title: The sMAF transcription factor MAFG is a potent driver of melanoma

city: Tampa

first: jiqiang

last: yao

address: 12902 USF Magnolia Drive

institute: moffitt cancer center

summary: While common mutations in potent proto-oncogenes and tumor suppressors induce melanoma formation, no recurrent mutations associated with the late stages of melanomagenesis have been identified. Instead, non-mutational mechanisms such as the deregulation of transcriptiona or epigenetic programs typically promote the malignant progression and metastasis of melanoma. We have previously identified the small MAFG transcription factor MAFG as a critical target of the melanoma suppressor miRNA miRN-29, prompting an in-depth evaluation of the role of MAFG in melanoma. MAFG expression is elevated in melanoma compared to melanocytes and increases with tumor stage. Ectopic expression of MAFG in human melanocytes and melanoma cells enhances proliferation in virto and promotes melanoma growth in vivo. Moreover, MAFG overexpression in BRAFV600E; PTEN+/- mice accelerated melanomagenesis, significantly shortening overall survival. Despite being a critical NRF2 dimerization partner, NRF2 was dispensable for the effects of MAFG. Moreover, MAFG overexpression in melanoma had no effect on the activity of NRF2 transcriptional reporters, nor did it induce transcriptional programs associated with NRF2. RNA sequencing and pathway analysis revealed MAFG-mediated effects on melanoma-associated processes such as pigmentation. Indeed, MAFG downregulated MITF and its target genes in the pigmentation pathway in melanoma cells. We next analyzed the connection between MAFG and MITF and found that MAFG binds to MITF to impede its binding to melanocytic differentiation genes, thus promoting a less differentiated and more aggressive phenotype. Our study establishes MAFG as a potent driver of melanoma development through the relocation of MITF.

overall_design: Chromatin Immune Capture and sequencing (Cut&Run) performed on WM164 cells either overexpressing GFP or MAFG, using the following primary antibodies: anti-histone H3K4me3, anti-MAFG and anti-MITF. Secondary antibody anti-rabbit IgG was used as a negative control.

first: Michael

last: Martinez

first: Xiaoqing

last: Yu

first: Florian

last: Karreth


title: Long non-coding RNA LINC00680 functions as a ceRNA to promote esophageal squamous cell carcinoma progression through the miR-423-5p/PAK6 axis

status: Public on Oct 10 2024

city: Xiamen

first: Jiancheng

last: Ding

state: Fujian

address: School of Pharmaceutical Sciences, Xiamen University Xiang`an South Road

country: China

institute: Xiamen University

type: Expression profiling by high throughput sequencing

summary: Background: Esophageal squamous cell carcinoma (ESCC) is a common invasive malignancy worldwide with poor clinical outcomes. Increasing amount of long non-coding RNAs (lncRNAs) have been reported to be involved in cancer development. However, lncRNAs that are functional in ESCC and the underlying molecular mechanisms remain largely unknown.   Methods: Transcriptomic analysis was performed to identify dysregulated lncRNAs in ESCC tissue samples. The high expression of LINC00680 in ESCC was validated by RT-qPCR, and the oncogenic functions of LINC00680 was investigated by cell proliferation, colony formation, migration and invasion assays in ESCC cells in vitro and xenografts derived from ESCC cells in mice. RNA-seq, competitive endogenous RNA (ceRNA) network analysis, and luciferase reporter assays were carried out to identify LINC00680 target genes and the microRNAs (miRNAs) bound to LINC00680. Antisense oligonucleotides (ASOs) were used for in vivo treatment.  Results: Transcriptome profiling revealed that a large number of lncRNAs was dysregulated in ESCC tissues. Notably, LINC00680 was highly expressed, and upregulation of LINC00680 was associated with large tumor size, advanced tumor stage, and poor prognosis. Functionally, knockdown of LINC00680 restrained ESCC cell proliferation, colony formation, migration, and invasion in vitro and inhibited tumor growth in vivo. Mechanistically, LINC00680 was found to act as a ceRNA by sponging miR-423-5p to regulate PAK6 (p21-activated kinase 6) expression in ESCC cells. The cell viability and motility inhibition induced by LINC00680 knockdown was significantly reversed upon PAK6 restoration and miR-423-5p inhibition. Furthermore, ASO targeting LINC00680 substantially suppressed ESCC both in vitro and in vivo.  Conclusions: An oncogenic lncRNA, LINC00680, was identified in ESCC, which functions as a ceRNA by sponging miR-423-5p to promote PAK6 expression and ESCC. LINC00680/miR-423-5p/PAK6 axis may serve as promising diagnostic and prognostic biomarkers and therapeutic targets for ESCC.

first: Songtao

last: Xue

first: Bin

last: Zheng

first: Shi-qiang

last: Cao

first: Guo-sheng

last: Hu

first: Wen

last: Liu

first: Chun

last: Chen


title: Bile acid synthesis impedes tumor-specific T cell responses during liver cancer [RNA-seq1]

status: Public on Oct 09 2024

city: La Jolla

first: Kathryn

last: Lande

state: CA

address: 10010 N Torrey Pines Rd

department: IGC

country: USA

institute: Salk Institute

summary: The tumor's metabolic landscape significantly influences anti-tumor immunity, but it remains unclear how metabolic pathways commonly active in the organ of tumor-origin influence immunosurveillance. This study focused on the liver and identified accumulation of primary conjugated and secondary bile acids (BAs) as metabolic features associated with hepatocellular carcinoma and liver cancer models. Inhibiting conjugated BA synthesis through BAAT-deletion in hepatocytes enhanced tumor-specific T cell responses, reduced growth, and sensitized tumors to immune checkpoint blockade. BAs regulated CD8+ T cells differently; primary BAs like TCDCA induced oxidative stress and secondary BAs like LCA impaired T cell function through ER stress, which was countered by UDCA. Indeed, dietary UDCA provision suppressed tumor progression. These findings demonstrate how manipulating organ-specific metabolites affects antitumor immunity and modifying BA synthesis or dietary intake could enhance immunotherapy in liver cancer.

overall_design:
To investigate the effect of bile acid treatment on T Cell gene expression.
Gene expression profiling analysis using data obtained from RNA-seq of T Cells treated with different bile acids.in vitro activated CD8+ T cells  were exposed with different bile acids for 24 hours following with RNA was isolated using Qiagen RNA isoaltion kit.

first: Siva Karthik

last: Varanasi

first: Susan

last: Kaech


title: tRNA regulation and amino acid usage bias reflect a coordinated metabolic adaptation in Plasmodium falciparum [RNA-Seq III]

city: Stockholm

first: Sherwin

last: Chan

address: Solnavagen 9

country: Sweden

institute: Karolinska Institutet

summary: An adaptive feature of malaria-causing parasites is the digestion of host hemoglobin (HB) to acquire amino acids (AAs). Here we describe a link between nutrient availability and translation dependent regulation of gene expression as an adaptive strategy. We show that tRNA expression in Plasmodium falciparum does not match the decoding need expected for optimal translation. A subset of tRNAs decoding AAs that are insufficiently provided by HB are lowly expressed, wherein the abundance of a protein-coding transcript is negatively correlated with the decoding requirement of these tRNAs. Proliferation-related genes have evolved a high requirement of these tRNAs, thereby proliferation can be modulated by repressing protein synthesis of these genes during nutrient stress. We conclude that the parasite modulates translation elongation by maintaining a discordant tRNA profile to exploit variations in AA-composition among genes as an adaptation strategy. This study exemplifies metabolic adaptation as an important driving force for protein evolution.

overall_design: Plasmodium falciparum blood stage cultures were subjected to amino acid starvation and the change in mRNA profiles were monitored in a time course experiment upon blockage of transcription

first: Qian

last: Li

first: Leonie

last: Vetter

first: Ola

last: Larsson


title: Immunoresponsive Gene 1 Facilitates TLR4-agonist-Induced Augmentation of Innate Antimicrobial Immunity

city: Nashville

first: Margaret

middle: Ann

last: Mcbride

state: Tennessee

address: 1211 Medical Center Drive

department: Anesthesiology

institute: Vanderbilt University Medical Center

summary: Treatment with the toll-like receptor (TLR) 4 agonist monophosphoryl lipid A (MPLA) conditions innate immunocytes to respond robustly to subsequent infection, a phenotype termed innate immune memory. Our published studies show that metabolic reprogramming of macrophages is a prominent feature of the memory phenotype. We undertook studies to define the functional contributions of tricarboxylic acid (TCA) reprogramming to innate immune memory.  We observed that MPLA potently induced expression of immunoresponsive gene 1 (Irg1) and accumulation of its byproduct, the TCA cycle metabolite itaconate, in macrophages. Treatment with MPLA enhanced microbial clearance in wild type (WT), but not Irg1-deficient, mice, demonstrating that itaconate contributes to the antimicrobial phenotype in vivo. Treatment with MPLA equally augmented innate leukocyte recruitment, phagocytosis, respiratory burst and cytokine responses in WT and Irg1-deficient innate leukocytes after infection. However, the ability of Irg1-deficient macrophages to kill Pseudomonas aeruginosa was impaired. We further observed that itaconate is directly antimicrobial against P. aeruginosa at pH 5, which is characteristic of the phagolysosome, and is facilitated by reactive oxygen species. RNA sequencing revealed suppressed transcription of genes associated with phagolysosome function in Irg1 deficient macrophages. This study identifies a contribution of itaconate to MPLA-induced augmentation of innate antimicrobial immunity via facilitation of microbial killing.

overall_design: Bone marrow-derived macrophages from WT or Irg1KO (Jackson labs) C57BL/6 mice were treated with monophosphoyl lipid A (MPLA), lipopolysaccaride (LPS), vehicle (DMSO), or multiple treatments in sequence. We then performed gene expression profile analysis from 1-4 biological replicates per treatment at multiple timepoints. The effects on the transciptome were compared between groups. This data comes from 2 batches of RNA sequencing, and analysis of each batch was perfomed relative to its own vehicle controls. Samples 1-6, 15-20 and  are a batch, as are samples 7-14, 21-26.

middle: A

last: McBride

first: Edward

middle: R

last: Sherwood


title: ITPK1 sensitizes tumor cells to IgA-dependent neutrophil killing in vivo [RNA-seq]

city: Amsterdam

first: Connor

middle: Antonio

last: Richterich

address: Plesmanlaan 121

country: Netherlands

institute: Netherlands Cancer Institute

summary: Neutrophils can efficiently trigger cytotoxicity towards tumor cells and other target cells upon engagement of the IgA receptor CD89. However, the cell-intrinsic factors that influence the induction of cell death upon exposure to neutrophil effector mechanisms in vivo remain largely unknown. To uncover genetic regulators that influence target cell sensitivity to IgA-induced neutrophil-mediated killing, we used a human CD89 (hCD89) transgenic mouse model in which IgA-mediated killing of Her2-positive CD47-deficient murine target cells is mediated by neutrophils. Using a genome-wide in vivo screening approach, we demonstrate that deletion of the gene encoding inositol-tetrakisphosphate 1 kinase, ITPK1, increases survival of target cells in anti-Her2 IgA-treated mice. Moreover, we show that this effect depends on neutrophil activity and on the ITPK1 kinase domain. Notably, ITPK1 deficiency did not measurably impact survival of IgA-opsonized target cells in in vitro systems, underscoring the importance of in vivo screening systems to uncover physiologically relevant regulators of neutrophil killing.

overall_design: Three separate cultures of Ba/F3 cells were harvested, and washed once in PBS. RNA was subsequently extracted using the RNeasy Mini Kit (Qiagen #74004), according to the manufacturer’s instructions. Samples were processed using the TruSeq Stranded mRNA Kit (Illumina #20020594) for whole transcriptome sequencing. Samples were multiplexed and sequenced (50bp, single-end) on an Illumina NovaSeq6000 system (S1 flow cell). Reads were aligned to the reference genome (GRCm38, pre-built HISAT2 index, genome_snp_tran), and transcript counts were obtained using an in-house generated pipeline (GenSum, https://github.com/NKI-GCF/gensum).

first: Meike

last: Logtenberg

first: Marco

last: Jansen


title: Sensor Systems of KEAP1 Uniquely Detecting Oxidative and Electrophilic Stresses Separately In Vivo

city: Sendai

first: Takafumi

last: Suzuki

state: Miyagi

address: 2-1 Seiryo-machi, Tohoku University

country: Japan

institute: Tohoku University

summary: In the KEAP1-NRF2 stress response system, KEAP1 acts as a sensor for oxidative and electrophilic stresses through formation of S-S bond and C-S bond, respectively. Of the many questions left related to the sensor activity, following three appear important; whether these KEAP1 sensor systems are operating in vivo, whether oxidative and electrophilic stresses are sensed by the similar or distinct systems, and how KEAP1 equips highly sensitive mechanisms detecting oxidative and electrophilic stresses in vivo. To address these questions, we conducted a series of analyses utilizing KEAP1-cysteine substitution mutant mice, conditional selenocysteine-tRNA (Trsp) knockout mice, and human cohort whole genome sequence (WGS) data. The Trsp-knockout provokes severe deficiency of selenoproteins and compensatory activation of NRF2. However, mice lacking homozygously a pair of critical oxidative stress sensor cysteine residues of KEAP1 fail to activate NRF2 in the Trsp-knockout livers. Secondly, this study provides evidence for the differential utilization of KEAP1 sensors for oxidative and electrophilic stresses in vivo. Thirdly, theoretical calculations show that the KEAP1 dimer equips quite sensitive sensor machinery in which modification of a single molecule of KEAP1 within the dimer is sufficient to affect the activity. WGS examinations of rare variants identified seven non-synonymous variants in the oxidative stress sensors in human KEAP1, while no variant was found in electrophilic sensor cysteine residues, supporting the fail-safe nature of the KEAP1 oxidative stress sensor activity. These results provide valuable information for our understanding how mammals respond to oxidative and electrophilic stresses efficiently.

overall_design: To examine the contribution of KEAP1-Cys226/613 to the NRF2 activation in selenoprotein deficient livers, we generated compound mutant mice by crossing conditional liver-specific Trsp knockout (Trspfl/fl::AlbCre) mice and KEAP1C226S&C613S mutant mice. We performed RNA-sequence analysis using RNA samples from the livers. We utilized RNA samples from livers of Trspfl/fl, Trspfl/fl::AlbCre, KEAP1C226S&C613S/C226S&C613S::Trspfl/fl and KEAP1C226S&C613S/C226S&C613S::Trspfl/fl::AlbCre mice, in age of 20 day-old.

first: Miu

last: Sato

first: Nahoko

last: Yaguchi

first: Takuya

last: Iijima

first: Aki

last: Muramatsu

first: Liam

last: Baird

first: Masayuki

last: Yamamoto


title: Modulation of NOX2 causes Obesity- Mediated Atrial Fibrillation

city: Chicago

first: Jordan

last: Jousma

state: Illinois

address: 909 South Wolcott

department: Pharmacology and Regenerative Medicine

institute: University of Illinois

summary: Obesity is linked to an increased risk of atrial fibrillation (AF) via increased oxidative stress. While NADPH oxidase II (NOX2), a major source of oxidative stress and reactive oxygen species (ROS) in the heart predisposes to AF, the underlying mechanisms remain unclear. Here, we studied NOX2-mediated ROS production in obesity-mediated AF using Nox2-knock-out (KO) mice and mature human induced pluripotent stem cell-derived atrial cardiomyocytes (hiPSC-aCMs). Diet-induced obesity (DIO) mice and hiPSC-aCMs treated with palmitic acid (PA) were infused with a NOX blocker (apocynin) and a NOX2-specific inhibitor, respectively. We showed that NOX2 inhibition normalized atrial action potential duration and abrogated obesity-mediated ion channel remodeling with reduced AF burden. Unbiased transcriptomics analysis revealed that NOX2 mediates atrial remodeling in obesity-mediated AF in DIO mice, PA-treated hiPSC-aCMs, and human atrial tissue from obese individuals by upregulation of paired-like homeodomain transcription factor 2 (PITX2). Furthermore, hiPSC-aCMs treated with hydrogen peroxide, a NOX2 surrogate, displayed increased PITX2 expression, establishing a mechanistic link between increased NOX2-mediated ROS production and modulation of PITX2. Our findings offer insights into possible mechanisms through which obesity triggers AF and support NOX2 inhibition as a potential novel prophylactic or adjunctive therapy for patients with obesity-mediated AF.

overall_design: We then performed an unbiased global transcriptomic analysis by RNA sequencing (RNA-Seq) separately on lean vs obese HAT, control hiPSC-aCMs versus PA-hiPSC-aCMs, and PA-hiPSC-aCMs versus PA-GSK-hiPSC-aCMs.


title: Negative feedback of cyclic di-GMP levels optimizes switching between sessile and motile lifestyles in Vibrio cholerae [ChIP-seq]

city: Ann Arbor

first: Lydia

last: Freddolino

state: MI

address: 1136 Catherine St

department: Biological Chemistry

institute: University of Michigan

type: Genome binding/occupancy profiling by high throughput sequencing

summary: The signaling molecule cyclic di-GMP (cdG) controls the switch between bacterial motility and biofilm production, and fluctuations in cellular levels of cdG have been implicated in Vibrio cholerae pathogenesis. Intracellular concentrations of cdG are controlled by the interplay of diguanylate cyclase (DGC) enzymes, which synthesize cdG to promote biofilms, and phosphodiesterase (PDE) enzymes, which hydrolyse cdG to drive motility. To track the complete regulatory logic of how V. cholerae responds to changing cdG levels, we followed a time course of overexpression of either the V. harveyi diguanylate cyclase QrgB or a variant of QrgB lacking catalytic activity (QrgB*). We find that QrgB increases cdG levels relative to QrgB* for 30 minutes after overexpression, but the effect of QrgB on cdG levels plateaus at 30 minutes, indicating tight adaptive control of cdG levels. In contrast, loss of VpsR, a master regulator activating biofilm formation upon binding to cdG, leads to higher baseline levels of cdG and continuously increasing cdG through 60 minutes after QrgB induction, revealing the existence of a negative feedback loop on cdG levels operating through VpsR. Through a combination of RNA polymerase ChIP-seq, RNA-seq, and genetic approaches, we show that the PDE CdgC is activated by VpsR at high cdG concentrations, mediating this negative feedback on cdG levels. We further identify a transcript encoded within, and antisense to, the cdgC open reading frame which we name sRNA negative regulator of CdgC (SnrC). RNA polymerase ChIP-seq and RNA-seq demonstrate SnrC to be expressed specifically under conditions of high cdG in the absence of VpsR. Ectopic SnrC expression increases cdG levels in a manner depending on CdgC, demonstrating that its effect on cdG levels is likely through interference with CdgC production. Further, although cells lacking cdgC exhibit enhanced biofilm formation, these mutants are outcompeted by wild type V. cholerae in colonization assays that reward a combination of attachment, dispersal, and motility. These results underscore the importance of negative feedback regulation of cdG to maintain appropriate homeostatic levels for efficient transitioning between biofilm formation and motility, both of which are necessary over the course of the V. cholerae infection cycle.

overall_design: Chromatin immunoprecipitation (ChIP) of RNA polymerase in Vibrio cholerae in the following genotypes: WT, delta-vpsR, delta-vpsT

first: Aathmaja

last: Rangarajan

first: Jeremy

middle: W

last: Schroeder

first: Rebecca

middle: L

last: Hurto

first: Geoffrey

middle: B

last: Severin

first: Meng-Lun

last: Hsieh

first: Christopher

middle: M

last: Waters


title: GETomics based bulk-RNA sequencing revealed HIF-3α prevents COPD by inhibiting alveolar epithelial cell ferroptosis via the HIF-3α-GPx4 axis.

city: Beijing

first: Junchao

last: Jiang

address: Dongdan Santiao, Doncheng District

institute: Peking Union Medical College

summary: COPD patients are largely asymptomatic until later stages when prognosis is generally poor. Here, by shifting the focus forward to pre-COPD and smoking stages, we found the peak of reactive oxygen species (ROS) level and hypoxia character are enriched in pre-COPD samples, and hypoxia inducible factor (HIF)-3α is the key factor. Smoking induced regional tissue hypoxia and emphysema have been found in COPD patients. However, the mechanisms underlying hypoxia especially HIF-3α and COPD have not been investigated. In this study, by leveraging scRNA-seq and EpCAM co-localization analysis we identified HIF-3α is downregulated in alveolar epithelial cells in COPD. In vitro experiments using lentivirus transfection, bulk-RNA seq and RSL3 we found the activation of HIF-3α-GPx4 axis inhibits alveolar epithelial cell ferroptosis under the treatment of cigarettes smoking extracts (CSE). Further results from SftpcCreert2/+R26LSL-Hif3a knock-in mice demonstrated overexpression of HIF-3α inhibits alveolar epithelial cells ferroptosis and prevents the declining of lung function. Our data suggest the activation of HIF-3α-GPx4 axis prevents COPD by inhibiting alveolar epithelial cells ferroptosis. By leveraging this comprehensive analysis method, more promising treatment targets will be found and enhance our understanding to the pathogenesis.

overall_design: To explore the decades long COPD pathogenesis process by including non-smokers, smokers, pre-COPD and COPD patients.Using transcriptomics technology to delineate the changing chracter.

first: Ting

last: Yang

first: Chen

last: Wang


title: Cell-Type-Specific Epigenetic Priming of Gene Expression in Nucleus Accumbens by Cocaine

city: New York

first: Aarthi

last: Ramakrishnan

state: New York

address: 1425 Madison Avenue

department: Neuroscience

institute: Icahn School of Medicine at Mount Sinai

summary: A hallmark of addiction is the ability of drugs of abuse to trigger relapse after periods of prolonged abstinence. Here, we describe a novel epigenetic mechanism whereby chronic cocaine exposure causes lasting chromatin and downstream transcriptional modifications in the nucleus accumbens (NAc), a critical brain region controlling motivation. We link prolonged withdrawal from cocaine to the depletion of the histone variant H2A.Z, coupled with increased genome accessibility and latent priming of gene transcription, in D1 dopamine receptor-expressing medium spiny neurons (D1 MSNs) that relate to aberrant gene expression upon drug relapse. The histone chaperone ANP32E removes H2A.Z from chromatin, and we demonstrate that D1 MSN-selective Anp32e knockdown prevents cocaine-induced H2A.Z depletion and blocks cocaine’s rewarding actions. By contrast, very different effects of cocaine exposure, withdrawal, and relapse were found for D2 MSNs. These findings establish histone variant exchange as an important mechanism and clinical target engaged by drugs of abuse to corrupt brain function and behavior.

overall_design: we performed ChIP-seq for H2A.Z and, simultaneously, for total histone H3 in 4 conditions - Saline Saline, Saline Cocaine, Cocaine Saline and Cocaine-Cocaine.

first: Philipp

last: Mews

first: Yentl

last: Van der Zee

first: Ashik

last: Gurung

first: Eric

middle: J

last: Nestler


title: Cell-Type-Specific Epigenetic Priming of Gene Expression in Nucleus Accumbens by Cocaine [ATAC-seq]

overall_design: We examined ‘open’ chromatin regions in both D1 and D2 MSN nuclei genome-wide by use of assay for transposase-accessible chromatin with sequencing (ATAC-seq) At baseline (SS), acute cocaine (SC), cocaine relapse (CC) and following chronic cocaine exposure and prolonged withdrawal (CS).


title: Low Dose Pro-resolving Mediators Reset the Resolution Response to Microbial Inflammation

city: Boston

last: Chiang

state: MA

address: 60 Fenwood Rd. 3016Q

institute: Brigham and Women's Hospital

summary: With murine zymosan peritonitis, repetitive dosing of a SPM panel (RvD1, RvD2, RvD5, MaR1 and RvE2 at 1 ng/mouse, i.p.) was given to mice, followed by zymosan challenge to initiate peritonitis. Leukocyte composition and resolution indices were calculated. RNA-sequencing was carried out

overall_design: Mice were given "treatment" i.e., a SPM panel (RvD1, RvD2, RvD5, MaR1 and RvE2, 0.1 ng each/mouse, i.p.) or "vehicle" controls on days 0, 2, 5, 7 and 9. On day 12, zymosan A (1 mg/mouse, i.p.) was given together with either the SPM panel or vehicle. Peritoneal exudates were collected at 12, 24 and 48h, and leukocyte subtypes were assessed using flow cytometry. Aliquots of exudate leukocytes collected from 12h time point were used for RNA-seq.

first: Charles

middle: N

last: Serhan


title: Host transcriptome in human 239T cells infected with HPIV3 expressing wild-type virus versus its matrix-deleted counterpart

first: Chuan-Tien

last: Hung

address: 1468 Madison Avenue

department: Microbiology

summary: Paramyxoviruses (PMVs) exploit the host's translation machinery to enhance their replication. We found that the PMV matrix protein is crucial in this process, inhibiting host protein synthesis while boosting viral protein production. This occurs through interactions with the core exon-junction complex (cEJC), a key player in mRNA biogenesis. Disruption of this interaction using siRNA led to increased viral replication but did not affect other viruses like SARS-CoV-2. Our study unveils a novel mechanism by which PMVs hijack host cell resources, offering new targets for antiviral therapy.

sample_organism:
Homo sapiens
Human respirovirus 3

platform_organism:
Homo sapiens
Human respirovirus 3

overall_design: HEK 293T cells were mock-infected or infected with either HPIV3-WT or HPIV3 delta M at an MOI of 3 for 48 hours. Total RNA was extracted from the cells and subjected to polysome profiling followed by RNA sequencing analysis using poly(A) enrichment.

first: Benhur

last: Lee


title: ITPK1 sensitizes tumor cells to IgA-dependent neutrophil killing in vivo

first: Connor

last: Richterich

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

overall_design: Refer to individual Series


title: ITPK1 sensitizes tumor cells to IgA-dependent neutrophil killing in vivo [CRISPR focused]

overall_design: For the focused screen, sensBa/F3 cells were lentivirally transduced with the focused sgRNA library in the lenti-Guide-Puro vector (Addgene #52963) at a transduction rate of approximately 50% and a coverage of >2000 fold. The focused library targets 35 genes with 2 sgRNAs each, and with target genes selected based on observed enrichment scores in a primary screen (and on RNA-expression levels. In addition, high scoring genes encoding certain transcription factors (e.g. Rhox11 and Gtf2e2) were excluded, and a number of presumed interactors of top-scoring genes (i.e. Kctd10, Cul3, and Wipf2 with Itsn2) were included. Following transduction, the cells were cultured for >7 days in the presence of 5ug/ml puromycin. Before injection into mice, a library reference sample was taken. Next, hCD89-transgenic mice and littermate controls were injected intraperitoneally (i.p.) with 2x107 cells in 200uL PBS. Directly after target cell injection, mice received PBS (control treatment) or 100ug anti-hHer2 IgA1 antibody in PBS by intraperitoneal injection. 16h after injection, mice were euthanized and the peritoneal cavity was washed with PBS containing 5 mM EDTA to obtain target cells and murine immune cell infiltrates. To estimate target cell killing efficiency by flow cytometry, a small aliquot was taken from each sample and pooled per treatment group (PBS or anti-hHer2 IgA1). Next, cells were stained with APC-conjugated anti-hHer2 and anti-hCD19 antibodies and target cells were positively selected for by MACS using anti-APC magnetic beads (MiltenyiBiotec #130-090-855), according to the manufacturer’s protocol. Thereafter, DNA was isolated from enriched target cells using the DNeasy Blood & Tissue Kit (QIAGEN #69504). In the focused screen, 2 of the DNA samples derived from the treated group were pooled. SgRNA sequences in the pooled or unpooled sample groups were amplified using NEBNext High-Fidelity 2X PCR Master Mix (New England Biolabs #M0541S), following the manufacturer’s instructions and using primers containing sample barcodes. After PCR, prepared libraries were pooled at equimolar amounts and sgRNA distributions were analyzed by deep sequencing using the Illumina HiSeq2500 platform. In brief, sequence reads were aligned to the Brie mouse library, reads containing mismatches were removed, sgRNA distributions were quantified, and genes were scored using MAGeCK RRA (version 0.5.6) (27).

first: Meike

last: Logtenberg

first: Marco

last: Jansen


title: A correctable immune niche for epithelial stem-cell reprogramming and post-viral lung diseases

city: St. Louis

first: Michael

last: Holtzman

state: MO

address: 4939 Childrens Pl

department: Pulmonary & Critical Care

institute: Washington University in St. Louis School of Medicine

summary: Epithelial barriers are programmed for defense and repair but are also the site of long-term structural remodeling and disease. In general, this paradigm features epithelial stem cell (ESCs) that are called on to regenerate damaged tissues but can also be reprogrammed for detrimental remodeling. Here we identify a Wfdc21-dependent monocyte-derived dendritic cell (moDC) population that functions as an early sentinel niche for basal-ESC reprogramming in mouse models of epithelial injury after respiratory viral infection. Niche function depends on moDC delivery of ligand Gpnmb to basal-ESC receptor CD44 so that properly timed antibody blockade of ligand or receptor provides long-lasting correction of reprogramming and broad disease phenotypes. These same control points work directly in mouse and human basal-ESC organoids, and corresponding biomarkers track with comparable lung diseases in humans. Together, the findings identify a mechanism to explain and modify what is otherwise a stereotyped but sometimes detrimental response to epithelial injury.

overall_design: Lung tissue RNA was purified from four mice per sample condition (moDCs or monocytes for 12 d after SeV infection or PBS control challenge) and subjected to RNAseq.

first: Kangyun

last: Wu

first: Yong

last: Zhang

first: Huiqing

last: Yin-DeClue

first: Kelly

last: Sun

first: Dailing

last: Mao

first: Kuangying

first: Stephen

last: Austin

first: Erika

middle: C

last: Crouch

first: Steven

last: Brody

first: Derek

middle: E

last: Byers

first: Christy

last: Hoffmann

last: Hughes


title: The effect and mechanism of freeze-dried powder of Poecilobdella manillensis on improving inflammatory injury of rat glomerular mesangial cells through TXNIP / NLRP3 pathway.

first: Xi

address: Gongchen Street, Fangshan District

institute: Beijing University of Chinese Medicine

summary: In this study, transcriptome technology was used to explore the mechanism of Poecilobdella manillensis lyophilized powder to improve the inflammatory injury of rat mesangial cells.In general, this study found that TXNIP / NLRP3 signaling pathway is one of the effective mechanisms of Poecilobdella manillensis lyophilized powder to improve the inflammatory injury of diabetic nephropathy.

overall_design: The cell samples were HBZY-1 rat mesangial cells, which were divided into three groups : control group, LPS-induced model group and high-dose water extract of Poecilobdella manillensis lyophilized powder group for transcriptome sequencing, with three biological replicates in each group.


title: ITPK1 sensitizes tumor cells to IgA-dependent neutrophil killing in vivo [CRISPR genome-wide]

overall_design: For the genome-wide screen, sensBa/F3 cells were lentivirally transduced with the pooled Brie CRISPR KO library (Addgene #73632/3) in the lenti-Guide-Puro (Addgene #52963) vector at a transduction rate of approximately 50% and a coverage of >1800 fold. Following transduction, the cells were cultured for >7 days in the presence of 5ug/ml puromycin. Before injection into mice, a library reference sample was taken. Next, hCD89-transgenic mice and littermate controls were injected intraperitoneally (i.p.) with 2x10^7 cells in 200uL PBS. Directly after target cell injection, mice received PBS (control treatment) or 100ug anti-hHer2 IgA1 antibody in PBS by intraperitoneal injection. 16h after injection, mice were euthanized and the peritoneal cavity was washed with PBS containing 5 mM EDTA to obtain target cells and murine immune cell infiltrates. Next, cells were stained with APC-conjugated anti-hHer2 and anti-hCD19 antibodies and target cells were positively selected for by MACS using anti-APC magnetic beads (MiltenyiBiotec #130-090-855), according to the manufacturer’s protocol. Thereafter, DNA was isolated from enriched target cells using the DNeasy Blood & Tissue Kit (QIAGEN #69504). In the genome-wide screen 3-4 samples within each group were pooled based on recovered DNA amounts, yielding 3 anti-hHer2-treated samples and 3 PBS-treated genomic DNA samples. SgRNA sequences in the pooled or unpooled sample groups were amplified using NEBNext High-Fidelity 2X PCR Master Mix (New England Biolabs #M0541S), following the manufacturer’s instructions and using primers containing sample barcodes. After PCR, prepared libraries were pooled at equimolar amounts and sgRNA distributions were analyzed by deep sequencing using the Illumina HiSeq2500 platform. In brief, sequence reads were aligned to the Brie mouse library, reads containing mismatches were removed, sgRNA distributions were quantified, and genes were scored using MAGeCK RRA (version 0.5.6)


title: Docosahexaenoic acid (DHA); a driving force regulating gene expression in bluefin tuna (Thunnus thynnus) larvae development.

city: Eilat

first: Eran

last: Yanowski

address: North beach

country: Israel

institute: National Center for Mariculture

summary: This study elucidated the role of DHA-modulated genes in the development and growth of bluefin tuna (Thunnus thynnus) larvae ingesting increasing levels of DHA in their rotifer prey. The effect of feeding low, medium, and high rotifer (Brachionus rotundiformis) DHA levels (2.0, 3.6 and 10.9 mg DHA g-1 DW, respectively) was tested on 2-15 days post hatching (dph) bluefin tuna larvae. Larval DHA content markedly (P < 0.05) increased in a DHA dose-dependent manner (1.5, 3.9, 6.1 mg DHA g-1 DW larva, respectively), that was positively correlated with larval prey consumption, and   growth (P < 0.05). Gene ontology enrichment analyses of DEGs demonstrated dietary DHA significantly (P < 0.05) affected different genes and biological processes at different developmental ages.  The number of DHA up-regulated DEGs was highest in 10 dph larvae (408), compared to 5 (11) and 15 dph fish (34), and were mainly involved in neural and synaptic development in the brain and spinal cord. In contrast, DHA in older 15 dph larvae elicited fewer DEGs but played critical roles over a wider range of developing organs. The emerging picture underscores the importance of DHA-modulated gene expression as a driving force in bluefin tuna larval development and growth.

overall_design: To evaluate the function of DHA on the development and growth of Blue-fin tuna larvae, we fed BFT larvae with enriched rotifers at 3 doses of DHA (2.0, 3.6 and 10.9 mg DHA g-1 DW) and evaluated larval ingestion rates, growth and whole-larvae transcriptome at 5, 10 and 15 days post hatch.

first: William

last: Koven

first: Luke

middle: D

last: Gardner

first: Oria

last: Nixon

first: Barbara

last: Block


title: Bulk RNA-Seq of cardiomyocyte Runx1 overexpression in neonatal mouse ventricles

city: Milwaukee

first: Michaela

last: Patterson

state: WI

address: 8701 Watertown Plank Road

institute: Medical College of Wisconsin

summary: The effect of Runx1 expression in neonatal mouse ventricles was assessed with overexpression of Runx1 in cardiomyocytes using inducible Myh6-Cre.

overall_design: Littermates with either Myh6-CreER alone (n=2) or Myh6-CreER and Rosa-Runx1 overexpression allele (n=4) were injected with tamoxifen at P0 and P1. Whole ventricles were then collected at P10.

first: M

first: K

last: Atkins

first: S

middle: K

last: Swift

last: Flinn

first: C

last: O'Meara


title: SPP1+ macrophages in HR+ breast cancer are associated with tumour-infiltrating lymphocytes.

city: Seoul

first: SUMIN

last: CHA

address: 88, Olympic-ro 43-gil, Songpa-gu

country: South Korea

institute: Asan Institute for Life Sciences, Asan Medical Center

summary: Breast cancer, categorised into hormone receptor-positive (HR+), HER2-positive (HER2+), and triple-negative (TNBC) subtypes, exhibits varied outcomes based on the number of tumour-infiltrating lymphocytes (TILs). Increased TIL levels are associated with better outcomes in HER2+ and TNBC, while HR+ individuals with high TIL levels show shorter survival but greater neoadjuvant chemotherapy response. To explore the divergent roles of TIL levels across various subtypes and their effect on immune cell composition, we employed single-cell RNA sequencing on 31 patients with breast cancer. HR+ breast cancer with high TIL levels (TIL-high) revealed increased SPP1+ macrophages, increased SPP1 expression in other monocytes/macrophages (mono/macro) subgroups, and enriched pathways associated with extracellular matrix (ECM) remodelling in mono/macro. Moreover, cell–cell interaction analyses revealed enhanced SPP1, MIF, and FN1 signalling in the interaction between SPP1+ macrophages and T-cells in TIL-high HR+ breast cancer. Spatial transcriptomics data highlighted the close proximity of SPP1+ macrophages, CD8+ T-cells, and CD4+ T-cells in TIL-high HR+ breast cancer. Our findings unveil the novel influence of SPP1+ macrophages on T-cells in TIL-high HR+ breast cancer, potentially explaining the poor prognosis and offering insights for targeted interventions.

overall_design: Single cell dissociation of breast cancer tissue and sorting CD45+ cells (n=21)


title: Serial single-cell RNA sequencing unveils drug resistance and metastatic traits in stage IV breast cancer

city: Koto-ku

first: Reo

last: Maruyama

state: Tokyo

department: Cancer Institute

institute: Japanese Foundation For Cancer Research

summary:
Background: Drug resistance and metastasis are complex processes that remain poorly understood on the molecular level with single-cell resolution. We profiled the transcriptomic changes at the single-cell level associated with cancer cell progression, chemotherapy resistance, and metastasis in samples from a single de novo Stage IV breast cancer patient.
Method: Pretreatment biopsy samples and posttreatment surgical specimens from the primary tumor and distant metastases were collected for single-cell RNA sequencing and subsequent cell clustering, copy number variation (CNV) estimation, transcription factor activity estimation, and pseudotime analyses.
Results: CNV estimation analysis identified a small pretreatment cancer cell population that resisted chemotherapy and subsequently expanded. In the posttreatment primary tumors, new clones emerged, which we refer to as potential Metastatic Precursor Cells (MPCs), exhibiting the CNV profiles similar to metastatic cells. MPCs appeared to originate from hypoxic regions in the necrotic core of the tumor and exhibited expression profiles indicative of epithelial–mesenchymal transition. Comparison of MPCs to metastatic cancer cells also revealed dynamic changes in transcription factor activities and calcitonin pathway gene expressions.
Conclusion: These findings demonstrate the utility of single-patient clinical sample analysis for clonal adaptations underlying tumor drug resistance, regrowth, and metastasis.

overall_design:
Pre- and post-treatment and metastatic specimens from a single stage IV breast cancer patient were subjected to scRNA-seq analysis.

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Submitter states that missing raw files are due to patient privacy concerns.
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first: Kazutaka

last: Otsuji


title: Transcriptomic Landscape of Leptospira Forming Biofilm Reveals Adaptation to Starvation and General Stress while Maintaining Virulence.

city: Paris

first: Monot

last: Marc

address: 25, rue du docteur roux

department: Genomes and Genetics

country: France

institute: Institut Pasteur

summary: The life-threatening pathogen Leptospira interrogans navigates a dual existence: surviving in environmental reservoirs and infecting mammalian hosts. Leptospira biofilm formation is thought to be an important survival strategy in environmental contexts and may also contribute to the persistence of leptospirosis in maintenance hosts. Examining the correlation between biofilm formation and the virulence of pathogenic strains might improve our comprehension of the epidemiology of leptospirosis. To further explore Leptospira’s survival strategy, our study focused on elucidating the biological state of pathogenic Leptospira within biofilms, particularly aiming to uncover the adaptations and regulatory mechanisms that are involved in such complex microenvironments. To determine the transcriptional profile of pathogenic Leptospira in biofilm, we compared the genome-wide transcriptomic profiles in late biofilms (21 days old) with those in exponential planktonic cultures (5 days old), revealing a pronounced transcriptomic shift. While genes linked to motility, energy production, and metabolism were downregulated, those governing the general stress response, defense against metal stress, and redox homeostasis showed a significant upsurge, hinting at a tailored defensive strategy against stress in late biofilms. A standout finding was the increased expression of the csoR, copZ, and copA locus, integral to copper ion stress response in other bacterial genera, suggesting a unique adaptation to metal-induced stress. Further, despite a reduced metabolic state in biofilms, their disruption swiftly restored metabolic activity. Crucially, bacteria either in late biofilms or resulting from biofilm disruption retained virulence in a hamster infection model, defying the notion that biofilm maturation abolishes pathogenicity. In summary, our study highlights Leptospira's adaptive equilibrium in biofilms: minimizing cellular energy expenditure to conserve resources, potentially aiding in withstanding stresses while maintaining its pathogenicity. These insights are important for explaining the survival strategies of Leptospira, revealing that a biofilm lifestyle may confer an advantage in maintaining virulence. This understanding is essential for managing leptospirosis across both environmental reservoirs and mammalian hosts.

first: Marc

last: Monot

first: Grégoire

last: Davignon


title: ITPK1 sensitizes tumor cells to IgA-dependent neutrophil killing in vivo [CRISPR Epistasis]

first: Connor

last: Richterich

overall_design: Epistasis_Screen: A set of 76 genes involved in cell death processes was assembled based on literature research, including gene-ontology terms related to apoptosis, necroptosis, ferroptosis, pyroptosis and TNF-mediated cell death. Furthermore, genes reported to be involved in cytokine-induced cell-death were included. An sgRNA library targeting these 76 genes (3 sgRNAs per gene) was ordered as an oligonucleotide pool (IDT DNA) and cloned into the lenti-Guide-Puro vector (Addgene #52963). SgRNA sequences were picked using CRISPick (Broad Institute GPP Webportal). In order to generate sensBa/F3 cells deficient of ITPK1, sensBa/F3 cells were electroporated (Lonza #V4XP-3024) (Program E0-117) with sgRNA targeting ITPK1 (in lenti-Guide-Puro, Addgene #52963) and selected for 48h in 10g/mL puromycin. Next single cell clones were screened for ITPK1 inactivation by DNA sequencing and western blot analysis, and for lentiviral integration. Six clones with ITPK1 deletions, but without lentiviral integration were pooled. Subsequently, both sensBa/F3 cells and sensBa/F3 ITPK1 KO cells were transduced with the sgRNA library targeting the 76 cell death-related genes. Next, the resulting sensBa/F3 and sensBa/F3 ITPK1 KO cell death libraries were cultured for 10 days to remove cells containing knockouts of essential genes, and sgRNA distributions were assessed using Illumina MiSeq sequencing. The expanded sensBa/F3 cell death libraries were subsequently stained with CellTrace (CT) violet or yellow (ThermoFisher #C34554), respectively and pooled at a 1:1 ratio. Thereafter, the stained cell pool was intraperitoneally injected into hCD89-transgenic mice (n=8, 20M cells each) that were subsequently treated with anti-hHer2 IgA or PBS (n=4 each). After 16h, mice were sacrificed and cells obtained by intraperitoneal lavage, and stained with Near-IR Dead Cell Staining Kit (ThermoFisher #L10119). Subsequently, viable tumor cells obtained from each mouse were sorted into CT violet- and yellow-positive populations. Next, genomic DNA of each sample was obtained using DirectPCR Lysis reagent (Viagen Biotech #301-C) according to the manufacturer’s instructions. SgRNA sequences were subsequently amplified using NebNext High-Fidelity 2X PCR Master Mix (New England Biolabs #M0541S), following the manufacturer’s instructions, using primers containing sample barcodes. After PCR, prepared libraries were pooled at equimolar amounts and sgRNA library distributions were analyzed by Illumina MiSeq sequencing. In brief, sequence reads were aligned to a reference library, reads containing mismatches removed, and sgRNA distributions quantified and scored using MAGeCK RRA (version 0.5.6) (27). Downstream analysis focused on comparison of the positive log2-fold changes of sgRNAs between treated and untreated mice in either ITPK1-proficient or -deficient backgrounds.

first: Marco

last: Jansen

first: Meike

last: Logtenberg


title: NF-κB- and TET2-mediated macrophage reprogramming overrides the anti-inflammatory effects of hypoxia and improves responses in human cancer (RNA-Seq II).

city: Badalona, Barcelona

first: Esteban

last: Ballestar

state: N/A = Not Applicable

address: Ctra de Can Ruti, Camí de les Escoles s/n

country: Spain

institute: Josep Carreras Research Institute (IJC)

summary: Macrophages orchestrate various aspects of immunity and tissue homeostasis in health and disease. In cancer, the presence of macrophages is largely associated with poor prognosis, due to their reprogramming into immunosuppressive cells in the tumor microenvironment (TME). In this study, we investigated the effects of hypoxia, a key feature of the TME, on the functional, epigenetic, and transcriptional status of macrophages. Surprisingly, we found that hypoxia boosts the immunogenicity of macrophages in vitro, in a process that is partially regulated by DNA methylation. Specifically, we find a cluster of pro-inflammatory genes that undergo DNA demethylation and transcriptional upregulation during macrophage activation in hypoxia. These genes are regulated by NF-κB, while HIF1α contributes to the transcriptional program through DNA methylation-independent mechanisms. In cancers such as bladder and ovarian carcinomas, the signatures of hypoxic inflammatory macrophages are found in immune-rich tumors, where they correlate with better patient prognoses. The NF-κB-associated DNA hypomethylation is recapitulated in an in vivo-equivalent subset of hypoxic inflammatory macrophages, isolated from ovarian tumors, validating the translation of the in vitro results. Functionally, co-culture assays and putative cell-cell communication analyses suggest that hypoxic-activated macrophages enhance T cell-mediated responses. Our results challenge existing paradigms regarding the effects of hypoxia on macrophages and highlight novel target cells to ameliorate cancer responses.

overall_design: RNA-seq analysis of distinct populations of macrophages (FOLR2, IL4I1, TREM2) sorted from human ovarian cancer tumors through flow cytometry.

first: Carlos

last: de la Calle-Fabregat

first: Josep

last: Calafell-Segura

first: Margaux

last: Gardet

first: Garett

last: Dunsmore

first: Kevin

last: Mulder

first: Laura

last: Ciudad

first: Aymeric

last: Silvin

first: Joaquim

last: Moreno-Càceres

first: Ángel

last: Corbí

first: Judith

last: Michels

first: Sébastien

last: Gouy

first: Cristina

last: Muñoz-Pinedo

first: Charles-Antoine

last: Dutertre

first: Javier

last: Rodríguez-Ubreva

first: Florent

last: Ginhoux


title: Effect of a FOXO1 Inhibitor on Trophoblast Differentiation from Human Pluripotent Stem Cells

city: Kobe City

first: Michiyo

last: Koyanagi-Aoi

address: Chuo-ku, Kusunoki-cho 7-5-1

institute: Kobe University

summary: We compared global gene expressions of naive iPSCs, TE, CT, ST and EVT treated with or without AS1842856 in TE differentiation

overall_design: Comparison of the gene expression profiles of iPSCs, iPS-derived differentiated cells and placenta

first: Erika

last: Tanaka

first: Takashi

last: Aoi


title: Collagen bioinks redefined: Optimizing ionic strength and growth factor delivery for cartilage tissue engineering

city: Plovdiv

first: Yordan

last: Sbirkov

address: 15 A Vassil Aprilov Blvd

department: Medical Biology

country: Bulgaria

institute: Medical University of Plovdiv

summary: This study investigates the impact of ionic strength on the gelation kinetics of collagen biomaterial inks and evaluates the efficiency of TGFβ-1 sequestration within these hydrogels, alongside their compatibility with bioprinting live chondrocyte and adipose-derived stem cell lines for cartilage tissue engineering. By adjusting sodium chloride and phosphate-buffered saline (PBS) concentrations, we demonstrate that reduced ionic strengths accelerate gelation, facilitating high-fidelity bioprinting while supporting high cell viability and post-printing proliferation of chondrocytes. Furthermore, at a 1% collagen concentration, the hydrogel effectively immobilized TGFβ-1, with less than 0.5% released over two weeks, indicating potent sequestration capabilities. Using adipose-derived mesenchymal stem cells, histomorphological and transcriptomic analyses reveal that the presence of TGFβ-1 significantly enhances chondrogenesis. These findings underscore the critical role of ionic strength in optimizing collagen ink properties for advanced bioprinting applications and highlight the potential of collagen hydrogels as effective carriers for sustained growth factor delivery, paving the way for successful cartilage tissue engineering strategies.

overall_design: Compare 3D bioprinted SVF cells in collagen vs collagen+TGFb bioinks in terms of chondrogenesis. We also compare these two bioprinting approaches to the gold standard of 3D spheroids (micromasses). All conditions are compared to SVF cells grown in standard 2D (monolayer) cultures.


title: NF-κB- and TET2-mediated macrophage reprogramming overrides the anti-inflammatory effects of hypoxia and improves responses in human cancer

first: Esteban

last: Ballestar


title: NF-κB- and TET2-mediated macrophage reprogramming overrides the anti-inflammatory effects of hypoxia and improves responses in human cancer (RNA-Seq I).

overall_design: RNA-seq analysis of human blood monocytes and monocyte-derived macrophages (unstimulated or activated with LPS) in vitro in normoxia (21% O2) or hypoxia (1% O2).

first: Carlos

last: de la Calle-Fabregat

first: Josep

last: Calafell-Segura

first: Margaux

last: Gardet

first: Garett

last: Dunsmore


title: NF-κB- and TET2-mediated macrophage reprogramming overrides the anti-inflammatory effects of hypoxia and improves responses in human cancer (DNA methylation I).

overall_design: DNA methylation analysis of human blood monocytes and monocyte-derived macrophages (unstimulated or activated with LPS) in vitro in normoxia (21% O2) or hypoxia (1% O2).

first: Kevin

last: Mulder

first: Laura

last: Ciudad

first: Aymeric

last: Silvin

first: Joaquim

last: Moreno-Càceres


title: The molecular mechanism and clinical significance of IGF2BP3 in adenocarcinoma of the esophagogastric junction [RNA-seq]

city: guangzhou

first: Yongbo

last: Pan

address: 106 Zhongshan 2 Lu, Guangzhou

department: Guangdong Provincial People's Hospital

institute: Guangdong Academy of Medical Sciences

summary: To explore the major function of IGF2BP3 in Adenocarcinoma of the esophagogastric junction, we conducted RNA sequencing (RNA-seq) in IGF2BP3 KD OE-19 cells.

overall_design: Adenocarcinoma of the esophagogastric junction OE-19 cells were KD IGF2BP3, then subjected to RNA-sequencing analysis.


title: NF-κB- and TET2-mediated macrophage reprogramming overrides the anti-inflammatory effects of hypoxia and improves responses in human cancer ChIP-Seq).

overall_design: ChIP-seq of HIF1a and p65 in monocyte-derived macrophages (unstimulated or activated with LPS for 2h) in vitro in normoxia (21% O2) or hypoxia (1% O2).

first: Ángel

last: Corbí

first: Judith

last: Michels

first: Sébastien

last: Gouy

first: Cristina

last: Muñoz-Pinedo

first: Charles-Antoine

last: Dutertre


title: NF-κB- and TET2-mediated macrophage reprogramming overrides the anti-inflammatory effects of hypoxia and improves responses in human cancer (DNA methylation II).

first: Javier

last: Rodríguez-Ubreva

first: Florent

last: Ginhoux


title: The molecular mechanism and clinical significance of IGF2BP3 in adenocarcinoma of the esophagogastric junction [RIP-seq]

summary: To reveal the potential targets of IGF2BP3 in  Adenocarcinoma of the esophagogastric junction, we performed IGF2BP3 Immunoprecipitation sequencing (RIP-seq) in OE-19 cells.

overall_design: OE-19 cells lysate was Immunoprecipitation by IGF2BP3, then subjected to RNA-sequencing analysis


title: TIme-cource single-cell RNA-seq analysis of bleomycin and LPS-induced lung injury and fibrosis

city: Chiba

first: Shigeyuki

last: Shichino

address: 2641, Yamasaki, Noda-shi

department: Department of Molecular regulation of Inflammatory and Immune Diseases

institute: Tokyo University of Science

summary: Pulmonary fibrosis (PF) is a progressive fibrotic disease with a poor prognosis and suboptimal therapeutic options. To construct comprehensive single-cell atlas and identify disease-relevant cell subsets and gene signatures of the murine models of PF pathogenesis and lung repair, time-course singel-cell RNA-seq analysis of bleomycin (BLM) and LPS-induced lung injury/fibrosis models were performed. We sampled mouse lungs of untreated, day3, 7, 14, 28, 42, 63 (BLM model, 1.25 mg/kg), day3, 7, 14 (3 mg/kg), and day7, 14, 42 (LPS model, 1mg/ml, 50ul/head) with 5 biological replicates (except BLM 3mg/kg at day 14 which includes 2 biological replicate)  and processed by using TAS-Seq protocol (Shichino et al. Commnue Biol 2022). We found that some of the early phase-specific markers, and repair phase-specific cell subsets and associated gene signatures. Of these, we found that Sftpb was an early marker of lung injury that was specifically expressed on AT2 cells, and SFTPC protein also highly expressed in the serum extracellular vesicles from prognosis-worsen PF-ILD patients. Our dataset might useful to clarify lung injury and repair mechanisms.

overall_design: We sampled mouse lungs of untreated, day3, 7, 14, 28, 42, 63 (BLM model, 1.25 mg/kg), day3, 7, 14 (3 mg/kg), and day7, 14, 42 (LPS model) with 5 biological replicates (except BLM 3mg/kg at day 14 which includes 2 biological replicate) and processed by using TAS-Seq protocol (Shichino et al. Commnue Biol 2022).

first: Bin

last: Wu

first: Satoshi

last: Ueha

first: Kouji

last: Matsushima


title: The molecular mechanism and clinical significance of IGF2BP3 in adenocarcinoma of the esophagogastric junction [LACE-seq]

summary: To reveal the potential targets of IGF2BP3 in  Adenocarcinoma of the esophagogastric junction, we performed linear amplification of complementary DNA ends and sequencing (LACE-seq) in OE-19 cells.


title: Impact of G167R mutation in CHUK gene, coding for IKKα protein, on gene expression in PBMCs

city: Adelaide

first: Nick

last: Warnock

state: South Australia

address: UniSA Bradley Building, North Terrace

department: Centre for Cancer Biology

country: Australia

institute: SA Pathology

summary: IKKα is a critical regulator of the non-canonical NF-KB signalling pathway. In patients with combined immunodeficiency, we identified a homozygous missense mutation in CHUK gene, coding for IKKα protein, leading to G167R amino acid change in the kinase domain of the protein. This mutation impairs the kinase activity of IKKα, which results in a range of aberratins in innate and adaptive immunity.

overall_design: To investigate the impact of G167R mutation in IKKα, we performed RNA-sequencing on PBMCs from 2 patients and 3 healthy controls.

first: Gokhan

last: Cildir

first: Baran

last: Erman

first: Damon

last: Tumes

first: Nicholas

middle: I


title: RNA-seq of NSC-34 cells expressing SOD1

city: Hummelstown

first: Esther

middle: S

last: Choi

state: PA

address: 1214 Research Blvd

department: Pharmacology

institute: Penn State College of Medicine

summary: Misfolded soluble trimeric species of superoxide dismutase 1 (SOD1) are associated with increased death in neuron-like cell models and greater disease severity in amyotrophic lateral sclerosis (ALS) patients compared to insoluble protein aggregates. The mechanism by which structurally independent SOD1 trimers cause cellular toxicity is unknown but may be a driver of disease pathology. Here, we uncovered the SOD1 trimer transcriptome. We identified key pathways using transcriptomic data from motor neuron-like cells (NSC-34s) expressing SOD1 trimers. We discovered differential gene expression in cells that express SOD1 trimers with selective enrichment of genes responsible for protein localization to membranes and a global upregulation of cellular senescence pathways. Our investigation highlights key protein factors and pathways within each system, revealing a plausible intersection of genetic and pathophysiological mechanisms in ALS through interactions involving SOD1 trimers.

overall_design: We cultured NSC-34 cells and transfected the cells with native SOD1 dimer or non-native SOD1 trimer. We confirmed transfection and differentiated the cells. After differentiation, we harvested the cell lysate and collected the total RNA. We then sent the samples to Novogene for library construction, sequencing, and data analysis.

first: Brianna

last: Hnath

first: Nikolay

last: Dokholyan


title: SRF promotes long-range chromatin loop formation and stem cell pluripotency

city: Berlin

first: Pavel

last: Tsaytler

address: Ihnestrasse  63-73

country: Germany

institute: MPI Molecular Genetics

summary: Serum response factor (SRF) is a transcription factor essential for cell proliferation, differentiation, and migration, and is required for primitive streak and mesoderm formation in the embryo. The canonical roles of SRF are mediated by a diverse set of context-dependent cofactors. Here we show that SRF physically interacts with CTCF and cohesin subunits at TAD boundaries and loop anchors. SRF reinforces the insulation of TADs and promotes the formation of long-range chromatin loops. In ES cells, SRF associates with Oct4, Sox2, and Nanog and contributes to the formation of 3D pluripotency hubs. Our findings reveal new roles of SRF in higher-order chromatin organization.

overall_design: Hi-C analysis in WT and SRF, CTCF, or SRF/CTCF double knock-out non-differentiated or cardiac differentiated mouse ESCs. SRF HiChIP in mouse ESCs. Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for SRF in non-differentiated or cardiac differentiated mouse ESCs. Transcriptomes (RNA-Seq) and chromatin accessibility (ATAC-Seq) from non-differentiated or cardiac mesoderm differentiated SRF wild-type and SRF knock-out mouse ESCs.


title: Deciphering the novel regulatory mechanisms and biological implications of ARID1A C-terminal missense mutations in cancer [ChIP-Seq]

city: Shanghai

first: Fang

last: Liu

address: 227, south chongqin road

institute: Shanghai Jiao Tong University School of Medicines

summary: ARID1A (the AT-rich interaction domain 1A, also known as BAF250a), a pivotal component of the SWI/SNF chromatin remodeling complex, is frequently mutated in various cancers. In this study, we discovered that mutations such as ARID1A V2084D or V2087E lead to the binding of ARID1A mutant with the nucleocytoplasmic shuttling protein XPO1, facilitating ARID1A mutant transport to the cytoplasm. Subsequently, the E3 ubiquitin ligase STUB1 recognizes and ubiquitinates ARID1A mutant protein, marking it for degradation. This reduction in protein levels due to the mutation impedes the assembly of the cBAF complex and the expression of tumor-suppressive target genes, ultimately contributing to a malignant phenotype. Knocked down STUB1 or inhibit XPO1 activity stabilizes the ARID1A mutant protein, retaining it in the nucleus, which restores the assembly of the cBAF complex, chromatin remodeling function, and the expression of critical genes, thereby decrease the tumor burden.

overall_design: ARID1A knocked out Huh7 cells with lentiviral stably expressed ARID1A WT/V2084D-Flag and knocksown STUB1 were analyzed by ChIP-seq analysis.

first: Jun

last: Ying

first: Xiaonan

last: Wang

first: Xuxu

last: Sun


title: Deciphering the novel regulatory mechanisms and biological implications of ARID1A C-terminal missense mutations in cancer [RNA-seq]

overall_design:
ARID1A knocked out Huh7 cells with lentiviral stably expressed ARID1A WT/V2084D and knockdown STUB1 were analyzed by RNA-seq analysis.
ARID1A knocked out 293T cells with lentiviral stably expressed ARID1A WT/V2084D/G2087E/G2087R/K2025E were analyzed by RNA-seq analysis.


title: Changes in heterochromatin distribution during the initial stages of sexual development  in Plasmodium falciparum

city: Barcelona

first: SANDRA

last: NHIM

address: 153 Carrer del Rossello

institute: Isglobal

summary: In the malaria parasite Plasmodium falciparum, changes in the distribution of heterochromatin at specific loci regulate the expression of clonally variant genes (CVGs), which play fundamental roles in host-parasite interactions. In addition to differences in heterochromatin between individual parasites associated with variant expression of CVGs, changes in heterochromatin distribution associated with different developmental stages have been described. However, while stage II/III gametocytes (sexual precursors) have an altered distribution of heterochromatin compared with asexual blood stages, it is unclear if heterochromatin changes already occur at the very early stages of sexual development. Here we used a transgenic parasite line in which sexual conversion can be conditionally induced to characterise the changes in heterochromatin distribution associated with the earliest stages of sexual development. We found that there were essentially no changes in sexual rings or stage I gametocytes compared with asexual blood stages, and remodelling of heterochromatin started at stage II or III of gametocyte development, with changes similar to those previously described in parasites of a different genetic background. Therefore, global redistribution of heterochromatin does not appear to play a role in the initial steps of sexual development. We also found that, for several genes, changes in expression associated with sexual development precede changes in the heterochromatin at their locus.

overall_design: Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for H3K9me3 mark in Plasmodium falciparum asexual or early sexually converted parasites

first: Sandra

last: Nhim

first: Elisabet

last: Tintó-Font

first: Núria

last: Casas-Vila

first: Lucas

last: Michel-Todó

first: Alfred

last: Cortés-Closas


title: RNA analysis of the circadian rhythm of choroidal neovascularization

city: Nantong

first: Aimin

last: Sang

address: xisi road

department: Department of Ophthalmology

institute: Affiliated Hospital of Nantong University

type: Non-coding RNA profiling by high throughput sequencing

summary: Many diseases that affect the heart, brain, and even the eyes originate from vascular pathology, emphasizing the role of vascular regulation. In age-related macular degeneration (AMD), excessive growth of abnormal blood vessels in the eye (choroidal neovascularization) ultimately leads to detachment of retinal pigment epithelium and decreased vision, indicating the importance of choroidal neovascularization in the treatment of age-related diseases. The circadian clock in the mammalian retina regulates various retinal functions, enabling the retina to adapt to the light dark cycle. Emerging evidence suggests a link between circadian clock and retinopathy, but the causal relationship has not yet been determined.

overall_design: In this study, we investigated the regulatory mechanism of ceRNAs in the formation of choroidal neovascularization based on the different light dark cycles of the retina, choroid, and sclera in a mouse model of RPE.

first: Ying

last: Yang


title: Covalent Targeting of Splicing in T Cells

city: New York

first: Jahan

last: Rahman

address: 417 E 68th St

department: Center for Hematologic Malignancies

institute: Memorial Sloan Kettering

summary: Despite significant interest in therapeutic targeting of splicing, few chemical probes are available for the proteins involved in splicing. Here, we show that elaborated stereoisomeric acrylamide chemical probe EV96 and its analogues lead to a selective T cell state-dependent loss of interleukin 2-inducible T cell kinase (ITK) by targeting one of the core splicing factors SF3B1. Mechanistic investigations suggest that the state-dependency stems from a combination of differential protein turnover rates and availability of functional mRNA pools that can be depleted due to extensive alternative splicing. We further introduce a comprehensive list of proteins involved in splicing and leverage both cysteine- and protein-directed activity-based protein profiling (ABPP) data with electrophilic scout fragments to demonstrate covalent ligandability for many classes of splicing factors and splicing regulators in primary human T cells. Taken together, our findings show how chemical perturbation of splicing can lead to immune state-dependent changes in protein expression and provide evidence for the broad potential to target splicing factors with covalent chemistry.

overall_design: Expanded primary human T cells were pelleted (600 g, 5 min, 4 °C) and re-suspended in fresh T cell media at 2 x 106 cells/mL for compound treatment. T cells were treated with WX-02-23 (5 µM), WX-02-43 (5 µM), PladB (10 nM)) or DMSO control in 6-well plates (2 mL/well, 4 x 106 cells/treatment) with or without CD3/CD28 stimulation for 8 h. For treatment conditions that required T cell activation, plates were pre-coated with 1.5 mL PBS containing CD3 (5 µg/mL, BioXCell) and CD28 (2 µg/mL, BioXCell) overnight at 4 °C or for 2 h at 37 °C and washed with PBS (3 mL, 3 times) prior to treatment.

first: Kevin

last: Scott

first: Hiroyuki

last: Kojima

first: Nathalie

last: Ropek

first: Charles

last: Warren

first: Tiffany

last: Zhang

first: Simon

last: Hogg

first: Henry

last: Sanford

first: Caroline

last: Webster

first: Xiaoyu

first: Bruno

last: Melillo

first: Benjamin

middle: F

last: Cravatt

first: Jiankun

last: Lyu

first: Omar

last: Abdel-Wahab

first: Ekaterina

middle: V

last: Vinogradova


title: RNA sequencing analysis of isoginkgetin and thapsigargin treated HCT116 and HCT116 ATF4 deficient cells

city: Ottawa

first: John

last: Stead

state: ON

address: 1125 Colonel By Drive

department: Department of Neuroscience

country: Canada

institute: Carleton University

summary: Activating transcription factor 4 (ATF4) is activated during cellular stress through a pathway called the integrated stress response (ISR). We had previously reported that the splicing inhibitor isoginkgetin (IGG) activates ATF4 and ATF4-dependent transcripts. To determine the role of ATF4 in the transcriptional response to IGG, we used tandem CRISPR cas9 gene editing to create an ATF4 deficient HCT116 (colon cancer) cell line. We completed RNA sequencing on HCT116 parental and HCT116 ATF4 deficient cells treated with IGG, and thapsigargin (Tg), a positive control for ATF4 activation. We found that IGG led to the differential expression of 76 transcripts, and 58 of these were dependent on ATF4. Tg led to a far more robust transcriptional response, which appeared to be less ATF4 dependent.

overall_design: Total RNA was collected from mock-treated, vehicle control-treated (DMSO), IGG (30µM) and Tg (2µM) drug treated HCT116 and HCT116 ATF4 deficient cells following an 8h exposure, of a 2h vehicle control (DMSO) and 2h Tg (2µM) treatment. Four biological experiments were included in both cell lines. RNA was quantified using the Qubit 4 Fluorometer with the Qubit RNA Broad range Assay kit. RNA quality was determined using the 4200 TapeStation System. Library prep was completed following the Illumina stranded mRNA prep reference guide, and pooled samples were loaded into a NextSeq 2000 Reagent Cartridge and sequenced using an Illumina NextSeq 2000. The data was analyzed using the Omics data analysis framework for regulatory application (R-ODAF) pipeline (https://github.com/R-ODAF/R-ODAF_Health_Canada/) and differentially expressed genes were identified using the DESeq2 package.

first: Erin

last: van Zyl

first: Claire

last: Peneycad

first: Carole

last: Yauk

first: Bruce

last: McKay


title: RNA-seq of peptide RSM3 treatment in prostate cancer cells

city: Changsha

first: Dingxiao

state: Hunan

address: Yuelushan

department: School of BioMedical Sciences

institute: Hunan University

summary: METTL3 play nonnegligible function in PCa progression.RSM3,RSM3, a peptide inhibitor targeting METTL3. Here, we seek to explore the effect of RSM3  on transcriptome profile in PC3 cells

overall_design: RSM3 treatment versus control in PC3 cell lines, three replicates for each group.

first: Zenghui

last: Li

first: Yuqing

last: Feng


title: METTL3 in prostate cancer cells

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

overall_design: Refer to individual Series


title: m6A seq in prostate cancer cell lines

summary: In order to reveal the biological role of m6A in prostate cancer, we conducted m6A-seq on two pairs of cancer/paracancer tissues.

overall_design: two replicates for each group.


title: RIP seq in prostate cancer cell lines

summary: The function of METTL3 as an m6A writer is widely known to involve binding RNA and catalyzing m6A modification.To reveal the target gene landsCAPe of METTL3 in prostate cancer cell lines, we applied RIP-seq with a specific METTL3 antibody in PC3 and LNCAP cells.


title: Intracellular zinc protects tumors from T cell-mediated cytotoxicity

city: Melbourne

first: Jane

last: Oliaro

state: Victoria

address: 305 Grattan St

institute: Peter MacCallum Cancer Centre


title: Serum Apolipoprotein H Alters Cellular Lipid Composition to Inhibit Ferroptosis

first: Xiang

last: He

address: 280 South Chongqing Road, Shanghai, China

institute: Shanghai Jiaotong University School of Medicine

summary: Ferroptosis is an iron-dependent regulated cell death caused by the accumulation of lipid peroxidation for the uncontrolled metabolism. Serum, as the major medium for the cultured cells, resembles the contents of the extracellular fluid in vivo and provides biomolecules for cellular metabolism. The efficiency of ferroptosis induction is influenced by several factors including the extracellular environment. However, the effect of serum on ferroptosis remains largely unclear. We found that cells cultured in different serums have varying efficiencies in ferroptosis induction. By purifying and identifying active serum components, we discovered that serum protein apolipoprotein H (APOH) play essential role in inhibiting ferroptosis. Moreover, APOH activates the phosphoinositide 3-kinase (PI3K)/AKT-Sterol regulatory element-binding proteins (SREBPs) pathway. SREBPs upregulate the stearoyl-CoA desaturase (SCD) increasing cellular monounsaturated fatty acid-containing phospholipids (MUFA-PLs), leading to ferroptosis inhibition. Our findings indicate that APOH, as an extracellular protein, plays an important role in cellular lipid metabolism and inhibition of ferroptosis, thus may having therapeutic applications in cancer treatment and ferroptosis-related diseases.

overall_design: Given the impact of serum on cellular sensitivity to ferroptosis, we aimed to elucidate the mechanisms by which R-FBS (resistance FBS to ferroptosis)mediates cellular resistance to ferroptosis. We analyzed the transcriptome of MEFs cultured in Rcm(R-FBS complete medium), Scm (sensitive to ferroptosis, S-FBS, the S-FBS complete), and exchanged medium for 12h (referred to as R-Scm-12h or S-Rcm-12h, respectively) and 24h (referred to as R-Scm-24h or S-Rcm-24h, respectively) by RNA sequencing (RNA-seq)

first: Jiahui

last: Zhang

first: Masha

last: Huang

first: Jie

last: Wang

first: Simin

last: Yang

last: Yu

first: Yingjie

last: Xu

first: Wen


title: RNA-seq of METTL3 knockdown in prostate cancer cells

summary: Prostate cancer (PCa) is the most frequently diagnosed malignancy in men worldwide. METTL3 play nonnegligible function in PCa progression. Here, we seek to explore the effect of METTL3  on transcriptome profile in PC3 cells via RNA-seq.

overall_design: METTL3 knockdown versus control in PC3 cell lines, two replicates for each group.


title: Dynamic changes in gene expression of growing nonhuman primate antral follicles

city: East Lansing

first: Peter

last: Schall

address: 474 S. Shaw Lane

department: CMIB ANS

institute: Michigan State University

summary: Ovaries from adult female rhesus monkeys were obtained on a serendipitous basis from animals going to necropsy for various reasons, none of which would be expected to impact ovarian function. The follicle size categories to be collected were determined based on the fact that in rhesus monkeys, follicles that produce mature oocytes are generally larger than 1mm and in each pair of naturally cycling ovaries there were generally fewer than five follicles in this category. Three size categories of follicles were established: less than 0.5, 0.5 to 1.0 and greater than 1.0.   All samples from each size category were pooled for each animal, resulting in three pools of granulosa, three of cumulus and three of oocytes for each animal. Samples were processed for RNA extraction followed by RNAseq on the Illumina HiSeq 2000 platform.

overall_design: RNA-sequencing of rhesus monkey cumulus, granulosa, and oocytes from three antral follicle sizes.

first: Catherine

middle: A

last: VandeVoort

first: Charles

last: Chaffin

first: Schall

middle: Z

last: Peter

first: Keith

last: Latham


title: Pancreatic ductal adenocarcinoma tumor growth requires SREBP pathway activity

city: Baltimore

last: Espenshade

state: Maryland

address: 725 N Wolfe Street, Physiology 107

department: Cell Biology

institute: Johns Hopkins University

summary: Tumor growth outstrips local nutrient supply, making metabolic reprogramming a necessary component of oncogenesis and cancer progression. The supply of lipids such as cholesterol and fatty acids is required for continued tumor cell division. Sterol regulatory element-binding protein (SREBP) transcription factors control cellular lipid homeostasis by activating genes required for cholesterol and fatty acid synthesis and uptake. SREBPs have been implicated in the progression of multiple cancers, including glioblastoma, breast, colon, liver and prostate. However, the role the SREBP pathway and its central regulator SREBP cleavage activating protein (SCAP) in pancreatic ductal adenocarcinoma (PDAC) has not been studied in detail. Here, we demonstrate that SREBP target genes are upregulated in PDAC tumors, and SREBPs are upregulated in patient-derived PDAC cell lines under low serum conditions that mimic the tumor microenvironment. Chemical or genetic inhibition of the SREBP pathway prevented PDAC cell growth under low serum conditions due to a lack of lipid supply. Using subcutaneous and orthotopic xenograft models, we showed that SCAP is required for PDAC tumor growth. Pancreas-specific knockout of Scap had no effect on mouse pancreas development or function, allowing examination of the role for Scap in the murine KPC model of PDAC. Notably, heterozygous loss of Scap significantly prolonged survival in KPC mice, and homozygous loss of Scap impaired PDAC tumor inception. Collectively, these results demonstrate that SCAP and SREBP pathway activity are essential for PDAC cell and tumor growth in vitro and in vivo, identifying SCAP as a potential therapeutic target for PDAC.

overall_design: Total RNA isolated from patient derived pancreatic adenocarcinoma cells cultured in 0.5% FBS compared to same cells cultured in 10% FBS.

first: Chiaki

last: Ishida

first: Stephanie

last: Meyers

first: Wei

last: Shao

first: Meredith

middle: R

last: McGuire

first: Chune

last: Liu

first: Casie

last: Kubota

first: Theodore

last: Ewachiw

first: Debaditya

last: Mukhopadhyay

first: Suqi

last: Ke

first: Hao

first: Zeshaan

last: Rasheed

first: Robert

last: Anders


title: Deciphering the novel regulatory mechanisms and biological implications of ARID1A C-terminal missense mutations in cancer [ATAC-Seq]

first: Fang

overall_design: ARID1A knocked out Huh7 cells with lentiviral stably expressed ARID1A WT/V2084D and knocksown STUB1 were analyzed by ATAC-seq analysis.

first: Jun

last: Ying

first: Xiaonan

last: Wang

first: Xuxu

last: Sun


title: Single cell RNA-sequence reveals the transcriptional dynamics of tumor immune micrenvironment by LDHA inhibition in cancer cells

city: Fukuoka

first: Chikanori

last: Tsutsumi

address: 3-1-1 Maidashi, Higashi-ku

department: Department of Surgery and Oncology, Graduate School of Medical Sciences

institute: Kyushu University

summary: In this study, we investigeted the impact of LDHA inhibition in cancer cell on the tumor immune microenvironment (TIME) of orthotopic syngeneic PDAC tumors. Our results indicate that the TME with high-glucose and low-lactate can promote the antigen-presenting function of DCs and enhance the activity of CD8+ T cells.

overall_design: 1 sample integrated from 5 orthotopic syngeneic PDAC tumors transplanted KPC shNC. 1 sample integrated from 5 orthotopic syngeneic PDAC tumors transplanted KPC shLDHA.

first: Bo

last: Zhang

first: Kenoki

last: Ohuchida


title: Single-cell RNA sequencing reveals peripheral immunological responses and cell-specific biomarker in patients with Parkinson’s disease

city: Chengdu

first: Li

last: Chen

state: Sichuann

address: No.17, South Renmin Road

institute: Sichuan University West China Hospital

summary: Parkinson's disease (PD) is a neurodegenerative disease characterized by bradycardia, static tremor, rigidity and postural instability. At present, the current knowledge remains inadequate for the connection between the disease severity and peripheral immune response. In the present study, using single-cell RNA sequencing (scRNA-seq), we profiled peripheral blood mononuclear cells (PBMCs) from clinical cases, including 2 early PD, 2 advance PD and 2 matched controls, representing five major immune cell subsets: T cells (CD4+, CD8+), monocytes, B cells, natural killer (NK) cells and hematopoietic stem cells (HSCs). The characteristic alterations of cell subset proportions were disentangled, in which T cells and NK cells were declined in the progression of PD. We further enlarged the sample size and collected peripheral blood samples of 14 normal individuals, 21 early and 14 advance PD patients. T cells and NK cells showed an obvious reduction in the blood samples as the disease proceeded. Moreover, NK cells infiltration into cerebral motor cortex indicated by immunostaining of human brain sections evidenced the close communication existing between peripheral immune response and central nervous system. Importantly, NK cells participated in numerous common immune-related biological processes. Strikingly, NK cell-specific XCL2 that was specifically expressed in cluster 6 was further identified and positively correlated with the PD severity. All these findings delineated the critical role of peripheral adaptive immune response, especially mediated by NK cells, in the pathogenesis of PD, providing clues of immune-related biomarkers or therapeutic targets during PD progression in future investigations.

overall_design: In this study, we presented 5 major immune cell subsets, T cells (CD4+, CD8+), B cells, NK cells, monocytes and HSCs, and displayed the distinct proportion of cell subsets and different gene expression patterns in the progression of PD by scRNA-seq analysis. Among these cell subsets, T cells and NK cells were found decreased as the disease proceeded, which was evidenced by the decreased percentage of T cells and NK cells in PD patients with the increase of severity with the clinical sample size enlarged. Moreover, the NK cell type-specific key genes were proved to be involved in PD related signal transduction and immune regulation. Correlation analysis revealed the number of NK cells was negatively correlated with the severity of PD. Strikingly, NK cells were found expressed in the motor cortex of human brain sections from PD patients. Importantly, NK cell-specific crucial gene was identified in the present study (XCL2), which might present great potentials as PD-associated biomarkers.

first: Liu-Lin

last: Xiong

first: Lu-Lu

last: Xue

first: Ruo-Lan

last: Du

first: Rui-Ze

last: Niu

first: Xue-Yan

last: Huang

first: Chang-Yin

last: Yu

first: Jia

first: Ting-Hua


title: Elucidating the role of miRNA in the modulation of Survival Motor Neuron (SMN) genes in both fibroblasts and firboblast-derived iPSCs.

city: Subang Jaya

first: Wendy

middle: Wai Yeng

last: Yeo

state: Selangor

address: Jalan Lagoon Selatan, 47500 Bandar Sunway,

department: Pharmacy

country: Malaysia

institute: Monash University

summary: Spinal muscular atrophy (SMA) is a devastating neuromuscular disorder that affects the spinal motor neurons and leads to progressive muscle wasting and atrophy. It is caused by a reduction in SMN protein levels due to the mutations in the survival motor neuron 1 (SMN1) gene. Human are unique as they possess a homologous pseudogene known as survival motor neuron 2 (SMN2) gene. MicroRNAs (miRNAs) play a role in either translational repression or mRNA degradation. It has been highlighted that dysregulation of miRNA has been a common feature of motor neuron disease such as SMA. Moreover, it is speculated that the dysregulation of miRNAs expression contributes to the pathophysiology of SMA and the vulnerability of SMN protein can be altered by the modulation of specific miRNA. However, there are still lacking of studies on the dysregulation of miRNAs in human SMA patients using iPSC cell models and how the miRNAs correlate with the SMN protein. Hence, we utilized miRNA microarray to identify the miRNAs dysregulated in SMA patients as compared to normal controls in both fibroblast and its derivative induced pluripotent stem cells (iPSCs). Human fibroblasts and iPSCs were cultured and their respective RNA were extracted.

overall_design: Human fibroblast and iPSCs were cultured and their respective RNA were extracted. Each samples were grown with at least triplicates.

first: Gayatri

last: Gandhi

middle: W


title: Healthy aging of skeletal muscle: Insights from a combined morphological and multiomic study

city: Giessen

first: Marek

last: Bartkuhn

state: Hessen

address: Aulweg 132

department: Biomedical Informatics and Systems Medicine

institute: Justus-Liebig-University Giessen

summary: This study shows age-related differences in the molecular signatures of healthy skeletal muscle. Moreover, the findings assert that processes previously associated solely with aging and sarcopenia are indeed part of healthy aging. Additionally, the data underscores the importance of using appropriate age-matched controls for multi-omic molecular profiling of muscular disorders.

overall_design: Comparative age related transcriptomic analysis of human skeletal muscle samples

first: Anne

last: Schänzer

first: Alexenader

last: Schaiter


title: WNT2 blockage augments anti-tumor immunityby reducing myeloid-derived suppressor cell accumulation and function in colorectal cancer

city: Shenzhen

first: Albert

last: CUI

state: Guangdong

address: Xueyuan Avenue, Nanshan District, Shenzhen

institute: Shenzhen University

summary: To investigate the role of WNT2 on regulation of  bone marrow-derived MDSCs proliferation and function,  we extracted total RNA from untreated and mrWNT2-treated bone marrow-derived MDSCs for RNA sequencing.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for control cells and mrWNT2 treated cells.

last: Fu

first: Cheng

last: cui

first: Tian

middle: T


title: Next Generation Sequencing Facilitates Quantitative Analysis of WT and GSH or Ctrl and Cas9 mouse breast cancer cells Transcriptomes

city: Beijing

first: Chenxi

last: Zhao

address: 1 Tian Tan Xi Li

institute: Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences & Peking Union Medical College

summary: Purpose: Next-generation sequencing (NGS) has revolutionized systems-based analysis of cellular pathways. The goals of this study are to analysis the differiational genes and pathways in WT and WT+GSH or Ctrl-Cas9 and Scarb2-Cas9 mouse breast cancer cells by using NGS-derived transcriptome profiling (RNA-seq).  Methods:  4T1 cells treated with or without GSH (500 µM) and 4T1 stably expressing Ctrl-Cas9 or Scarb2-Cas9 were collected. These cells' mRNA profiles were generated by deep sequencing, in triplicate, using Illumina HiSeq 4000. The sequence reads that passed quality filters were analyzed at the transcript isoform level with  following methods: Alignment by using HISAT2 v2.1, IGV was used to to view the mapping result by the Heatmap, histogram, scatter plot or other stytle, FPKM was then calculated to estimate the expression level of genes in each sample, DEGseq v1.18.0 was used for differential gene expression analysis between two samples with non biological replicates and Function Enrichment Analysis including  GO enrichment analysis and KEGG .  Conclusions: Our study represents the first detailed analysis of WT and WT+GSH or Ctrl-Cas9 and Scarb2-Cas9 mouse breast cancer cells' transcriptomes, with biologic replicates, generated by RNA-seq technology. The optimized data analysis workflows reported here should provide a framework for comparative investigations of expression profiles. Our results show that NGS offers a comprehensive and more accurate quantitative and qualitative evaluation of mRNA content within a cell or tissue. We conclude that RNA-seq based transcriptome characterization would expedite genetic network analyses and permit the dissection of complex biologic functions.

overall_design: WT and GSH or Ctrl and Cas9 mouse breast cancer cells' mRNA profiles were generated by deep sequencing, in triplicate, using Illumina HiSeq 4000.

first: Ke

last: Li

first: Zhuorong

first: Tingting

last: Zhang


title: Negative feedback of cyclic di-GMP levels optimizes switching between sessile and motile lifestyles in Vibrio cholerae [RNA-seq]

city: Ann Arbor

first: Lydia

last: Freddolino

state: MI

address: 1136 Catherine St

department: Biological Chemistry

institute: University of Michigan

summary: The signaling molecule cyclic di-GMP (cdG) controls the switch between bacterial motility and biofilm production, and fluctuations in cellular levels of cdG have been implicated in Vibrio cholerae pathogenesis. Intracellular concentrations of cdG are controlled by the interplay of diguanylate cyclase (DGC) enzymes, which synthesize cdG to promote biofilms, and phosphodiesterase (PDE) enzymes, which hydrolyse cdG to drive motility. To track the complete regulatory logic of how V. cholerae responds to changing cdG levels, we followed a time course of overexpression of either the V. harveyi diguanylate cyclase QrgB or a variant of QrgB lacking catalytic activity (QrgB*). We find that QrgB increases cdG levels relative to QrgB* for 30 minutes after overexpression, but the effect of QrgB on cdG levels plateaus at 30 minutes, indicating tight adaptive control of cdG levels. In contrast, loss of VpsR, a master regulator activating biofilm formation upon binding to cdG, leads to higher baseline levels of cdG and continuously increasing cdG through 60 minutes after QrgB induction, revealing the existence of a negative feedback loop on cdG levels operating through VpsR. Through a combination of RNA polymerase ChIP-seq, RNA-seq, and genetic approaches, we show that the PDE CdgC is activated by VpsR at high cdG concentrations, mediating this negative feedback on cdG levels. We further identify a transcript encoded within, and antisense to, the cdgC open reading frame which we name sRNA negative regulator of CdgC (SnrC). RNA polymerase ChIP-seq and RNA-seq demonstrate SnrC to be expressed specifically under conditions of high cdG in the absence of VpsR. Ectopic SnrC expression increases cdG levels in a manner depending on CdgC, demonstrating that its effect on cdG levels is likely through interference with CdgC production. Further, although cells lacking cdgC exhibit enhanced biofilm formation, these mutants are outcompeted by wild type V. cholerae in colonization assays that reward a combination of attachment, dispersal, and motility. These results underscore the importance of negative feedback regulation of cdG to maintain appropriate homeostatic levels for efficient transitioning between biofilm formation and motility, both of which are necessary over the course of the V. cholerae infection cycle.

overall_design: Chromatin immunoprecipitation (ChIP) of RNA polymerase in Vibrio cholerae in the following genotypes: WT, delta-vpsR, delta-vpsT

first: Aathmaja

last: Rangarajan

first: Jeremy

last: Schroeder

first: Rebecca

middle: L

last: Hurto

first: Geoffrey

middle: B

last: Severin

first: Meng-Lun

last: Hsieh

first: Christopher

middle: M

last: Waters


title: Perinatal correlated retinal activity is required for the wiring of visual axons in non-image forming nuclei

city: San Joan d'Alacant

first: Eloísa

last: Herrera

address: Av Ramon y Cajal s/n

department: Instituto Neurociencias Alicante

institute: CSIC-UMH

summary: The formation of the visual system is a complex multistep process that includes the establishment of  proper connectivity of retinal ganglion cell (RGC) axon terminals with their relay neurons located in the brain. In mammals, the assembly of the different components of the visual circuit occurs at perinatal stages before eye opening. Upon reaching the target nuclei RGC axons extensively arborize and subsequently refine to establish the final connections. Spontaneous activity generated in the immature retina plays an essential role in the refinement of visual terminals at the main image-forming nuclei (IFN) that follow an eye-specific and retinotopic organization. However, the molecular mechanisms underlying spontaneous activity-dependent axon remodeling, and the influence of this activity in the connectivity of non-image forming nuclei (NIFN) that lack precise retinotopic maps and/or eye-specific segregation, are not well known. Here, by generating conditional mice with disturbed spontaneous retinal aneactivity and analyzing their retinal transcriptomic profiles, we identified novel players involved in axon refinement at the visual nuclei (e. g. Syt13).  The analysis of visual projections in the NIFN of these mice revealed that correlated-retinal activity shapes final connectivity in non retinotopic or eye-specific segregating visual nuclei.

overall_design: RNA-seq

first: Maria

last: López-Cascales

first: Santiago

last: Negueruela

first: Cruz

last: Morenilla Palao

first: Eloisa

last: Herrera González de Molina


title: Gene expression changes in human thoracic and abdominal aortic aneurysm tissue

city: Iowa City

first: Long-Sheng

last: Song

state: IA

address: 285 Newton Rd

department: Internal Medicine

institute: University of Iowa

summary: To determine how gene expression is altered in aorta tissue in response to aortic aneurysm disease. Thoracic or abdominal aorta tissue was isolated from patients requiring surgery due to aortic aneurysm or other (control) reason.

overall_design: Gene expression profiling of RNA-seq data was performed on total RNA isolated from 4 samples per group.

first: Duane

last: Hall

first: Jose

last: Gomez


title: Differences in the gene expression profile of HIV- and CMV-specific CD8 T cells in HIV infection

city: Amsterdam

first: Lisa

last: van Pul

address: Meibergdreef 9

institute: Amsterdam UMC

summary: Here we studied the transcriptional profile of virus specific CD8 T cells to gain molecular insights in CD8 T cell functionality in HIV infection. HIV- and CMV-specific CD8 T cells were isolated from HIV infected individuals participating in the Amsterdam Cohort Studies (HIV progressors, long-term non-progressors (LTNP; HLA-B*57 and non-HLA-B*57) and individuals carrying the MAVS minor genotype) and CMV seropositive blood donors (BD). The transcription profile of HIV-specific CD8 T cells of LTNP groups was associated with increased protein/RNA metabolism pathways, indicating that immune control of HIV infection in these individuals is associated with increased functionality. In contrast, CMV-specific CD8 T cells from progressors showed increased expression of genes related to effector functions and suggests recent CMV reactivation. Our study provides novel insights into molecular mechanisms involved in HIV and CMV control in chronic HIV infection.

overall_design: RNA sequencing of HIV- and CMV-specific CD8 T cells obtained from different groups (B*57 LTNP, non-B*57 LTNP, MAVS-/-, Progressors) of HIV-infected individuals and blood donors (for CMV-specific CD8 T cells only).

first: Neeltje

last: Kootstra


title: Transcriptional profiling of biomass degradation-related genes during Herbivorax saccincola A7 growth on different carbon sources

city: Tsukuba

first: Sirilak

last: Baramee

state: Ibaraki

address: 1-1 Ohwashi

institute: Japan International Research Center for Agricultural Sciences

summary: Herbivorax saccincola A7 is an alkali-thermophilic lignocellulolytic bacterium that was known to possess a strong xylan degradation ability. They can utilize a wide range of carbon sources, however the response and regulation mechanisms to different carbon sources, in terms of genes expression, are still not identified. In this study, H. saccincola A7 was cultured with cellobiose, cellulose, xylan, cellulose-xylan mixture, and empty fruit bunch (EFB) as the sole carbon source. We carried out a comparative expression analysis, focusing on biomass degradation-related genes. The cellulosomal scaffolding proteins and cellulosomal enzymes of H. saccincola A7 were drastically induced when a cellulose-containing substrate was used as a carbon source. On the other hand, xylan induced only some non-cellulosomal enzymes that might be necessary for xylan degradation. The identification of the carbohydrate-sensing mechanism in H. saccincola A7 appeared a similar system as previously described in Clostridium thermocellum. The lignocellulose degradation ability of H. saccincola A7 may be controlled enzyme expression by cellulose-containing substrate, not by xylan. Thus, recognition in the existence of cellulosic substrates is effective in enhancing a high xylan degradation ability in H. saccincola A7.

overall_design: The 5 samples were analyzed. H. saccincola A7 cell growth on different carbon sources were used to extracted total RNA for gene expression analysis. We examined the selection of the calibrator gene and found that Triosephosphate isomerase (Tpi) is the best calibrator gene for H. saccincola A7

first: Akihiko

last: Kosugi


title: RNA-seq analysis of BL21 (DE3) cells for AcK biosynthesis

status: Public on Oct 08 2024

city: Nanjing

first: Jinyuan

last: Xu

address: No. 138, Xianlin Street, Qixia District, Nanjing City, Jiangsu Province

institute: Nanjing University of Traditional Chinese Medicine

summary: We performed RNA-Seq to accurately assess the impact of AcK addition or biosynthesis on gene expression in E. coli cells.

overall_design: BL21 (DE3) cells were transformed with either NmGNAT or AcK or negative control.

first: Nanxi

last: Wang


title: PP2A B''a and B''b regulation gene expression in dark to red transition in Arabidopsis thaliana.

city: Austin

first: Enamul

last: Huq

state: TX

address: NHB 2.616, Stop A5000 100 East 24th St.

institute: University of Texas at Austin

summary: The mRNA expression profiles of the WT and b''ab''b  were analyzed in 1 hour 8 μmol/m2s  red-light exposure and darkness after 4-day dark-grown. The gene expression changes was analyzed and compared between dark and red-light treated samples.

overall_design: Detect the gene expression in WT and b''ab''b after red light teatment. Each samples for each condition we have 3 replicates. Totally, we have 12 samples

first: Xingbo

last: Cai