studies have characterized the membrane protein fusogens, Syncytin-1 and Syncytin-2, that are both necessary and sufficient for human trophoblast cell-cell fusion. However, identification and characterization of upstream transcriptional regulators regulating their expression has been limited. Here, using CRISPR knockout in an in vitro cellular model of syncytiotrophoblast development (BeWo cells), we find that the transcription factor TFEB, mainly known as a regulator of autophagy and lysosomal biogenesis, is required for cell-cell fusion of syncytiotrophoblasts. TFEB translocates to the nucleus, exhibits increased chromatin interactions, and directly binds the Syncytin-1 and Syncytin-2 promoters to control their expression during differentiation. While TFEB appears to play an important role in syncytiotrophoblast differentiation, ablation of TFEB largely does not affect lysosomal gene expression or lysosomal biogenesis in differentiating BeWo cells, suggesting that TFEB plays an alternative role in placental cells.

overall_design: We examined the differential expression of genes using RNASeq to determine the effects of TFEB on syncytiotrophoblast differentiation using BeWo cells. We compared gene expression in wild-type versus TFEB/TFE3 double KO or TFEB KO BeWo cells after no treatment, Forskolin-induced differentiation, or after treatment with the mTOR inhibitor Torin1.

middle: N

first: Xavier

last: Darzacq

first: Robert

last: Tjian


title: tRF-Val-TAC-004 protects against renal ischemia-reperfusion injury via attenuating Apaf1-mediated apoptosis

city: Changsha

first: Ni

last: Yin

state: Hunan

address: Yin Penling Street

institute: The Third Xiangya Hospital, Central South University

summary: tRNA-derived fragments (tRFs) play critical roles in cellular process, and we have previously reported that tRFs are involved in ischemia reperfusion injury induced acute kidney injury (IRI-AKI). However, the precise involvement of tRFs in IRI-AKI remains obscure. This study aims to elucidate the impact of tRF-Val-TAC-004 (tRF-Val) on IRI-AKI and uncover the underlying mechanisms. Our observations reveal a significant downregulation of tRF-Val in IRI-AKI mice and its overexpression mitigated renal dysfunction, morphological damage, and apoptosis in IRI-AKI mice, while its inhibition exacerbated these effects. Similar outcomes were replicated in CoCl2-treated BUMPT cells upon transfection with tRF-Val mimic or inhibitor. Mechanistically, dual-luciferase reporter assay and AGO-RIP qPCR analyses demonstrated that tRF-Val suppresses Apaf1 expression by targeting the 3’-UTR of Apaf1 mRNA. Furthermore, the protective efficacy of tRF-Val was notably weakened by Apaf1-overexpressing plasmids. In summary, these novel findings unveil the protective role of tRF-Val against IRI-AKI through inhibition of Apaf1-mediated apoptosis.

overall_design: To investigate the apototic downstream target gene of tRF-Val-TAC-004 in BUMPT cells

first: Wei

last: Zhang

first: Hao

first: Xian

last: Xie


title: Next Generation Sequencing For Quantitative Analysis Of Hepatic Transcriptomes In Response To Squalene

city: Zaragoza

first: Roubi Hamid

middle: Ahmad

last: AbuObeid

state: Spain

address: calle de Miguel Servet, 177

institute: university of Zaragoza

summary:
Purpose: The goal of this study was to determine squalene effect on hepatic transcriptome by comparing high-throughput data for tested group vs. control group in Wildtype New Zealand white rabbits.
Method: Hepatic mRNA profile of two Wildtype New Zealand white rabbits groups fed over 4 weeks with  1% of sunflower oil and the second fed with purified 1% of sunflower oil and 0.5% of squalene was analysed by Transcriptome sequencing using DNBseq platform. Sequence reads that passed quality filters were mapped onto reference genome, followed by SNP & INDEL calling and Gene expression detection.
Results:   Squalene intake did not significantly influence single nucleotide polymorphisms.   When alternative splicing events were tested, squalene administration had significant influence on splicing events, alternative 5' splicing, alternative 3' splicing Site and retained intron while no effect was found on skipped exons or mutually exclusive exons.  Splicing patterns led to variety of differentially splicing genes and a variety of different isoforms from one gene. Differentially expressed genes were 18,485 in the squalene group and 18,286 in the control group.
Conclusions: This study represents the first analysis of hepatic transcriptome in respond to squalene. Result data confirm that RNA-Sequencing is a sensitive and accurate tool that provides a framework for comparative investigations of known and novel DEGs.

overall_design: Testing the influence of 0.5% of squalene on hepatic gene expression in Wildtype New Zealand white rabbits groups fed with diet over 4 weeks

first: Roubi

last: Abuobeid

first: Roberto

last: Martínez-Beamonte

first: Jesus

last: Osada


title: Single Cell RNAseq of CD8 positive Lymphocyte T from Lung Tumor in Human patients

city: Paris

first: Sebastien

last: Mella

address: 28 rue du Dr Roux

department: Immunology

institute: Institut Pasteur

summary: Analysis at the single cell level of the CD8+ Lymphocytes T extracted from Non Small Cell Lung Tumor  patients after surgery and before chemoimmunotherapy

overall_design: CD8+T cells were sorted and then submitted to Single Cell RNAseq performed using the 10X genomics platform

first: Milena

last: Hasan

first: Bernd

last: Jagla

first: Valentina

last: Libri


title: Inhibition of the chemokine receptors CXCR1 and CXCR2 synergizes with docetaxel for effective tumor control and remodeling of the immune microenvironment of HPV-negative head and neck cancer models [mouse]

city: Bethesda

first: Thomas

middle: Joshua

last: Meyer

address: 41 Center Drive, Building 41, Room B620

department: National Cancer Institute

institute: National Institutes of Health

summary: Relapsed head and neck squamous cell carcinoma (HNSCC) unrelated to HPV infection carries a poor prognosis. Novel approaches are needed to improve the clinical outcome and prolong survival in this patient population which has poor long-term responses to immune checkpoint blockade. This study evaluated the chemokine receptors, CXCR1 and CXCR2, as potential novel targets for the treatment of HPV-negative HNSCC. Expression of IL-8, CXCR1, and CXCR2 was investigated in HNSCC tissues and human cell line models. Inhibition of CXCR1/2 with the clinical stage, small molecule inhibitor, SX-682, was evaluated in vitro and in vivo using human xenografts and murine models of HNSCC, both as a monotherapy and in combination with the taxane chemotherapy, docetaxel. High levels of IL-8, CXCR1, and CXCR2 expression were observed in HPV-negative compared to HPV-positive HNSCC tumors or cell lines. Treatment of HPV-negative HNSCC cell lines in vitro with SX-682 sensitized the tumor cells to the cytotoxic activity of docetaxel. In vivo, treatment of HNSCC xenograft models with the combination of SX-682 plus docetaxel led to strong anti-tumor control resulting in tumor cures. This phenomenon was associated with an increase of microRNA-200c and a decreased expression of its target, tubulin beta-3, a protein involved in resistance to microtubule-targeting chemotherapies. In vivo treatment of a murine syngeneic model of HNSCC with SX-682 plus docetaxel led to potent anti-tumor efficacy through a simultaneous decrease in suppressive CXCR2+ polymorphonuclear, myeloid-derived suppressor cells and an increase in cytotoxic CD8+ T cells in the combination therapy treated tumors compared to controls. This study reports, for the first time, mechanistic findings through which the combination of CXCR1/2 inhibition and docetaxel chemotherapy exhibits synergy in models of HPV-negative HNSCC. These findings provide rationale for the use of this novel combination approach to treat HPV-negative HNSCC patients and for future combination studies of CXCR1/2 inhibition, docetaxel, and immune-based therapies.

overall_design: MOC1 - To investigate the effect of docetaxel plus CXCR1/CXCR2 inhibition on the murine MOC1 HNSCC model, control, SX-682, docetaxel, and SX-682 plus docetaxel treated MOC-1 tumors from C57BL/6 mice were collected, RNA was purified from a piece of total tumor tissue, and RNA sequencing was performed.

first: Lucas

middle: A

last: Horn

first: Claudia

last: Palena


title: Inhibition of the chemokine receptors CXCR1 and CXCR2 synergizes with docetaxel for effective tumor control and remodeling of the immune microenvironment of HPV-negative head and neck cancer models

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

overall_design: Refer to individual Series


title: CAR T Cells Secreting NGF-Neutralizing scFv Enhance Efficacy in Clear Cell Renal Cell Carcinoma by Relieving Immunosuppression through Immunosympathectomy [scRNA-seq]

city: Nanjing

first: peiwei

last: yang

state: Jiangsu

address: 639 Longmian Avenue

institute: China pharmaceutical university

summary: This study investigated the role of sympathetic denervation in enhancing CAR T cell anti-tumor efficacy in renal clear cell carcinoma. By blocking the NGF pathway with antibodies, we demonstrated reduced sympathetic nerve distribution and delayed tumor progression. Furthermore, CAR T cells engineered to secrete NGF scFv achieved tumor immunosympathectomy, leading to improved anti-tumor effects. RNA sequencing revealed that this enhancement was linked to reduced terminal exhaustion in CD8 T cells and inhibited macrophage polarization from M1 to M2, promoting a robust anti-tumor immune state. Additionally, splenic T cells exhibited a stronger immune effector phenotype following the infusion of NGF scFv-secreting CAR T cells. These findings suggest that immunosympathectomy may represent a novel strategy to mitigate tumor-induced immunosuppression and improve CAR T cell efficacy in solid tumors.

overall_design: Tumor cells were inoculated into the left axilla of Balb/c mice. When tumors reached approximately 80 mm3, mice were intraperitoneally injected with 200 mg/kg cyclophosphamide. Two days later, the mice were randomly divided into three groups and intravenously infused with either UT, 1×107 V28z, or 1×107 V28z/αNGF cells. 500,000 U of human recombinant IL-2 was administered intraperitoneally, followed by continuous injections twice daily for two days.


title: Inhibition of the chemokine receptors CXCR1 and CXCR2 synergizes with docetaxel for effective tumor control and remodeling of the immune microenvironment of HPV-negative head and neck cancer models [human]

overall_design: UM-SCC-74B - To investigate the effect of CXCR1/CXCR2 inhibition on the UM-SCC-74B HNSCC model, control and SX-682 treated xenografts from NSG mice were collected, RNA was purified from a piece of total tumor tissue, and RNA sequencing was performed.


title: Integrative analysis of transcriptome and metabolism reveals functional roles of redox homeostasis in low light and salt combined stress in Leymus chinensis

city: Yangzhou

first: Mingnan

last: Qu

address: Pazhou Dadao Rd, 17-19

institute: Yangzhou Univerisity

summary: Soil salinity (Salt) is one of the major limiting factors of Leymus chinensis (named sheepgrass) performance and alters their primary metabolic process and gene expression. Salt stress could accelerate inhibitive effects concomitant with low light (LL-Salt). However, little is known about physiological and molecular mechanisms under such LL-Salt in sheepgrass. This study aims to uncover the key reprogrammed metabolic pathways induced by LL-Salt through an integrated analysis of transcriptome and metabolism. Plant seedlings were exposed to six combinations of light intensity (moderate light, ML, and LL) and NaCl concentrations (0, 50, and 200mM) for 20 days. Results suggest that the growth of sheepgrass seedlings was dramatically inhibited with severe lodging phenotype, especially when exposed to LL-Salt combined conditions. Activities of an antioxidant enzyme, catalase, were significantly increased in LL but significantly decreased in salt stress, leading to a reverse pattern for H2O2 in LL and salt stress. Transcriptome analysis reveals 4921 downregulated differentially expressed genes (DEGs) with carbon metabolism pathways were significantly enriched compared to light response without salt (ML_0 vs LL_0). There are 194 overlapped downregulated DEGs induced by two salt treatments (ML_50 vs ML_0 and ML_200 vs ML_0), where the carbon metabolism pathway was also significantly enriched. In terms of interactive effects of LL-Salt treatments, we found that there are 16 overlapped DEGs, including a phytochrome-interacting transcription factor 4 (PIF4) with upregulation in LL treatment while downregulation in salt treatment, which was validated by qPCR.Collectively, we found that serval antioxidant redox pathways, including photorespiration, GSG/GSSH redox, and ABA signaling, participated in response to LL and salt combined events and highlighted the roles of cellular redox homeostasis in LL-Salt response in sheepgrass.

overall_design: To better identify the key regulatory genes in sheepgrass induced by salt and low light, we carried out transcriptome deep sequencing of sheepgrass leaves from exposed to either salt stress and low light

first: Jikai

last: Li


title: An extra-genital cell population contributes to urethra closure during mouse penis development

city: Durham

first: Humphrey

middle: Hung-Chang

last: Yao

state: NC

address: 111 TW Alexander Drive

institute: NIEHS/NIH

summary: Penis, the organ that bears reproductive and psychological importance, is susceptible to birth defects such as hypospadias, or incomplete closure of urethra along the penis shaft. We discover that proper urethral closure in mouse embryos requires a unique mesenchymal cell population originated from outside of the penis. These “extra-genital” cells, marked by a lineage marker Nr5a1, migrate from the inguinal region into the embryonic penis, and facilitate urethra closure by interacting with adjacent peri-urethral cells via the epidermal growth factor pathway. Ablation of Nr5a1+ cells leads to severe hypospadias, and alters cell differentiation in the penis. This discovery highlights the indispensable role of Nr5a1+ extra-genital cells in urethra closure, shedding light on the biology of penis formation and potential implications for human hypospadias.

overall_design: To investigate the transcripitional changes during caused by the lost of Nr5a1+ cells, we conducted single cell mRNAseq on penises collected at embryonic day 16.5. Comparisons were made between Nr5a1cre+ x DTA f/+ (DTA+) and Nr5a1cre- x DTA f/+ (DTA-) genitalia.

first: Ciro

last: Amato

first: Xin

last: Xu


title: Transcriptome analysis of jejunal mucosal tissue in breeder hens exposed to acute heat stress

city: Nakhon Ratchasima

first: Yongcai

last: Zhu

address: School of Animal Technology and Innovation, Institute of Agricultural Technology, Suranaree University of Technology, Nakhon Ratchasima, 30000, Thailand

country: Thailand

institute: Suranaree University of Technology

summary: Heat stress (HS) can damage the integrity of the intestinal mucosal barrier, leading to decreased poultry productivity. This study aimed to identify candidate genes related to acute HS in breeder hens and provide insight into the molecular mechanisms underlying acute HS in gut health. Fifty 28-week-old breeder hens were divided into two groups (25 hens each) raised under thermoneutral zone (23 °C) and acute HS (36 °C, 6 hours) conditions. The heart rate and cloacal temperature were measured in all hens, and jejunal mucosa tissues were randomly collected from 12 hens per treatment for RNA-sequencing analysis. The results indicated a significant increase in the heart rate and cloacal temperature in hens exposed to acute HS (P < 0.05). Transcriptome analysis identified 138 differentially expressed genes (DEGs) in heat-stressed breeder hens, including 75 upregulated DEGs containing heat shock protein (HSP), energy homeostasis metabolism-related gene (PDK4), and fat metabolism-related genes (PPARA and CD36), and 63 downregulated DEGs containing the bile acid transporter gene (SLC10A2). Gene ontology analysis revealed significant enrichment in biological processes related to heat response and cholesterol biosynthesis. Kyoto Encyclopedia of Genes and Genomes analysis highlighted several significant pathways, including steroid biosynthesis, steroid hormone biosynthesis, protein processing in endoplasmic reticulum, the peroxisome proliferator-activated receptor (PPAR) signaling pathway, and the adipocytokine signaling pathway. Protein-protein interaction network analysis involving two large networks: one containing several upregulated HSPs and genes related to energy homeostasis and fat metabolism (PDK4, PPARA, and CD36) and glucose transporter (SLC2A5), and the other containing downregulated DEGs related to cholesterol biosynthesis. Overall, acute HS might affect energy metabolism, fat metabolism, and glucose transport in the jejunal mucosa of breeder hens. Heat-stressed hens could restore the nutritional function of the jejunal mucosa by increasing the expression of HSPs. These findings provide a theoretical framework for further investigation into the molecular regulatory mechanisms responsible for HS-induced changes in the gut health of poultry.

overall_design: This study aimed to identify candidate genes related to acute HS in breeder hens and provide insight into the molecular mechanisms underlying acute HS in gut health. Fifty 28-week-old breeder hens were divided into two groups (25 hens each) raised under thermoneutral zone (23 °C) and acute HS (36 °C, 6 hours) conditions.Breeder hens jejunal mucosa tissues were randomly collected from 12 hens per treatment for RNA-sequencing analysis.

first: Satoshi

last: Kubota

first: Phocharapon

last: Pasri

first: Sutisa

last: Khempaka


title: Identification and characterization of an antiviral cytokine regulated by NF-κB signaling pathway in Bombyx mori

city: Chongqing

first: Fei

last: Wang

address: #216 Tiansheng Road, Beibei District

institute: Southwest University

summary: Insect NF-κB-like factor, Relish, is activated by viral infection and induces the production of antiviral proteins. In this study, we performed a transcriptomic analysis of BmE cells expressing the active form of BmRelish (BmRelishact) and identified BmVago as the most strongly-induced secreted-protein. Expression of Bmvago was specifically triggered by Bombyx mori nucleopolyhedrovirus (BmNPV) infection and regulated by BmSTING-BmRelish pathway. Incubating the fresh culture of cells with supernatant medium of BmVago-expressing cells or recombinant BmVago protein (rBmVago) significantly increased antiviral resistance. On the contrary, reducing the expression of Bmvago by RNA interference (RNAi) in BmE cells as well as in silkworm larvae impaired antiviral response. Furthermore, we constructed transgenic silkworm line over-expressing BmVago (BmVagoOV) and found they had markedly lower viral load and higher survival rate after BmNPV infection compared with the wild-type control. Co-immunoprecipitation assay showed Bmintegrin β1 interacts with BmVago and it was involved in BmVago-mediated antiviral response. Finally, we found the expression level of signaling molecules in Jak-Stat pathway increased in rBmVago-treated cells and BmVagoOV silkworm larvae but decreased in RNAi-treated cells. In summary, our research uncovered an inducible antiviral response in silkworm mediated by cytokine BmVago, which is the downstream effector of BmSTING-BmRelish pathway and functions as an antiviral cytokine.

overall_design: To identify which antiviral protein were induced by BmRelishact, we transfected BmE cells with BmRelishact or EGFP-expressing plasmid and performed transcriptomic analysis. We then performed structural domain and signal peptide prediction using Smart and SignalP-5.0 programs to find which secreted protein(s) was transcriptionally regulated by BmRelishact.

first: Dongmei

last: Wei

first: Jinming

last: Liu

first: Jie

last: Hu

first: Beilei

last: Zhang

first: Yumeng

last: Pan

first: Qingyou

last: Xia


title: Exercise promotes skeletal muscle growth in adolescents via modulating Mettl3-mediated m6A methylation of MyoD in muscle satellite cells

city: Shanghai

first: Shujing

last: Feng

address: Qingyuan ring road

institute: Shanghai university of sport

summary: Exercise promoted skeletal muscle growth in adolescents through facilitating MyoD mRNA stability of MuSCs in a Mettl3-mediated m6A dependent manner. The methyl donor betaine could be a potential alternative to exercise for promoting adolescent skeletal muscle growth by directly augmenting the global levels of m6A methylation. These findings may provide a theoretical foundation for encouraging daily fitness exercise and ensuring healthy growth in adolescents.

overall_design: To investigate the underlying mechanisms governing the effects of exercise on adolescent skeletal muscle growth, we conducted RNA-seq comparing TA from exercised mice with those from control mice. The exercise model was established in accordance with a modified protocol as described [24, 25]. Briefly, mice underwent a familiarization phase on the treadmill for 5 days, and then the speed was gradually increased from 8m/min to 12m/min, for 30 minutes daily. Subsequently, the mice were subjected to four weeks of treadmill training at a speed of 12m/min for 45min/day, 5 days/week.

first: Hao

last: Zhou

first: Xingzuan

last: Lin

first: Siyuan


title: CAR T Cells Secreting NGF-Neutralizing scFv Enhance Efficacy in Clear Cell Renal Cell Carcinoma by Relieving Immunosuppression through Immunosympathectomy [bulk RNA-seq]

first: peiwei

last: yang


title: Gene expression profile at single cell level of HS and normal skin

city: Birmingham

first: Mohammad

last: Athar

state: AL

address: 1670 University Blvd

department: Dermatology

institute: University of Alabama at Birmingham

summary: We used single cell RNA sequencing (scRNA-seq) to analyze infiltration and diversity of various immune and other types of cell populations.

overall_design: The Institutional Review Board of the University of Alabama at Birmingham approved the protocol (IRB-300005214) for obtaining surgically discarded skin tissues from heathy and HS (Hurley II/III stage) subjects. Surgical excisions from 6 patients with HS (axillary and groin lesions) and 2 healthy controls (discarded tissue from breast or abdominoplasty reduction surgery) were collected.

first: Mahendra

middle: P

last: Kashyap

first: Bharat

last: Mishra

first: M Shahid

last: Mukhtar

first: Chander

last: Raman