title: A smooth muscle cell lncRNA controls angiogenesis in chronic limb-threatening ischemia through a miR-143-3p/HHIP signaling axis

city: Boston

first: Ming

last: Zhai

state: Massachusetts

address: Avenue Louis Pasteur

department: Cardiovascular Division, Department of Medicine

institute: Brigham and Women's Hospital

summary: Peripheral artery disease (PAD), affecting over 200 million people globally, often advances to chronic limb-threatening ischemia (CLTI), resulting in severe complications such as limb amputation and heightened mortality. Despite the potential of therapeutic angiogenesis, the mechanisms of cell-cell communication and transcriptional changes driving PAD are not fully understood. Profiling long non-coding RNAs (lncRNAs) from human gastrocnemius muscles of human subjects with or without CLTI revealed that a vascular smooth muscle cell (SMC)-enriched lncRNA CARMN was reduced with CLTI. This study explored the role of how SMC-driven regulation of angiogenesis in limb ischemia is conferred via a unique lncRNA-miRNA coupled signaling pathway.To investigate this interesting project, we perform three RNA-seq project. The first one was performed between hypoxia and normoxia VSMCs to mimic the HLI disease in patients. The second one was performed between CARMN KO SMC and CARMN WT SMC to find the potential downstream signaling pathway. The third one was about the RNA-seq of the gastro muscle obtained from CARMN KO and CARMN WT mice after HLI surgery.

overall_design: To explore this intriguing research question, we conducted three RNA-seq experiments. The first experiment compared VSMCs under hypoxic and normoxic conditions to simulate the HLI disease state observed in patients. The second experiment analyzed CARMN knockout (KO) SMCs versus CARMN wild-type (WT) SMCs to identify potential downstream signaling pathways. The third experiment involved RNA-seq analysis of gastrocnemius muscle tissue obtained from CARMN KO and CARMN WT mice following HLI surgery.

first: Anurag

last: Jamaiyar

first: Winona

first: Emre

last: Bektik

first: Mark

middle: W

last: Feinberg


title: Noncanonical role of NCOR1 as a facilitator of DNA mismatch repair and its deficiency sensitizes cancers to immune checkpoint blockade therapy

status: Public on Mar 01 2025

city: Beijing

first: Xuejiao

last: Ren

address: Beijing

country: China

institute: Nanjing Medical university

type: Expression profiling by high throughput sequencing

summary: Tumor genomic heterogeneity significantly influences the efficacy of immune checkpoint blockade (ICB) therapy. To elucidate the underlying mechanisms, we pioneered the development of an ICB response score (IRS) based on machine learning analysis of microenvironmental features. Through a pan-cancer genomic screening, we identified the nuclear receptor corepressor NCOR1 as a key regulator, the tumors with genomic deficiency of which display a tumor infiltrating lymphocyte (TIL)-inflamed microenvironment. Genetic ablation of Ncor1 delays the tumorigenesis and metastasis in syngeneic and spontaneous tumor models in immunocompetent mice. Mechanistically, NCOR1 guards DNA mismatch repair by directly interacting with MSH2 and recruiting MutSa to chromatin for mismatch recognition, independent of its transcriptional regulatory activity coupling with histone deacetylase. Ncor1 depletion in hepatocellular carcinoma and melanoma cells promotes tumor immunogenicity by increasing tumor mutation burden (TMB), neoantigen load, and TIL infiltration and activation, thereby enhancing anti-PD-1 therapy efficacy. Our findings reveal that NCOR1 loss represents a novel mechanism underlying TMB-high tumors with intact MMR components, broadening the application of ICB.

overall_design: RNA was extracted from mouse Hepa1-6 and B16 cell lines using the Quick-RNA column purification kit (Zymo, USA). RNA-seq libraries were constructed using the VAHTS mRNA-seq v2 Library Prep Kit (Illumina, USA) according to the manufacturer’s protocol. The libraries were sequenced using Novaseq 6000 (Illumina, USA) at Bioguoke Corporation (Beijing, China). The quality of raw sequencing reads was evaluated using FastQC (v0.12.1). Adaptor sequences and low-quality score bases were trimmed using TrimGalore (v.0.6.7). These reads were then aligned to the mm10 reference mouse genomes using Bowtie2 (v.2.2.5). The fragments per kilobase of exon per million mapped reads (FPKM) and gene counts were computed using RSEM (v.1.2.8), and the differentially expressed genes (DEGs) were analyzed using the DESeq2 (v.1.37) in R package. The functional enrichment analysis of the DEGs was performed using the clusterProfiler (v.4.2.2) in R package, in which the gene ontology (GO) annotations and canonical pathways/gene sets (KEGG and Reactome) were obtained from MSigDB (www.broadinstitute.org/gsea/msigdb; v2022.1.Mm).

first: Gangqiao

last: Zhou


title: Nitrite Protects Against Cardiac Surgery-Associated AKI via Inflammation Suppression: Insights from a Rat CPB Model

city: Sapporo

first: Sho

last: Ohno

address: South1 West 16 291

department: Anesthesiology

country: Japan

institute: Sapporo Medical University

summary: Background: Cardiac surgery-associated acute kidney injury (CS-AKI) is reported in 20-30% of patients undergoing cardiac surgery and, leading to poor outcomes and increased healthcare costs. Because the pathophysiology of CS-AKI is complex and has not been fully elucidated, no effective preventive and interventional strategy has been reported. Nitrite, a nitric oxide (NO) donor under hypoxic conditions, has shown to exert organ-protective effects in various kind of preclinical models, however, its efficacy in CS-AKI remains unclear. Objectives: This study aimed to generate a rat cardiopulmonary bypass (CPB) model to induce AKI, identify key molecular pathways involved in its pathophysiology, and evaluate the renoprotective effects of nitrite. Methods: A CPB model was established using rats and AKI was confirmed through molecular and histological analysis. RNA-seq was performed to identify differentially expressed genes and enriched pathway. Nitrite (2 mg/kg) was intravenously administered prior to CPB, and its effects on kidney injury markers and inflammation-related pathways were evaluated. Results: AKI was successfully produced in our rat CPB model, as evidenced by increased KIM-1 and NGAL expression and renal histological damage. RNA-seq revealed significant activation of multiple inflammation-related pathways, including TNF and NFκB signaling pathway. Nitrite administration significantly reduced KIM-1 and NGAL expression and suppressed these pathways associated with development of AKI. Conclusion: This study underscores the role of inflammation in CS-AKI pathophysiology and demonstrates the renoprotective potential of nitrite. These findings provide a foundation for further investigations into nitrite's mechanisms and its potential clinical application in cardiac surgery.

overall_design: RNA-seq profiling of acute kidney injury after CPB and the renoprotective effects of nitrite using a rat CPB model

first: Yusuke

last: Yoshikawa


title: Lipid droplets impair anti-tumor immunity by disrupting IFNGR1 trafficking via TGN diacylglycerol depletion

city: Nanjing

first: Lingyan

last: Xu

address: Guzhou Road

department: Department of Oncology and Cancer Rehabilitation Centre

institute: The First Affiliated Hospital of Nanjing Medical University

summary: Lipid droplets (LDs) are dynamic organelles that regulate cellular metabolism, yet their role in tumor immune evasion remains unclear. Here, we demonstrate that LDs impair anti-tumor immunity by disrupting the membrane localization of interferon-gamma receptor 1 (IFNGR1). Tumor cells with reduced LD content exhibit heightened susceptibility to immune-mediated cytotoxicity in vitro, in murine immunotherapy models, and in patients undergoing immune checkpoint blockade. Mechanistically, IFNGR1 trafficking to the plasma membrane is dependent on diacylglycerol (DAG) in the trans-Golgi network (TGN). However, LDs sequester DAG, impeding IFNGR1 trafficking and attenuating JAK2-STAT1 signaling. Notably, genetic or pharmacological depletion of LDs enhances tumor sensitivity to anti-PD-1 therapy. These findings establish LDs as immunosuppressive organelles that compromise interferon signaling, highlighting their potential as therapeutic targets to improve cancer immunotherapy outcomes.

overall_design: RNA-seq of HT29 shCTRL and shFITM2 colon cancer cells.

first: Xiaofei

last: Li

first: Yufeng

first: Wen

last: Jiang

first: Xiaohan

last: Wu

first: Qiang

first: Wenjie

last: Guo

first: Yanhong

last: Gu


title: Differential Expression of Small Non-Coding RNAs in Uterine Leiomyomas

city: torrance

first: Omid

last: Khorram

state: CA

address: 1124  w carson st

country: USA

institute: The lundquist Institute at UCLA medical center

type: Non-coding RNA profiling by high throughput sequencing

summary: This study aimed to characterize the expression profile of small non-coding RNA transcripts in leiomyomas compared to their paired myometrium.

overall_design: Comparative analysis of sncRNA expression profiles from RNA-seq data in 19 paired myometrium and leiomyoma specimens

first: Tsai-Der

last: Chuang

first: Nhu

last: Ton

first: Shawn

last: Rysling

first: Daniel

last: Baghdasarian


title: Saturation functional assays of MUTYH resolve clinical variants of uncertain significance

city: Ann Arbor

first: Jacob

last: Kitzman

state: MI

address: 4811 Med Sci II, 1241 E Catherine St

department: Department of Human Genetics

institute: University of Michigan

summary: Variants of uncertain significance (VUS) limit the actionability of genetic testing. A prominent example is MUTYH, a base excision repair factor associated with polyposis and colorectal cancer, which has a pathogenic variant carrier rate approaching 1 in 50 individuals in some populations. To systematically interrogate variant function in MUTYH, we coupled deep mutational scanning with a DNA repair reporter containing its lesion substrate, 8OG:A. Our variant-to-function map covers >97% of all possible MUTYH point variants (n=10,941) and achieves 100% accuracy classifying pathogenicity of known clinical variants (n=247). Leveraging a large clinical registry, we observe significant associations with colorectal polyps and cancer, with more severely impaired missense variants conferring greater risk. We recapitulate known functional differences between pathogenic founder mutations and highlight sites of complete missense intolerance, including residues that intercalate DNA and coordinate essential Zn2+ or Fe-S clusters. This map provides a resource to resolve the 1,122 existing clinical missense VUS in MUTYH and demonstrates a scalable strategy to interrogate other clinically relevant DNA repair factors.

overall_design: To measure missense variant impacts, MUTYH cDNA was split into 10 tiles and each underwent saturation mutagenesis. HEK293 MUTYH KO cells were transduced with each library, then transfected with an 8OG:A DNA repair construct. Repair-positive (GFP+) and negative (GFP-) cells were isolated, and the MUTYH library was deeply sequenced from each.

first: Shelby

last: Hemker

first: Grace

last: Clark

first: Alyssa

last: Bashir

first: Krystal


title: Integrating microbial GWAS and single cell transcriptomics reveals associations between host cell populations and the gut microbiome

city: Wenzhou

first: Chenghao

state: Zhejiang

address: Wenzhou Medical University, Xueyuan Xi Road

institute: Wenzhou Medical University

summary: Microbial genome-wide association studies (GWAS) have uncovered numerous host genetic variants associated with gut microbiota. However, links between host genetics, the gut microbiome and specific cellular context remains unclear. Here, we use a computational framework, scBPS (single-cell Bacteria Polygenic Score), to integrate existing microbial GWAS and single-cell RNA-sequencing profiles of 24 human organs, including the liver, pancreas, lung, and intestine, to identify host tissues and cell types relevant to gut microbes. Analyzing 207 microbial taxa and 254 host cell types, scBPS-inferred cellular enrichments confirmed known biology such as dominant communications between gut microbes and the digestive tissue module and liver epithelial cell compartment. scBPS also identified a robust association between Collinsella and central-veinal hepatocyte subpopulation. We experimentally validated the causal effects of Collinsella on cholesterol metabolism in mice through single-nuclei RNA sequencing on liver tissue to identify relevant cell subpopulations. Mechanistically, oral gavage of Collinsella modulated cholesterol pathway gene expression in central-veinal hepatocytes. We further validated our approach using independent microbial GWAS data, alongside single-cell and bulk transcriptomic analyses, demonstrating its robustness and reproducibility. Together, scBPS enables a systematic mapping of the host-microbe crosstalk by linking cell populations to their interacting gut microbes.

overall_design: To investigate the direct effect of Collinsella on cholesterol metabolism, we conducted an in vivo experiment in which C57BL/6 mice were gavage with C. aerofaciens for four weeks (n=9, C.aero group) and compared with the control group (n=6, Ctrl group). We further performed single-nucleus RNA sequencing (snRNA-seq) on mouse liver tissue to identify cell-subpopulation-specific effects of C. aerofaciens.


title: Protective effect of Jianpi Qinghua Sanyu Yin (JPQHSYY) on raised erosive gastritis (REG) [ncRNA-Seq]

city: Fuzhou

first: Aling

last: Shen

state: Fujian

address: 1 Qiuyang road, Minhou Shangjie

institute: Fujian university of Traditional Chinese Medicine

summary: This study aims to comparatively analyze the differentially expressed lncRNA in tissue samples from patients with Raised Erosive Gastritis (REG), before and after treatment with Jianpi Qinghua Sanyu Decoction (JPQHSYY).

overall_design: RNA sequencing was performed with colonic mice in 2 groups: REG and JPQHSYY treatment (n=5 for each group) to identify the differently expressed lncRNA

first: Liya

last: Liu


title: Protective effect of Jianpi Qinghua Sanyu Yin (JPQHSYY) on raised erosive gastritis (REG) [miRNA-Seq]

summary: This study aims to comparatively analyze the differentially expressed small RNA in tissue samples from patients with Raised Erosive Gastritis (REG), before and after treatment with Jianpi Qinghua Sanyu Decoction (JPQHSYY).

overall_design: RNA sequencing was performed with colonic mice in 2 groups: REG and JPQHSYY treatment (n=5 for each group) to identify the differently expressed small RNA

first: Ying

last: Cheng


title: Deletion of sphingosine 1-phosphate receptor 1 in myeloid cells reduces hepatic inflammatory macrophages and attenuates MASH

city: Rochester

first: Harmeet

last: malhi

address: 200 First Street SW

institute: Mayo Clinic

summary: Background: Immune cell-driven inflammation is a key mediator of metabolic dysfunction-associated steatohepatitis (MASH) progression. We have previously demonstrated that pharmacological sphingosine 1-phosphate (S1P) receptor modulation ameliorates MASH and is associated with attenuated accumulation of intrahepatic macrophage and T cell subsets. Although S1P receptors are expressed on several immune cell types, given the prominent role of monocyte-derived recruited macrophages in the sterile inflammation of MASH, we hypothesized that deletion of S1P receptor 1 (S1P1) on myeloid cells may ameliorate MASH by reducing the accumulation of proinflammatory monocyte-derived macrophages into the liver. Methods: The LyzMCre approach was used to generate myeloid cell-specific knockout mice, termed S1pr1MKO. Littermate S1pr1loxp/loxp mice were used as wild-type (WT) controls. MASH was established by feeding mice a high-fat, fructose, and -cholesterol (FFC) diet for 24 weeks which leads to the development of steatohepatitis and MASH-defining cardiometabolic risk factors. Liver injury and inflammation were determined by histological and gene expression analyses. Intrahepatic leukocyte populations were analyzed by mass cytometry and immunohistochemistry. Results: Histological examination demonstrated a reduction in liver inflammatory infiltrates and fibrosis in FFC-fed S1pr1MKO compared to WT. There was a corresponding reduction in alanine aminotransferase, a sensitive marker for liver injury. As determined by mass cytometry, a significant decrease in recruited macrophages was noted in the livers of FFC-fed S1pr1MKO mice compared to WT. Gene ontology pathway analysis revealed a significant suppression of the peroxisome proliferator-activated receptor gamma (PPARγ) and mitogen activated protein kinase (MAPK) pathways in S1pr1MKO consistent with attenuated MASH in mice. Conclusion: Deletion of S1P1 in myeloid cells is sufficient to attenuate intrahepatic accumulation of monocyte-derived macrophages and ameliorate murine MASH.

overall_design: S1pr1 floxed mice, S1pr1loxp/loxp, (The Jackson Laboratory, stock #019141) were crossed with Lyz2-Cre mice (The Jackson Laboratory, stock #004781) to generate myeloid cell specific knockout mice S1pr1loxp/loxp;Lyz2-Cre/0, designated as S1pr1MKO henceforth. Littermate controls, S1pr1loxp/loxp referred to as wildtype (WT), were employed for all experiments. Starting at 12 weeks of age, mice were placed on a high-fat, -fructose, and -cholesterol (FFC) diet (AIN-76A Western Diet, originally manufactured as D12079B, TestDiet, St. Louis, MO) or standard rodent chow diet (CD) for 24 weeks. Cryopreserved mouse liver tissues were recovered and homogenized in TRIzol reagent (Ambion). After tissue debris was centrifuged down, the supernatant was transferred into separate tubes. Total RNA was extracted using the Quick-RNA MiniPrep Kit (Zymo Research, R1055). RNA quality and yield was assessed by NanoDrop ND1000 (Thermo Scientific, Waltham, MA), then reverse transcribed into cDNA with the iScript cDNA Synthesis Kit (Bio-Rad Laboratories, 1708891). Bulk RNA sequencing was performed at the Genome Analysis Core of Medical Genome Facility at Mayo Clinic, Rochester. mRNA sequencing libraries were prepared and sequenced on an Ilumina HiSeq 2000 instrument at the Mayo Clinic Center for Individualized Medicine Medical Genomics Facility. RNA-seq data were analyzed using the MAP-RSeq pipeline (17). In brief, paired-end reads were aligned to the mouse genome reference mm10 using TopHat (v2.1.0) and gene counts were estimated using the feature Counts (v1.4.6) software based on the Ensembl gene definition files. Gene expression was quantified as reads per kilobase per million mapped reads (RPKM). Protein-coding genes with RPKM ≥1 in at least one sample were extracted and the 43295 genes with the largest between-sample variation were used in hierarchical clustering. A subset of protein-coding genes with RPKM ≥1 with intra-group statistical significance were selected for differential analysis (p-value ≤ 0.5). The differentially expressed genes were identified using the edgeR package (v3.18.1). Differentially expressed genes were selected using p-value ≤ 0.5 (1442 genes). Volcano plot was prepared with R-studio and the package ggplot2. Genes in the integrated analysis were analyzed with Ingenuity pathway analysis (IPA) to uncover common regulatory pathways. IPA mapped 1349 genes. The IPA filter for log Fold change used for further analysis was -0.5 for downregulated and +0.5 on upregulated genes. 505 genes were analyzed for top differentially activated pathways and results along with predicated upstream regulators were displayed.

first: Gopanandan

last: Parthasarathy

first: Nanditha

last: Venkatesan

first: Guneet

last: Singh Sidhu

first: Myeong

last: Jun Song

first: Chieh-Yu

last: Liao

first: Fanta

last: Barrow

last: Malhi


title: A circular RNA overcomes acquired resistance to BET inhibitors by antagonizing IGF2BP2-mediated c-MYC translation in triple-negative breast cancer

city: chengdu

first: PAN

last: YiTong

address: Wangjiang Street

institute: Sichuan University

summary: The bromodomain and extra-terminal domain (BET) proteins are promising therapeutic targets to treat refractory solid tumors including triple-negative breast cancer (TNBC); however, acquired resistance remains a major challenge in the clinic. Therefore, elucidation on the mechanism underlying acquired drug resistance is of practical significance. In this study, we found that abnormally upregulated RNA-binding protein IGF2BP2 acts as crucial contributor to triggering acquired BETi resistance in TNBC by facilitating the translational process of c-myc mRNAs. Considering that IGF2BP2 itself are not ideal drug target for small-molecular compounds, we carried out an RIP-seq experiment and identified circRNA-BISC as a potent IGF2BP2 repressor, which was able to repress both c-MYC translation process and acquired BETi resistance. BISC possesses a typical ‘CAC-linker-XGGX’ motif to specifically bind with IGF2BP2 without interactions with IGF2BP1 and IGF2BP3. The effectiveness and specificity of BISC on IGF2BP2 prompted further evaluation of the BISC-based RNA therapeutics in TNBC. In vitro transcribed and circularized BISC, when combined with OTX-015, achieves impressive tumor remission of the BETi-resistant TNBCs without obvious toxicities. Collectively, our result support BISC as a potent IGF2BP2 repressor and highlight the feasibility of pursuing BISC restoration by circRNA therapeutics to overcome BETi resistance.

overall_design: Both MDA-MB-231 parental cells and acquired BETi resistant cells were probed to RIP-seq using anti-IGF2BP2 mAb or normal rabbit IgG. Enrichment of IGF2BP2-associating circRNAs was measured by deducting the circRNA RPMs in IgG. Delta enrichment of IGF2BP2-associating circRNAs were calculated by minusing enrichment in parental cells from acquired BETi resistant cells. Candidates harboring the minus delta enrichments were considered as circRNAs whose association with IGF2BP2 were decreased upon acquired BETi resistance establishment.

first: Jiawei

last: Guo

first: Yitong

last: Pan


title: Active DNA demethylation is upstream of rod photoreceptor fate determination and required for retinal development [Bisulfite-Seq]

city: St. Louis

first: Brian

middle: Stewart

state: Missouri

address: 660 S Euclid Ave; Campus Box #8096

department: Ophthalmology and Visual Sciences

institute: Washington University

type: Methylation profiling by high throughput sequencing

summary: The goal of this study is to investigate the role of TET enzyme-mediated DNA demethylation on retinal development and cell fate specification. We utilized an allelic series of TET enzyme retinal conditional mouse mutants using the Chx10-Cre-GFP transgene, removing the TET enzymes from retinal progenitor cells. We observe that inhibition of DNA demethylation results in abnormal retinal development and a loss of visual function. Immunohistological and genomic profiling (RNA-seq, scRNA-seq, WGBS, and bACE-seq) indicate that rod photoreceptor gene regulatory networks are inhibited when DNA demethylation is lost, indicating that DNA demethylation is required for NRL and NR2E3 expression, resulting in a retina with increased specification of cone photoreceptors.

overall_design: Whole Genome Bisulfite Sequencing (WGBS) to profile methylated cytosines (5mC and 5hmC) and Bisulfite-assisted APOBEC-coupled epigenetic sequencing (bACE-seq) to profile hydroxymethylated cytosines (5hmC) in post-natal day 21 (P21)  mouse retinas from Cre-negative control and Tet1/Tet2/Tet3 conditional mutant animals. Recombination of floxed TET exons was induced in retinal progenitor cells using the Chx10-Cre-GFP transgene. Experiments were performed on RNA-DNA extractions from retinas, performing biological triplicates for each genotype with paired RNA and DNA samples for each biological replicate being used for RNA-seq, WGBS, and bACE-seq analyses.

middle: S

first: Ismael

last: Hernández-Núñez

first: Alaina

last: Urman

first: John

middle: R

last: Edwards


title: Active DNA demethylation is upstream of rod photoreceptor fate determination and required for retinal development [Human RNA-Seq]

last: Clark

summary: The goal of this study is to investigate the role of TET enzyme-mediated DNA demethylation on retinal development and cell fate specification. We utilized an allelic series of TET enzyme retinal conditional mouse mutants using the Chx10-Cre-GFP transgene, removing the TET enzymes from retinal progenitor cells. We observe that inhibition of DNA demethylation results in abnormal retinal development and a loss of visual function. As human retinoblastoma displays reduced TET3 protein expression and a reduction in 5hmC similar to our TET mutant models, we compare the RNA expression changes across human retinoblastoma and TET mutant retinas to identify common pathways that are altered when 5hmC is lost.

overall_design: RNA-sequencing to perform gene expression patterns in human retinoblastoma samples compared to unaffected human control retinas. 3 retinoblastoma cases and 2 control retinal samples were obtained from a retrospective search of pathology archive records. Tissue blocks were sectioned and RNA isolated from dissected regions of tumor or unaffected retina.

first: Philip

middle: A

last: Ruzycki

first: Rajesh

middle: C

last: Rao

first: Sohini

last: Rebba


title: Active DNA demethylation is upstream of rod photoreceptor fate determination and required for retinal development [snRNA-seq]

overall_design: PIPseq single nucleus RNA-sequencing to profile gene expression patterns within individual retinal cells in post-natal day 21 (P21) mouse retinas from Cre-negative control and Tet1/Tet2/Tet3 conditional mutant animals. Recombination of floxed TET exons was induced in retinal progenitor cells using the Chx10-Cre-GFP transgene. Experiments were performed on pooled samples (> 4 retinas each) from 2 separate nuclear dissociations for each genotype


title: Active DNA demethylation is upstream of rod photoreceptor fate determination and required for retinal development [RNA-Seq]

overall_design: RNA-sequencing to perform gene expression patterns in post-natal day 21 (P21) mouse retinas from Cre-negative control, Tet1/Tet2/Tet3 triple heterozygous Cre-positive, and Tet1/Tet2/Tet3 conditional mutant animals. Recombination of floxed TET exons was induced in retinal progenitor cells using the Chx10-Cre-GFP transgene. Experiments were performed on RNA-DNA extractions from retinas, performing biological triplicates for each genotype.


title: An Interleukin 9-ZBTB18 axis promotes germinal center development of memory B cells

first: xiaocui

last: luo

address: Haidian District

institute: Tsinghua university

summary: Memory B cell (MBC) development from germinal centers (GCs) entail profound changes in cell cycling, localization and survival. Here we examined the mechanisms that induce the memory program, focusing on interleukin (IL)-9, given its importance for normal recall antibody responses. Using adoptive transfer and radiation chimera models, we found that T cell-derived IL-9 was required for MBC development and function. In contrast, B cells deficient in IL-9 generated functionally normal MBCs that support antibody recall normally. IL-9 induced expression of the transcriptional repressor ZBTB18 in GC memory precursor cells and MBCs. ZBTB18 was dispensable for naïve B cell activation and GC formation but required for development of GC-derived MBCs. ZBTB18 directly repressed expression of a suite genes encoding cyclin and cyclin-dependent kinases, pro-apoptotic genes Bid and Casp3, and the GC-retaining factor S1pr2. Lack of IL-9-mediated instruction or intrinsic programming by ZBTB18 impaired GC-derived MBC development and antibody recall. Thus, an IL-9-ZBTB18 axis instructs development of functional B cell memory from GCs.

overall_design: Mice were immunized intraperitoneally with 100 μg NP-KLH (conjugate ratios20, Biosearch Technologies) mixed with 1 μg lipopolysaccharide (LPS, Sigma) in alum (Thermo Scientific). Single-cell suspension of splenocytes was stained in MACS buffer (PBS supplemented with 1% FBS and 5 mM EDTA) containing Fc blocker (supernatant from 2.4G2 cell culture) for 20 min on ice before surface staining with primary antibodies in appropriate concentrations. GC B cells were enriched from pooled splenocytes of 3 to 4 Cd79acre/+Zbtb18fl/fl or Cd79acre/+ mice with anti-mouse GL7 (clone GL7, eBioscience) 28 days after NP-KLH immunization. Memory precursor cells (GL7+FAS+CD38+IgD-CD19+) were then sorted into 4 replicates of ~200 cells per PCR tube containing the lysis buffer. Samples were reverse transcribed with SuperScript™ II Reverse Transcriptase (Catalog:18064022, Invitrogen) and the complement DNA library was constructed with TruePrep DNA Library Prep Kit for Illumina (Catalog:TD501, Vazyme), following the Smart-seq2 protocol. Amplified products were purified with VAHTS DNA Clean Beads (Vazyme) and quantified using Qubit and 2100 Bioanalyzer (Agilent). Quantitative RT-PCR was used for quality control. All libraries were sequenced on a HiSeq X Ten sequencer (Illumina).

first: Hai

last: Qi

first: Xiaocui

last: Luo

first: XiaoXiao

last: Hou

first: Yifeng

last: Wang

first: Ye

last: Li

first: Shangcheng

last: Yu


title: Non-Invasive Detection of Allele-Specific CRISPR-SaCas9-KKH Disruption of TOR1A DYT1 Allele in A Xenograft Mouse Model

city: Charlestown

first: Koen

last: Breyne

state: MA

address: Building 149 149 13th Street

institute: MGH

summary: DYT1 dystonia is a neurological movement disorder characterized by a dominant 3-base pair  deletion (dGAG) in the TOR1A gene. This study demonstrates a gene editing approach that  selectively targets the dGAG mutation in the TOR1A DYT1 allele while safeguarding the wild-type (WT) TOR1A allele. We optimized an adeno-associated virus (AAV) vector-compatible variant of  the Staphylococcus aureus Cas9 nuclease ortholog (SaCas9-KKH) in DYT1 patient-derived  human neuroprecursor cells (hNPCs). On-target editing of the TOR1A DYT1 allele was confirmed  at the genomic level from brain tissue in a xenograft mouse model. To avoid brain biopsy for  demonstrating TOR1A DYT1 editing, we developed a non-invasive monitoring method using  extracellular RNA (exRNA). TOR1A exRNA was retrieved from the EV secretions of hNPCs and  plasma samples, indicating whether the donor was a TOR1A DYT1 carrier. This technique  enabled us to assess AAV-mediated disruption of the TOR1A DYT1 allele in the brains of mice  using blood samples

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Next Generation CRISPR sequencing results analysed by CRISPResso2 software to detect allele edits or lack thereof

first: Katia

middle: E

last: Maalouf

first: Dawn

middle: M

last: Frederick

first: Nutan

last: Sharma

first: Edwina

last: Abou Haidar

first: Tianhe

last: Xiao

first: Justin

last: Seungkyu Han

first: Mohammed

last: Mahamdeh

first: Roy

middle: J

last: Soberman

first: David

last: Rufino- Ramos

first: Benjamin

middle: P

last: Kleinstiver

first: Hyder

last: Jinnah

first: Christine

last: Vaine

first: D Cristopher

last: Bragg


title: Next generation sequencing to identify KDM4A regulating signaling pathway in CRC

city: Guangzhou

first: Tanxing

last: Cai

state: Guangdong

address: 26 Erheng Road, Yuancun, Tianhe District

department: Department of General Surgery

institute: The Sixth Affiliated Hospital, Sun Yat-sen University

summary: We performed next generation sequencing between control CRC cells and CRC cells with KDM4A overexpression to identify KDM4A regulating signaling pathway

overall_design: RNA-seq profiling of control group and KDM4A overexpression group using HCT8 cells

first: Liang

last: Kang

first: Li

last: Xiong

first: Zhenxing

last: Liang


title: Single-cell landscape analysis reveals distinct regression trajectories and novel prognostic biomarkers in primary neuroblastoma

city: Chongqing

first: Shan

last: Wang

address: No. 136 Zhongshan 2nd Road, Yuzhong District, Chongqing

institute: Children’s Hospital of Chongqing Medical University

summary: Neuroblastoma (NB) is the most common extracranial malignant solid tumor in children, and its clinical manifestations and prognosis vary greatly. NB has a unique ability to spontaneously differentiate and regress, which provides potential directions for therapeutic interventions. To investigate the heterogeneity of tumors, we performed single-cell transcriptome sequencing on 19 tumor samples and 3 adrenal tissue samples, including 2 cases of familial neuroblastoma.

overall_design: we performed scRNA-seq on different stages of NB and peri-tumoral adrenal gland (AG) to investigate the developmental trajectory for spontaneous regression of NB and explore the potential value of the immune infiltrating cells. The study population consisted of 22 samples. Among them, 11 samples were NBL (high malignancy; groups 1-4), including three stage IV without MYCN amplification (group 1), two stage IV with MYCN amplification (group 2), three stage IVS (group 3), and three stage I/II (group 4); three samples were GN (benign; group5); and three samples were GNB (intermediate malignancy.group 6). Three AG samples comprising group 7 served as the control group. and F2 are two cases of family neuroblastoma


title: Dlk1 is a novel adrenocortical stem/progenitor cell marker that predicts malignancy in adrenocortical carcinoma [ssRNA-Seq]

city: London

first: James

last: Pittaway

address: Charterhouse Square

department: Centre for Endocrinology

country: United Kingdom

institute: Queen Mary University of London

summary: Adrenocortical carcinoma (ACC) is characterized by disruption of tissue development and homeostatic pathways. Here, using genetic fate mapping and murine ACC models, we have identified a population of adrenocortical stem cells that express Delta-like non-canonical Notch ligand 1 (DLK1). These cells are active during development, near dormant postnatally but are re-expressed in ACC. DLK1 expression is widespread in human ACC samples and independently influences recurrence-free survival. Spatial transcriptomic analysis has identified DLK1 positive ACC cells to have increased steroidogenic potential, a finding also observed in both human and murine ACC cell lines. Additionally, the cleavable DLK1 ectodomain is detectable in patients’ serum and effectively distinguishes ACC from other adrenal pathologies, offering potential diagnostic and follow-up utility for ACC patients. These data demonstrate a prognostic role for DLK1 in ACC, elucidate its hierarchical expression in homeostasis and cancer and suggest its utility as a biomarker in this malignancy.

overall_design: To investigate the transcriptomic differences of human ACC cell line H295R with different doses of DLK1 - over-expressing, knock-down or wild type.

middle: F

first: Katia

last: Mariniello

first: Leonardo

last: Guasti

first: Charlotte

last: Hall

last: Tourigny


title: Male-biased gene cyp17a2 enhances virus resistance in fish by promoting STING expression and degrading viral protein

city: Wuhan

first: Shengchi

last: Shi

address: Donghu South Road #7

institute: Institute of Hydrobiology, Chinese Academy of Sciences

overall_design: In the EPC cell line, overexpress the control plasmid and the cyp17a2 plasmid, then add SVCV infection after 24 hours, and perform RNA-seq on both groups.

first: Longfeng

last: Lu


title: CAR-macrophage therapy for HER2-overexpressing advanced solid tumors: a phase 1 trial

city: Philadelphia

first: Michael

last: Klichinsky

state: Pennsylvania

address: 3675 Market Street Ste. 401

institute: Carisma Therapeutics

summary: Biopsy samples from 4 sites were collected for RNA-Seq and single-cell RNA-Seq from patients with HER2 positive tumors enrolled in a Phase 1 clinical trial of anti-HER2 CAR-macrophage cell therapy. Biopsies of solid tumors were aimed to be collected at screening, 8 days after treatment, and 4 weeks after treatment. Patients in Group 1 received dose escalation of intravenous administration of up to 500 million cells, 1.5 billion cells, and 3 billion cells, on Day 1, 3, and 5, respectively. Patients in Group 2 received their full dose of cells on Day 1. This trial aimed to determine the feasibility of manufacturing CAR-macrophages, as well as assess the safety and tolerability of CAR-macrophage therapy, and determine the severity of potential adverse events.

overall_design:
TCR Sequencing was performed by Adaptive Biotechnologies on patient tumor biopsies at Screening, 8 days post CAR-Macrophage infusion, and 4 weeks post-infusion. TCR-Sequencing was performed on patient blood samples at Screening, 8 days, 15 days, and 4 weeks post-infusion.

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Adaptive Biotechnologies does not provide raw data.
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title: TMEM41B is an endoplasmic reticulum Ca2+ release channel maintaining naive T cell quiescence and responsiveness

first: Gang

last: Jin

address: Zhongguancun North street

department: Institute for Immunology

institute: Tsinghua University

summary: In mammalian cells, endoplasmic reticulum (ER) passively releases Ca2+ under steady state, but channels involved remain elusive. Here, we report that TMEM41B, an ER-resident membrane protein critical for autophagy, lipid metabolism, and viral infection, functions as an ER Ca2+ release channel. Biochemically, purified recombinant TMEM41B forms a concentration-dependent Ca2+ channel in single-channel electrophysiology assays. Cellularly, TMEM41B deficiency causes ER Ca2+ overload, while overexpression of TMEM41B depletes ER Ca2+. Immunologically, ER Ca2+ overload leads to upregulation of IL-2 and IL-7 receptors in naive T cells, which in turn increases basal signaling of JAK-STAT, AKT-mTOR, and MAPK pathways. This dysregulation drives TMEM41B-deficient naive T cells into a metabolically activated yet immunologically naive state. ER Ca2+ overload also downregulates CD5, lowering the activation threshold of TMEM41B-deficient T cells and leading to heightened T cell responses during infections. In summary, we identify TMEM41B as a concentration-dependent ER Ca2+ release channel, revealing an unexpected role of ER Ca2+ in naive T cell quiescence and responsiveness.

overall_design: To investigate how TMEM41B regulates naive T cell quiescence, we performed single-cell RNA sequencing (scRNA-seq) of naive T cells (TCR+CD62L+CD44-CD25-) isolated from  control and TMEM41B-deficient mice.

first: Yuying

last: Ma

first: Min

last: Peng


title: Single-cell RNA sequencing of the kidneys from adult old mouse

city: Seoul

first: Donghwan

last: YUN

state: Jongno-gu

address: 103 Daehakro

country: South Korea

institute: Seoul National University Hospital

summary: This study aims to investigate the impact of aging on the kidney through the analysis of single-cell RNA sequencing datasets.

overall_design: We prepared scRNA-seq datasets from whole kidneys of male WT mouse (n=1; 22 weeks old)

first: Chaelin

first: Yun

last: Donghwan

first: Han

last: Seung Seok


title: The Therapeutic Potential of mRNA-Mediated Type III Collagen Delivery for Photoaged Skin

city: City

first: Lianhao

last: Song

state: Guangzhou

address: Jinsui road

department: Zhongshan Ophthalmic Center

institute: Sun Yat-sen University

summary: Background: Skin aging can be classified into intrinsic and extrinsic types, with photoaging representing a significant form of extrinsic aging. Photoaging is caused by prolonged exposure to ultraviolet (UV) radiation, resulting in cellular damage, collagen degradation, and the development of wrinkles and pigmentation changes; severe cases can even lead to skin cancer. This condition affects individual aesthetics and health and imposes a broader societal burden, including increased healthcare costs and reduced quality of life. Addressing photoaging is critical, as effective treatments can enhance skin health, improve mental well-being, and alleviate the burden on healthcare systems, highlighting the importance of preventive and therapeutic strategies. Objective: This study aimed to develop a novel drug based on human type III collagen mRNA (hCOL3A1 mRNA) and to investigate its role in treating UVB-induced skin photoaging. Methods: We designed and synthesized the complete hCOL3A1 mRNA sequence and encapsulated it in lipid nanoparticles for skin delivery. The expression of the hCOL3A1 mRNA drug in human fibroblasts was examined by in vitro experiments, and its potential therapeutic effects on UVB-induced aging, oxidative stress, and inflammation were evaluated. In addition, we evaluated the efficacy of the hCOL3A1 mRNA drug on UVB-induced skin photoaging by an in vivo model. Results: It was found that the hCOL3A1 mRNA drug was effective in reducing UVB-induced skin photoaging, suggesting that the hCOL3A1 mRNA drug could be used as a therapeutic intervention for UVB-induced photoaging and also providing a novel approach to restore aging skin by increasing endogenous collagen production and reducing oxidative and inflammatory damage Conclusion: hCOL3A1 mRNA-LNP, as a novel protein therapeutic approach, overcomes the limitations of retinoic acid (RA) therapy, provides a simpler and more efficient therapeutic option, and is expected to be a new treatment for skin photoaging.

overall_design: RNA-seq profilling of balb/c-nude mice and UVB irradiation on their dorsal skin every other day for 8 weeks and tissue collection at day 28 ,The UV irradiation protocol started at an initial intensity equivalent to the minimal erythemal dose (MED) of 1 MED (60 mJ/cm²) for the first 2 weeks. The intensity was then gradually increased to 2 MED (120 mJ/cm²) in the third week, 3 MED (180 mJ/cm²) in the fourth week, and maintained at 4 MED (240 mJ/cm²) from the fifth to eighth week of the experiment.

first: Jiamin

last: Zhang

first: Shuqi

last: Chen

first: Zhi

last: Xie


title: Profiling the Cannabis-Sclerotinia pathosystem at the mRNA level using dual RNA sequencing

city: Winnipeg

first: Mark

last: Belmonte

state: MB

address: 50 Sifton Road

country: Canada

institute: University of Manitoba

summary: Known to infect more than 600 plant species worldwide, Sclerotinia sclerotiorum is a necrotrophic fungal pathogen, and the causative agent of white mold. With recent infection reports documented across North America, Cannabis sativa is known to be susceptible to Sclerotinia infection. Resulting from legal constraints associated with C. sativa, little is known about the Cannabis-Sclerotinia pathosystem, particularly in how the plant responds to pathogen attack at the cellular and molecular levels. Anatomical study revealed initial infection and degradation of the epidermis and cortical parenchyma, followed by widespread infection of the vascular phloem. Dual RNA sequencing provided a detailed transcriptomic profile of this pathosystem directly at the site of infection. Differential gene expression analysis revealed large-scale transcriptional shifts resulting from rapid infection. We identified the upregulation of 97 genes at 1 day post inoculation (dpi) and 6733 genes 5 dpi in C. sativa, while 3186 genes were identified in S. sclerotiorum 7 dpi. Gene ontology term enrichment identified processes associated with plant defense and signal transduction cascades during C. sativa infection while processes associated with redox control and sugar catabolism were enriched in S. sclerotiorum. Taken together, this study revealed transcriptional reprogramming in both the host plant and fungal pathogen associated with degradation of host cortical and vascular phloem tissues.

sample_organism:
Cannabis sativa
Sclerotinia sclerotiorum

overall_design: Cannabis sativa (cultivar 'Kona') inflorescences were inoculated with Sclerotinia sclerotiorum. Infected and uninfected tissues were collected at 1 day post inoculation (dpi), 3 dpi, 5 dpi and 7 dpi.

first: Natalie

middle: L

last: Cale

first: Rylee

last: Swiderek

first: Philip

last: Walker

first: Sean

last: Robertson

first: Dylan

last: Ziegler


title: Single-cell transcriptomic analysis reveals distinct cellular responses of cortical progenitors and neurons in human brain organoid modeling fetal hypoxic brain injury

city: shanghai

first: quanlong

last: jiang

institute: PICB

summary: Fetal hypoxia, characterized by reduced oxygen levels, adversely impacts fetal development and contributes to 23% of neonatal deaths worldwide. Notably, the developing brain, particularly in its immature stage, is highly susceptible to hypoxic insult, which increases the risk of neural defects, brain damage, and long-term cognitive impairments in offspring. Nevertheless, the specific cell types affected and the underlying molecular mechanisms that mediate hypoxic brain injury remain incompletely understood. In this study, we established a three-dimensional human brain organoid modeling fetal hypoxic brain injury. Utilizing single-cell transcriptomic analysis, we observed a global hypoxic response in cortical progenitors and neurons, characterized by enhanced HIF-1 signaling and increased expression of hypoxia-responsive genes following hypoxic insult. Under hypoxic conditions, glycolysis, reactive oxygen species pathway and oxidative phosphorylation were simultaneously upregulated. Furthermore, hypoxia led to a decrease in cell division and a suppression of neurotrophin signaling pathways. Regarding developmental stage, cortical progenitors exhibited minimal sensitivity to hypoxia, with ventral radial glia, outer radial glia, and intermediate progenitors (IPs) being largely unaffected. In contrast, glia progenitors displayed a slight tendency towards premature maturation. Conversely, hypoxia had a detrimental effect on the maturation of developing neurons, including glutamatergic and GABAergic neurons. Significantly, we identified two distinct populations of GABAergic neurons with differing characteristics. GABAergic neuron type 1 exhibited lower expression levels of GAD1 and GAD2 and greater resistance to hypoxia compared to GABAergic neuron type 2. Notably, GABAergic neuron type 2 was the most susceptible to hypoxia among all cell types, showing increased ferroptosis, disrupted axon development, and impaired synapse function, potentially due to decreased Hedgehog signaling. Additionally, specific responses to hypoxia in developing neurons included selective reduction in fatty acid synthesis in outer radial glia cells, glutamatergic neurons and GABAergic neuron type 1, selective downregulation of mTORC1 signaling in IPs, selective elevation of unfolded protein response in GABAergic neuron type 1, and selective activation of TNFα-signaling via NFκB in IPs, glutamatergic neurons and GABAergic neuron type 2. In summary, our study provides a comprehensive understanding of the distinct cellular responses among cortical progenitors and neurons in fetal hypoxic brain injury, providing potential therapeutic targets for this condition.

overall_design: scRNA-seq data of hypoxic cortical organoids (hCOs) exposed to hypoxic and normoxic conditions for 48 hours.


title: Functional interplay between RND efflux pumps and GacS in Pseudomonas aeruginosa

city: Norman

first: Justyna

middle: Weronika

last: Adamiak

state: OK

address: 101 Stephenson Parkway

department: Chemistry and Biochemistry

institute: University of Oklahoma

summary: Pseudomonas aeruginosa is an opportunistic human pathogen, which is ubiquitous in the environment. The environmental versatility of P. aeruginosa is often attributed to a large arsenal of active efflux pumps, low permeability of the outer membrane and extensive regulatory networks. In this study, we analyzed putative functional interplay between polyspecific drug efflux pumps of Resistance-Nodulation-Division (RND) superfamily of proteins and GacSA two-component regulatory system. RND transporters are the major contributors to antibiotic resistance and survival under various environmental stresses, whereas GacSA is a global regulator and governs critical lifestyles during human colonization. We found that the inactivation of either RND pumps or gacS induces broad, partially overlapping responses in lifestyle, virulence and metabolic programs that are more pronounced during exponential than stationary phase. GacSA and RND efflux pumps induced opposite responses in gene expression of P. aeruginosa, whereas the overexpression of GacS was additive with the deletion of efflux pumps in gene expression and under growth conditions typically associated with human infections such as elevated temperature and iron deprivation. We conclude that RND efflux and GacSA networks partially overlap with each other.

overall_design: RNAseq analysis of the exponential and stationary Pseudomonas aeruginosa PAO1 and its derivatives lacking 6 RND efflux pumps (PΔ6fs; ΔmexAB-oprM ΔmexCD-oprJ ΔmexEF-oprN ΔmexJKL ΔmexXY ΔtriABC) and overexpressing gacS (Δ6gacSfs::LAC-gacS). Samples were analyzed in duplicate.

first: Charles

last: Bergen

first: Valentin

middle: V

last: Rybenkov

first: Helen

middle: I

last: Zgurskaya


title: Csf1+ AD-MSCs promote stroke repair by activating the resident microglia

city: tianjin

first: huizhen

last: wang

address: tingjiang westroad

institute: Tasly Pharmaceutical Group Co., Ltd.

summary: We have employed a single cell sequencing approach using 10x Genomics scRNAseg to study the mechanism of AD-MSCs for treating the injured brain tissue. This study establishes a rat model of hemorrhagic stroke, followed by intravenous infusion of allogeneic AD-MSCs 24 hours post-model induction, in order to observe the therapeutic effect and analyze the underlying mechanism of action. Based on single-cell transcriptomic sequencing technology, we demonstrated that the re-infusion of AD-MSCs 24 hours after stroke mainly promotes the repair of injured brain tissue by activating brain tissue resident microglia and inhibiting bone marine-derived monocytes. Importantly, this role depends on the cell subsets of AD-MSCs that are highly expressing Csf1+. This study offers novel insights into the mechanisms underlying MSC-based stroke treatment and supports the potential for stable and efficacious MSC therapies.

overall_design: In this study, the collagenase-induced intracerebral hemorrhage (ICH) rat model was used for intravenous infusion of AD-MSCs 24 hours after modeling. At 72 hours after cell transfusion, the damaged brain tissue was sampled and sequenced by single cell transrecording.

first: Jiguang

last: Hou

first: Shuang

last: Luo

first: Jia

last: Liu

first: Xiao

last: Zuo

first: Fei

last: Ma

first: Huizhen

last: Wang

first: Pengfei

last: Han


title: Recognition of ubiquitinated histone H2A constitutes a treatment target for RSF1-driven ovarian cancer

city: RICHMOND

first: Hengbin

state: VA

address: 401 college st

institute: Virginia Commonwealth University

summary: RNA-seq results revealed that overexpression of RSF1, but not RSF1ΔUAB, induced a gene expression program enriched for cancer stem cell markers. Conversely, overexpression of a UAB domain decoy peptide reduced cell proliferation and blocked the cancer stem cell gene expression programs in OVCAR3 cells. Together, these studies reveal the critical role of recognizing H2Aub in RSF1-driven ovarian cancer and demonstrate that the interaction between RSF1 and H2Aub constitutes a druggable target for treating ovarian cancers with high levels of RSF1.

overall_design: Expression profiling by high throughput sequencing

first: Chunxu

last: Chen

first: Christiane

last: Morecock

first: Mikail

last: Bala


title: Agrimoniin synergistically works with erlotinib to inhibit the proliferation of pancreatic cancer by remodeling the tumor microenvironment and suppressing lactic acid expression

city: Nanjing

first: Xinyi

last: Xia

state: Jiangsu

address: 138 XianLin Road, Nanjing, JiangSu, China

institute: Nanjing University of Chinese Medicine

summary: Erlotinib, as an EGFR small molecule inhibitor for pancreatic cancer, its efficacy as monotherapy is limited. Combination of erlotinib and gemcitabine has shown improvement in multiple survival indicators. However, drug resistance remains a challenge. Agrimoniin, the main active ingredient in Agrimonia pilosa, has anticancer potential. Whether it can serve as a synergistic or resistance-reversing drug for erlotinib require further research. In this study, we performed subcutaneous xenograft tumor experiments, CCK8, EdU assays, flow cytometry, HE staining, immunohistochemical and western blot to explore the synergistic effects of agrimoniin and erlotinib in inhibiting pancreatic cancer. Single-cell RNA sequencing was used to explore the relationship between their synergistic effects and tumor microenvironment. Through western blot and colorimetric assays for lactic acid, we studied the connection between their synergistic effects and aerobic glycolysis as well as lactic acid expression. By employing signal pathway inhibitors and agonists, we studied the positive feedback regulatory mechanism between relevant signal pathways and lactic acid expression. We found that agrimoniin and erlotinib synergistically inhibits the proliferation and promotes the apoptosis of pancreatic cancer cells. Together, they effectively suppress signal interaction among tumor cells, stromal cells and macrophages within subcutaneous tumors. They synergistically inhibit aerobic glycolysis and lactic acid expression in pancreatic cancer. Notably, we uncover the vicious cycle between the PI3K/AKT//HIF-1α/HK2, MEK/ERK/HIF-1α/HK2 signaling pathways and lactic acid expression. Combination of agrimoniin and erlotinib remodels tumor microenvironment, reprograms aerobic glycolysis, disrupts the positive feedback loop between lactic acid and two signaling pathways, thus synergistically inhibits the proliferation of pancreatic cancer cells.

overall_design: We transplanted PANC1 cells into the subcutaneous skin of nude mice and treated them with the carrier, cyranin, agrimoniin, agrimoniin + erlotinib, respectively, and detect the effects of different drugs on tumor

first: Xiongfei

last: Zhang

first: Yan

first: Huanhuan

last: Zhao

first: Huiying

last: Xu

first: Yutong

first: Shaojun

first: Qinlong

last: Dai

first: Ling

last: Gu

first: Yiwen

first: Xinye

last: Dong

first: Beili

last: Xi

first: Li

last: Tao

first: Yang

last: Hua

first: Zhihan

first: Xinyue

last: Tian

first: Lei

last: Li

first: Yuyan

last: Ling

first: TingTing

first: Meijuan

first: Jingjing

first: Lifeng

last: Zhu


title: Altered polyadenylation site usage in SERPINA1 3’UTR in response to cellular stress affects A1AT protein expression

city: Greenwood

first: Lela

last: Lackey

state: SC

address: 114 Gregor Mendel Circle

department: Genetics and Biochemistry

institute: Clemson University

summary: Alternative polyadenylation results in different 3’ isoforms of messenger RNA (mRNA) transcripts. Alternative polyadenylation in the 3’ untranslated region (3’UTR) can alter RNA localization, stability and translational efficiency. The SERPINA1 mRNA has two distinct 3’ UTR isoforms, both of which express the protease inhibitor α-1-antitrypsin (A1AT). A1AT is an acute phase protein that is expressed and secreted from liver hepatocytes and upregulated during inflammation. Low levels of A1AT in the lung contributes to chronic obstructive pulmonary disease, while misfolding of A1AT in the liver contributes to liver cirrhosis. We analyzed the dynamics of alternative polyadenylation during cellular stress by treating the liver cell line HepG2 with the cytokine interleukin 6 (IL-6), ethanol or peroxide. SERPINA1 is transcriptionally upregulated after IL-6 treatment and has altered polyadenylation, resulting in an increase in long 3’UTR isoforms. We find that the long 3’UTR represses endogenous A1AT protein expression even with high levels of SERPINA1 mRNA. SERPINA1 expression and 3’ end processing are not affected by ethanol or peroxide. IL-6-induced changes in transcriptome-wide transcriptional regulation suggest changes to the endoplasmic reticulum and in secretory protein processing. Our data suggest that inflammation influences polyA site choice for SERPINA1 transcripts, resulting in reduced A1AT protein expression.

overall_design: We treated HepG2 cells with interleukin 6 (IL-6), ethanol or peroxide and performed polyA selection and 3' end specific library preparation (Lexogen QuantSeq Rev). We generated CRISPR mutant lines with a change in the first polyadenylation site of the SERPINA1 gene (APA-mut HepG2 cell line). We performed the same treatments on the APA-mut HepG2 cells.

first: FNU

last: Jiamutai

first: Abigail

last: Hatfield

first: Austin

last: Herbert

first: Debarati

last: Majumdar

first: Vijay

last: Shankar


title: Herbal-based Xuebijing injection ameliorated vascular endothelial dysfunction in sepsis-associated lung injury via inhibiting ACLY/MYB/RIG-I axis

city: 青岛市

first: Jiayin

last: Cui

state: 山东省

address: 市南区鱼山路5号

department: 医药学院

institute: 中国海洋大学

summary: Excessive endothelial pro-inflammatory response is an early hallmark in sepsis-induced acute lung injury (ALI). Xuebijing (XBJ) as a Chinese traditional medicine is widely used for treating sepsis in the clinic. However, the molecular mechanism of the beneficial effectiveness of XBJ remains unclear. Our present study showed that XBJ inhibited phosphorylation-dependent activation of ATP citrate lyase (ACLY), thereby suppressing the acetylation-dependent nuclear translocation of transcription factor MYB. Thus, as the downstream target gene of MYB, the expression of retinoic acid inducible gene I (RIG-I) was downregulated, leading to the inhibition of NF-κB signaling and EC pro-inflammatory and coagulation activation, which further alleviated sepsis-associated ALI. Moreover, XBJ compounds Quercetin, Ferulic Acid, Kaempferol and Paeoniflorin all showed inhibitory effects on the activation of the downstream MYB/RIG-I signaling by binding to ACLY protein. Our study revealed the novel regulatory mechanism of XBJ in sepsis-induced EC dysfunction and ALI via ACLY/MYB/RIG-I axis, providing theoretical basis for the clinical application of XBJ and shedding insight into novel therapeutic targets for treating sepsis.

overall_design:
In order to explore the protective mechanism of Xuebijing on endothelial cells in sepsis, we set up CT group, LPS group and XBJ+LPS group to sequence RNA.
Three biological replicates were set in each group, with a total of nine samples.
The cells used were HUVEC. The CT group did not deal with it. In LPS group, endothelial cells were stimulated with 1μg/ml LPS for four hours. In XBJ+LPS group, HUVEC was pretreated with 50μl/ml XBJ for 30 minutes, and then stimulated with LPS for 4 hours.

first: Ranran

first: Mei

last: Meng

first: Yuchao


title: Gene expression profile at single cell level of intestinal cells from drosophila posterior midgut 2

city: Dallas

first: Matt

last: Sieber

state: Texas

address: 5323 Harry Hines blvd, MC 9040 ND12.214

institute: UT Southwestern Medical Center

summary: We performed single cell RNA sequencing to investigate the gene expression changes upon maternal stress in progeny drosophila posterior midgut.

overall_design: We induced maternal metabolic stress by expressing InR-DN in the female germline. These InR-DN expressing females were mated it orgeon R males and allowed to grow to adult hood. We also rescues the effects of the InR-DN transgene by feeding mothers methionine. Posterior midguts of the progeny flies (maternally stressed,control and  rescued/methionine fed) progeny drosophila were dissected, diassociated, and put into a 10XGeneomics scRNA-Seq pipeline with assistance form the UTSW sequencing facility.

first: Helin

last: Hocaoglu

first: Farhan

last: Abu

first: Matthew

first: Benjamin

last: Ohlstein


title: Gene expression profile at single cell level of intestinal cells from drosophila posterior midgut 1

overall_design: We induced maternal metabolic stress by expressing InR-DN in the female germline. These InR-DN expressing females were mated it orgeon R males and allowed to grow to adult hood. We also rescued the effects of the InR-DN transgene by feeding mothers methionine. Posterior midguts of the progeny flies (maternally stressed,control and  rescued/methionine fed) progeny drosophila were dissected, diassociated, and put into a 10XGeneomics scRNA-Seq pipeline with assistance form the UTSW sequencing facility.


title: Single cell RNA-seq reveals GINIP-Expressing neurons as the main targets of focused ultrasoud.

city: Marseille

first: Thibaud

last: Parpaite

address: 51 Bd Pierre Dramard

department: Neural basis of the Somatosensory System

country: France

institute: Aix-Marseille University

summary: Focused ultrasound (US) is a recently-explored method for neurostimulation. One potential advantage of focused US compared to electrical stimulation is the ability to selectively target restricted subpopulations of neurons. This feature makes US neurostimulation a promising alternative tool for the design of future neural prostheses: for example, to restore hearing in deaf people, because contemporary cochlear implants – which work via electrical stimulation - mainly suffer from a lack of spatial selectivity. Focused US was recently shown to directly stimulate neurons in several in vitro and in vivo studies; however the mechanisms of this effect still remain poorly understood. The aim of this interdisciplinary PhD project is to understand the physical and molecular causes of US-induced neurostimulation. The molecular mechanisms will be identified by combining US, calcium imaging and single-cell RNA sequencing on cultured dorsal root ganglia (DRG) neurons. The physical mechanisms (thermal and mechanical effects of US action) will be measured using temperature monitoring and laser vibrometry and confronted to numerical simulations of the in vitro set-up.

overall_design: single-cell RNA-seq of ultrasound-reactive dorsal root ganglia sensory neurons after GCaMP calcium recorgings.

first: Aziz

last: Moqrich

first: Parpaite

last: Thibaud

first: Andrew

last: Saurin


title: Reawakening inflammation in the chronically injured spinal cord using lipopolysaccharide induces diverse microglial states

city: Edmonton

first: Jason

last: Plemel

state: AB

institute: University of Alberta

summary: Rehabilitative training is an effective method to promote recovery following spinal cord injury (SCI), with lower training efficacy observed in the chronic stage. The increased training efficacy during the subacute period is associated with an adaptive state induced by the SCI. A potential link is SCI-induced inflammation, which is elevated in the subacute period, and as injection of lipopolysaccharide (LPS) alongside training improves recovery in chronic SCI, suggesting LPS could reopen a window of plasticity late after injury. Microglia may play a role in LPS-mediated plasticity as they react to LPS and are implicated in facilitating recovery following SCI. However, it is unknown how microglia change in response to LPS following SCI to promote neuroplasticity. Here we used single-cell RNA sequencing to examine microglial responses in subacute and chronic SCI with and without an LPS injection. We show that subacute SCI is characterized by a disease-associated microglial (DAM) signature, while chronic SCI is highly heterogeneous, with both injury-induced and homeostatic states. With LPS injection, microglia shifted away from the homeostatic signature to a primed, translation-associated state and increased DAM in degenerated tracts caudal to the injury. Our results contribute to an understanding of how microglia and LPS-induced neuroinflammation contribute to plasticity following SCI.

overall_design: Female rats received a dorsolateral quadrant hemitransection injury, and then were perfused with HBSS(-/-). A 2 cm spinal cord section around the lesion site was extracted from rats at 8 days following injury (acute), 60 days following injury (chronic with saline), 60 days following injury and 3 days following an LPS injection (chronic with LPS). Spinal cords were manually cold disocciated using a dounce homogenizer into a single cell suspension and then fluorescent-activated cell sorting (FACS) was completed for live, nucleated and Cd11b+ cells.

first: Rebecca

middle: K

last: John

first: Sadie

last: Vogel

first: Sameera

last: Zia

first: Kelly

last: Lee

first: Antoinette

middle: T

last: Nguyen

first: Abel

last: Torres-Espin

first: Keith

last: Fenrich

first: Carmen

last: Ng

first: Emma

last: Schmidt

first: Romana

last: Vavrek

first: Pamela

last: Raposo

first: Keira

last: Smith

first: Karim

last: Fouad


title: METTL5 deficiency induces lung cancer regression via mitochondrial translation reduction

city: nanjing

first: Zhang

last: Jun

state: jiangsu

address: longmiandadao

institute: China Pharmaceutical University

summary: Cellular protein synthesis occurs through two types of translation: cytoplasmic and mitochondrial. These two types of translation operate independently but in coordination to regulate cell survival and death. Methyltransferase-like 5 (METTL5) has been found to play a role in directing the m6A modification of rRNA, which in turn controls cytoplasmic translation and impacts the decisions related to cell fate. In this study, we employed Ribo-seq to explore the effects of METTL5 deficiency on both cytoplasmic and mitochondrial translation.

overall_design: To investigate the potential role of METTL5 in the regulation of non-small cell lung cancer, a stable METTL5 knockout cell line was initially constructed using classical A549 and H1299 NSCLC cells using a CRISPR Cas9 sgRNA-based gene interference method.

first: Qiujie

last: Li

first: Jun

last: Zhang


title: LTO1 Complex as a Therapeutic Target for Enhancing Immunotherapy via Nonsense Mediated RNA Decay-Related MHC-I Upregulation in Tumor Cell

city: Guangzhou

first: Xiaoling

last: Wang

address: No. 195, West Dongfeng Road

institute: Guangzhou Medical University

summary: Immune checkpoint inhibitors benefit only a subset of patients, necessitating the development of new sensitizing agents strategies to improve tumor response and efficacy. Major histocompatibility complex class I (MHC-I), vital for tumor antigen presentation and cytotoxic T cell activation, has been an important target of immunotherapy. Nonsense-mediated RNA decay (NMD) is a highly conserved surveillance system that targets mRNAs with premature termination codons (PTCs) and monitors the quantity of about 10% of unmutated mammalian mRNAs. Tumor cells with aneuploidy heavily rely on NMD to compensate the imbalanced DNA levels and lower their antigenicity for immune evasion, making them sensitive to NMD inhibitors. Using an unbiased genome-wide analysis, we unexpectedly identified LTO1/YAE1 complex as novel NMD factors. LTO1/YAE1 complex and its downstream target for iron-sulfur cluster transfer, ABCE1, are widely overexpressed and negatively associated with immune cells infiltration in multiple cancer types. The RNA-sequencing results from these three genes led to the discovery that In tumor cells,compromised NMD is associated with MHC-I upregulation via increased expression of NLRC5, IRF1 and NF-κB in tumor cells. Importantly, we found that low doses of iron chelators, NMD inhibitors in clinical use in treatment of iron overload mimic , enhance MHC-I expression and improvingthese effects and improves immune checkpoint blockade (ICB) efficacy. Our findings suggest that LTO1/YAE1/ABCE1 could be a therapeutic target to enhance cancer immunotherapy by upregulating MHC-I and promoting immune infiltration.  Our workfindings links NMD toand iron homeostasis andto the tumor immune microenvironment. Our findings suggest that , highlighting that targeting the NMD could be a therapeutic strategy promote immune infiltration, potentially enhancing cancer immunotherapy.

overall_design: To explore the function of YAE1 and ABCE1,  A375 cells were transduced with lentivirus expressing CRISPR/Cas9 and indicated sgRNA, with irrelevant gRNAs targeting LacZ or Luciferase as negative controls. Seven days after selection with 10 µg/mL puromycin, RNA was isolated and its integrity is evaluated using Agilent 4200 TapeStation. 1 μg RNA per sample was used as input for the library preparation following standard protocols. Library preparations were sequenced on a NovaSeq 6000 platform (Chi-Biotech, Guangzhou, China) and 150 bp stand-specific paired-end reads were generated.

first: Wang

last: Xiaoling

first: Leng

last: Qibin

first: Qiu


title: Diabetes accelerates atherosclerosis through lactate induced GPR132-Src activation

city: Xiamen

first: Yijun

last: Lin

address: NO.2999, Jinshan Road, Huli District, Xiamen, China

institute: Xiamen Cardiovascular Hospital of Xiamen University, School of Medicine, Xiamen University

summary: Diabetes mellitus is one of the major independent risk factors for the development of ASCVD. Several studies have reported that diabetes increases inflammatary signaling in different cells, and the cytokines may influence plaque formation, but the signal pathway in circulating cells that links glucose metabolism and lipid metabolism remains unclear.We isolated peritoneal macrophages of diabetic and non-diabetic APOE-/- mice to investigate the diabetes influence on macrophages.

overall_design: APOE-/- mice were rendered diabetes from 8 weeks of age by intraperitoneal injections of streptozotocin (STZ, 40 mg/kg) or citrate buffer (nondiabetic) on 5 consecutive days, then the mice were fed with western diet for 8 weeks. In the 8th week, mice were i.p. with 1.5mL of 3% thioglycollate, and 3 days later, the peritoneal exudate was isolated for RNA isolation and sequencing.


title: IRF7 promotes systemic autoimmunity by transcriptionally regulating B cell differentiation through germinal center and plasma cell pathways [ChIP-seq]

city: Philadelphia

first: Michael

last: Gonzalez

address: 3535 Market St

department: Translational Medicine and Human Genetics

institute: University of Pennsylvania

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Little is known about how interferon regulatory factor 7 (IRF7) promotes systemic autoimmunity. Here performed scRNAseq on B cells purified from the spleens of 3 month old FcgRIIB-/-IRF7-/- and FcgRIIB-/- controls. We identified a role for IRF7 in promoting the differentiation of B cells through the GC and PC pathways during an autoimmune response. To determine which genes IRF7 specifically controlled for this response, we purified class-switched activated B cells from FcgRIIB-/- mice and performed IRF7 ChIPseq. We identified that IRF7 transcirptionally regulated genes assocaited with B cell protein translation and metabolism. In summary, we have identified a B cell intrinsic role for IRF7 in driving the transcriptoinal profile required for GC and PC differentiation in SLE-like autoimmunity.

overall_design: 4 single cell RNAseq libaries with paired gene expression and 2 IRF7 ChiPseq libraries

first: Adam

last: Fike

first: Ziaur

last: Rahman


title: AGAL misprocessing-induced ER stress and the unfolded protein response: lysosomal storage-independent mechanism of Fabry disease pathogenesis?

city: Prague

first: Viktor

last: Stranecky

state: Czech Republic

address: Ke Karlovu 2

department: 1st Faculty of Medicine

country: Czech Republic

institute: Charles University

summary: Classic Fabry disease (FD) is caused by GLA mutations that result in enzymatic deficiency of alpha-galactosidase A (AGAL), lysosomal storage of globotriaosylceramide, and a resulting multisystemic disease.  In non-classic later-onset FD, patients have some preserved AGAL activity and a milder disease course, though female carriers may also be affected.  While FD pathogenesis has been mostly attributed to catalytic deficiency of mutated AGAL, lysosomal storage and impairment of lysosomal functions, other pathogenic factors may be important, especially in non-classic later-onset FD.  Clinical findings in affected males revealed a milder clinical course with ~15% residual AGAL activity and borderline plasma lyso-Gb3Cer levels. Kidney biopsies did not show lysosomal storage. Laboratory investigations documented intracellular retention of mutated AGAL with resulting endoplasmic reticulum (ER) stress and the unfolded protein response (UPR), which were alleviated with BRD4780, a small molecule clearing misfolded proteins from the early secretory compartment. We observed similar findings of ER stress and UPR with several other classic and non-classic FD missense AGAL variants. We identified defective proteostasis of mutated AGAL resulting in chronic ER stress and UPR of AGAL expressing cells (hereafter referred to as AGALopathy) as an important contributor to FD pathogenesis. These findings provide insight into non-classic later-onset FD and may better explain clinical manifestations with implications for pathogenesis, clinical characterization and treatment of all FD forms.

overall_design:
We characterized the genetic, clinical, biochemical, molecular, cellular and organ pathology correlates of the p.L394P AGAL variant that was identified in six individuals with end-stage kidney disease by the Czech national screening program for FD and by further screening in an additional 24 family members. Among other material we used HEK cells stably producing either wt-AGAL_FLAG or mut-AGAL_FLAG tagged.
Preparation of cell lines stably expressing AGAL_FLAG in HEK cells: HEK 293 cells were maintained in Dulbecco's Modified Eagle Medium : Nutrient Mixture F-12 (DMEM/F-12) high glucose medium supplemented with 10% (vol/vol) fetal calf serum (Gibco), 100 U/ml penicillin G/streptomycin (Sigma, Prague, Czech Republic). HEK293 cells were transfected with 2.5 µg of plasmid DNA using  Lipofectamine 3000TM (Invitrogen, Paisley, UK). Three days post transfection, cells were trypsinized, diluted and cultured in selective medium containing 0.5 mg/ml G418 (Invitrogen-Gibco, Paisley, UK). For each selected clone, the presence and correct sequence of GLA_FLAG was confirmed by Sanger sequencing; the amount of GLA_FLAG transcript was determined by real-time PCR, and expression levels of AGAL_FLAG tagged proteins were assesed by Western blot with mouse monoclonal anti-FLAG antibody (F1804, SIGMA-Aldrich, Prague, Czech Republic). Clones demonstrating similar GLA_FLAG transcript and AGAL_FLAG protein amounts were selected for further analyses.

first: Martina

last: Živná

first: Veronika

last: Barešová

first: Dita

last: Mušálková

first: Lenka

last: Steiner-Mrázová


title: Effect of maternal metabolic stress on drosophila progeny intestines

first: Matt

last: Sieber

summary: In this experiment we exmained the impact of maternal metabolic stress on progeny gene expression in the drosophila intestine.

overall_design: To investigate the effect of maternal metabolic stress on progeny intestines, we crossed control females and females expressing InR-DN in the germline to Oregon R males. We isolated intestines from the adult progeny and preppeed RNA using a Tripure based RNA extraction procotol. the mRNA was then sequenced with assistance from the UTSW sequencing core.

first: Helin

last: Hocaoglu

first: Farhan

last: Abu

first: Matthew

first: Benjamin

last: Ohlstein


title: IRF7 promotes systemic autoimmunity by transcriptionally regulating B cell differentiation through germinal center and plasma cell pathways [scRNA-seq]


title: Nicotinamide mononucleotide mitigates neuroinflammation by enhancing GPX4-mediated ferroptosis defense in microglia

city: Yinchuan

first: Yin

address: 1160 Shengli St

institute: Ningxia Medical University

summary: Numerous neurological diseases involve neuroinflammation, a process in which immune cells, particularly microglia, contribute to neuronal death. Ferroptosis, a recently identified form of regulated cell death, is implicated in various diseases characterized by neuronal injury. Nicotinamide mononucleotide (NMN), a potent NAD+ precursor supplement, has been found to inhibit neuroinflammation and ferroptosis. However, the mechanisms of NMN in both ferroptosis and neuroinflammation remains unclear. The present study aimed to investigate the impact of NMN on neuroinflammation and the susceptibility of microglia to ferroptosis. Ferroptosis markers in microglia exposed to lipopolysaccharide (LPS) were analyzed using CCK8, flow cytometry, ELISA, and RT-qPCR. The effects of NMN on LPS-induced ferroptosis in microglia were evaluated through flow cytometry, western blotting, and immunofluorescence staining. RT-qPCR analysis assessed the inflammatory cytokine production of microglia subjected to ferrostatin-1-regulated ferroptosis. RNA sequencing elucidated the underlying mechanisms of NMN-associated microglia ferroptosis under LPS induction. In BV2 microglia, an inhibitor of Glutathione Peroxidase 4(GPX4), RSL3, was employed to suppress GPX4 expression. Intracerebroventricular injection of LPS was performed to evaluate neuroinflammation and microglia activation in vivo. LPS treatment resulted in decreased cell viability, accompanied by upregulation of ferroptosis markers SLC7A11 and GPX4, and elevated levels of malondialdehyde (MDA), 4-hydroxynonenal (4-HNE), and total iron in a dose-dependent manner. NMN effectively rescued LPS-induced ferroptosis and improved cell viability in microglia. Co-administration of NMN and ferrostatin-1 significantly reduced proinflammatory cytokine production in microglia following the introduction of LPS stimuli. Mechanistically, NMN facilitated glutathione (GSH) production, and promoted resistance to lipid peroxidation in a GPX4-dependent manner, repressing cytokine transcription and protecting cells from ferroptosis. RNA sequencing elucidated the underlying mechanism of NMN-associated microglia ferroptosis under LPS induction. Furthermore, simultaneous injection of NMN ameliorated LPS-induced ferroptosis and neuroinflammation in mouse brains. The data from the present study indicated that NMN enhances GPX4-mediated ferroptosis defense against LPS-induced ferroptosis in microglia by recruiting GSH, thereby inhibiting neuroinflammation. Therefore, therapeutic approaches targeting ferroptosis in diseases using NMN should consider both its anti-ferroptosis and anti-inflammatory effects to achieve optimal outcomes, presenting promising strategies for treating neuroinflammation-related diseases or disorders.

overall_design: To unravel the molecular mechanism underlying NMN-mediated rescue of LPS-induced ferroptosis in BV2 cells, RNA sequencing was conducted to identify differentially expressed genes between the LPS and LPS + NMN groups

first: Ruiqiong

last: Su

first: Xiaoyue

last: Pan

first: Qiuyuan

last: Chen


title: HSF1 promotes the repopulation of post-senescent fatty hepatocellular carcinoma cell via modulating HSP90α/P53 complex stability.

first: Wangl

middle: si

last: kai

address: fenglin road

institute: zhongshan hospital

summary: HSP90 is a molecular chaperone extensively studied in the context of tumor development. Although burgeoning studies demonstrate that the activity and modification status of HSP90 are involved in maintaining and evolving the malignant phenotype of tumor, both during radiotherapy and chemotherapy-induced stress responses, the mechanistic intricacies tying HSP90-regulated cell senescence, serving as a pro-survival strategy, to tumor progression remain elusive. In this paper, we applied RNA-seq of Huh7 cells with HSP90 inhibitor 17-AAG treatment and its parent counterparts to determine the effect of HSP90 on HCC cell and its transcriptome differences. Our results showed that HSF1-HSP90α-P53 axis facilitated switching lipid metabolism from catabolism to anabolism during repopulation post-senescence, thereby maintaining HCC cell survival, which offers a promising therapeutic strategy for tumor prevention.

overall_design: Cells were sorted as three groups based on 17-AAg treatment, including 17-AAG treatment group and untreated group (Control group). Three replicates were conducted to each group.

first: Kun

last: Guo

first: Sikai

last: Wang

first: Hongxia

last: Cheng


title: Nitidine chloride inhibits mTORC1 signaling through ATF4-mediated Sestrin2 induction and targets IGF2R for autophagic degradation

city: guangdong

first: chen

middle: feng

last: zhi

address: 深圳市光明区新湖街道中山大学医学园南721

institute: chen

summary: Nitidine chloride (NC), a natural phytochemical alkaloid derived from Zanthoxylum nitidum (Roxb.) DC, exhibits multiple bioactivities in cancer, infection disease, and other therapeutic areas. However, the primary targets of NC and the mechanism of action (MOA) have not been defined explicitly. Here, we report that NC inhibit mTORC1 activity by targeting amino acid sensing signaling through activating transcription factor 4 (ATF4)-mediated Sestrin2 induction, and it efficiently induces autophagy in cells and tumor xenograft. NC displays potent cytotoxicity against various cancer cells and inhibits xenograft tumor growth of B16 melanoma. Furthermore, we identified insulin-like growth factor 2 receptor (IGF2R) as a direct target of NC by drug affinity responsive target stability (DARTS) technique. NC may functional as a IGF2R antagonist to inhibit growth factor-mediated mTORC1 activation. This study suggested that NC targets mTORC1 signaling through nutrients sensing and growth factor receptor binding, providing mechanistic insights into the MOA of NC.

overall_design: HEK-293T cells were treated with DMSO and 10μM nitidine chloride for 12 hours, with 3 replicates set for each treatment. RNA was extracted and subjected to high-throughput sequencing.


title: Super-fast generation of all-X sperm grass carp by female germline stem cell transplantation

city: WuHan

first: Yi-Xuan

last: Tu

state: HuBei

address: 1 Shizishan Rd

department: College of Life Science and Technology

institute: Huazhong agricultural university

summary: Grass carp (Ctenopharyngodon idellus), the world’s largest aquaculture fish species, exhibits superior growth in females compared to males. However, the lengthy sexual maturation period of four to five years poses a significant obstacle to the genetic reproduction and breeding of grass carp. Consequently, classical methods such as gonadogenesis or sex reversal through steroid treatment, employed for breeding all-female grass carp, demand considerable time and effort. In this study, we developed an super-fast breeding strategy for generating all-female grass carp in a total of half a year, using a surrogate production method. We first characterized grass carp female germline stem cells (GSCs) from genetic female juveniles at three months post-fertilization (mpf). The female GSCs with XX chromosomes were then transplanted into germ cell-depleted zebrafish larvae at five days post-fertilization (dpf). The transplanted grass carp XX germ cells underwent rapid spermatogenesis in the zebrafish recipient. At three months after transplantation, all zebrafish recipients had developed into males capable of producing the all-X sperm of the grass carp. By using these sperm to fertilize wildtype grass carp eggs, we successfully produced an all-female grass carp offspring. This groundbreaking achievement highlights the potential of surrogate production in the genetic breeding of valuable fish species, and opens a new avenue for advancing genetic breeding in aquaculture.

sample_organism:
Danio rerio
Ctenopharyngodon idella

platform_organism:
Danio rerio
Ctenopharyngodon idella

overall_design: The GSCT (Germline stem cell transplantation), grass carp and zebrafish sperm were applied to RNA-seq analysis to further examine the genetic consistency among them.

first: Ding

last: Ye


title: Zygotic Arrest 1 Pathogenic Variants Disrupt Maternal mRNA Stability and Cause Oocyte Maturation Defects in Humans

city: Nanning

first: liu

last: shaoyuan

address: Guangxi province of china

institute: Guangxi university

summary: Zygotic arrest 1 (ZAR1) is one of the earliest identified maternal-effect genes that function during the maternal-to-zygotic transition (MZT) in mice. However, the role of human ZAR1 was unclear. This study investigated the association of ZAR1 gene variants and infertility in women, focusing on their impact on oocyte and early embryonic development. In five individuals with unexplained female infertility, defects in oocyte maturation and early embryonic arrest were observed during in vitro fertilization treatment. Whole-exon sequencing identified either homozygous or compound heterozygous variants of ZAR1 in the patients. Functional analysis revealed that the V118A mutation disrupted the localization of ZAR1 to the mitochondria-associated ribonucleoprotein domain (MARDO) structure, a key mRNA storage site in oocytes. In contrast, R397Q and S121* mutations impaired ZAR1’s RNA-binding capability. These variants disrupt MARDO formation and function, thereby negatively affecting oocyte maturation and early embryonic development. Additionally, transcriptome analysis of oocytes and embryos from ZAR1 variant carriers showed downregulation of maternal mRNAs and altered translation profiles, indicating a disrupted MZT. This study highlights the critical roles of ZAR1 and MARDO in human reproduction and provides insights into the molecular mechanisms underlying some forms of female infertility.

overall_design: Expression profiling by high throughput sequencing.RNA sequencing (RNA-seq) was applied to both healthy control and patient-derived oocytes at the germinal vesicle (GV) stage, as well as to embryos at the 1-cell stage.

first: Wei

last: Zheng

first: Qianqian

last: Sha


title: Zinc finger homeobox-3 (ZFHX3) exerts genome-wide regulation of dailygene expression in the suprachiasmatic nucleus

city: Didcot

first: Akanksha

last: Bafna

address: Harwell Campus

institute: MRC HARWELL

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

overall_design: Refer to individual Series


title: Zinc finger homeobox-3 (ZFHX3) exerts genome-wide regulation of dailygene expression in the suprachiasmatic nucleus [RNA-seq]

summary: The mammalian suprachiasmatic nucleus (SCN), situated in the ventral hypothalamus, directs daily cellular and physiological 24-hr rhythms across the body. The cell-autonomous timing mechanism is based on a self-sustaining transcriptional-translational feedback loop (TTFL) that in turn co-ordinates the expression of clock-controlled genes (CCGs) that direct circadian programmes of SCN cellular activity.  In the mouse, the transcription factor, ZFHX3 (zinc finger homeobox-3), is necessary for the development of the SCN and influences circadian behaviour in the adult.  The molecular mechanisms by which ZFHX3 affects the SCN at transcriptome and genome levels are, however, poorly defined. Here, we used chromatin immunoprecipitation (ChIP-seq) to map the genomic localization of ZFHX3 sites in the SCN. In parallel, we conducted a comprehensive SCN RNA sequencing at six distinct times-of-day, and then compared the transcriptional profile between the control and ZFHX3-conditional null mutants. We show that the genome wide occupancy of ZFHX3 occurs predominantly around the gene transcription start site (TSS), co-localizing with the known histone modifications, and preferentially partnering with the core-clock TFs (CLOCK, BMAL1) to regulate the clock gene(s) transcription. Correspondingly, we show that the conditional loss of ZFHX3 in the adult has a dramatic effect on the SCN transcriptome,  reducing the level of neuropeptides-encoding transcripts and abolishing rhythmic (24-h) oscillation of the clock TF, Bmal1.  In addition, various TTFL genes and CCGs exhibited altered circadian expression profiles, consistent with the advanced daily behaviour under 12hr light- 12hr dark conditions. Taken together, these findings reveal the extensive genome-wide regulation conferred by ZFHX3 in the central clock that orchestrates daily timekeeping in mammals.

overall_design: Gene expression profiling analysis of RNA-seq data in the SCN at six distinct time-points , ZT2,6,10,14,18,22 from control (Creneg), tamoxifen control (Pre-tam) and ZFHX3-KO (Post-tam) conditions. Each time-point had 2-4 biological replicates. Each biological replicate constituted 3-4 individual SCN samples.

first: Gareth

last: Banks

first: Michael

middle: H

last: Hastings

first: Patrick

last: Nolan


title: Transcriptional analysis of dendritic cells upon TLR stimulation reveals an immunosuppressive profile [cut&tag]

first: Ling

address: zhongshan road No.74

institute: Sun Yat-sen University

summary: Dendritic cells (DCs) simultaneously prime and dampen the immune response. The transcriptional program that regulates immunosuppression in DCs is currently unknown. Here we identified ETS homologous factor as a key transcription factor that regulates DC immunosuppression.

overall_design: cDCsFL-Notch were cultured for 6 days and stimulated with 100 nM TLR stimulation. Live B220- MHCII+ CD11c+ cells were FACS-sorted after 18 hours of TLR stimulation.

first: Xiaoli

last: Liu

first: Cliff

middle: Y

last: Yang


title: Transcriptional analysis of dendritic cells upon TLR stimulation reveals an immunosuppressive profile [PLF scRNA-seq]

overall_design: Mice were immunized intraperitoneally (i.p.)  with or without 10 µgTLR stimulation in 100 µL PBS, peritoneal cavity cells were obtained by peritoneal cavity lavage twice with 5 mL of ice-cold PBS. They were collected 6 hrs after injection for cDCs (live MHCII+ CD11c+) FACS-sorting for scRNA-seq library preparation.


title: Low expression of miR-182 caused by DNA hypermethylation accelerates acute lymphocyte leukemia development by targeting PBX3 and BCL2: miR-182 promoter methylation is a predictive marker for hypomethylation agents + BCL2 inhibitor venetoclax

city: Wenzhou

first: Shenmeng

last: Gao

state: Zhejiang Province

address: 1 Xuefubei Street, Ouhai District

department: Laboratory of Internal Medicine

institute: The First Affiliated Hospital of Wenzhou Medical University

summary: miR-182 promoter hypermethylation frequently occurs in various tumors, including acute myeloid leukemia, and leads to low expression of miR-182. However, whether adult acute lymphocyte leukemia (ALL) cells have high miR-182 promoter methylation has not been determined. miR-182 (miR-182-5P) expression was substantially lower in ALL blasts than in normal controls (NCs) because of DNA hypermethylation at the miR-182 promoter in ALL blasts but not in normal controls (NCs). Knockout of miR-182 (182KO) markedly accelerated ALL development, facilitated the infiltration, and shortened the overall survival in a BCR-ABL (P190)-induced murine B-ALL model. Furthermore, the 182KO ALL cell population was enriched with more leukemia-initiating cells (CD43+B220+ cells, LICs) and presented higher leukemogenic activity than the 182WT ALL population. Furthermore, depletion of miR-182 reduced the overall survival in a Notch-induced murine T-ALL model, suggesting that miR-182 knockout accelerates ALL development. Mechanistically, overexpression of miR-182 inhibited proliferation and induced apoptosis by directly targeting PBX3 and BCL2, two well-known oncogenes that are key targets of miR-182. Most importantly, DAC in combination with Ven had synergistic effects on ALL cells with miR-182 promoter hypermethylation, but not on ALL cells with miR-182 promoter hypomethylation. Collectively, we identified miR-182 as a tumor suppressor gene in ALL cells and low expression of miR-182 because of hypermethylation facilitates the malignant phenotype of ALL cells. DAC+Ven cotreatment might has been applied in the clinical try for ALL patients with miR-182 promoter hypermethylation. The frequency of methylation at the miR-182 promoter should be a potential biomarker for DAC+Ven treatment in ALL patients.

overall_design: To investigate the function of miR-182 in ALL, we constructed miR-182 knockout (182KO) and 182 wild type (182WT) B-ALL and T-ALL mice model induced by BCR-ABL (P190) and Notch, respectively. We then performed gene expression profiling analysis using data obtained from RNA-seq of 2 different leukemia cells isolated from bone marrow.

first: Danyang

last: Li

first: Yigang

last: Yuan

first: Chen

last: Meng


title: Transcriptional analysis of dendritic cells upon TLR stimulation reveals an immunosuppressive profile [PBMC scRNA-seq]

last: Wang

overall_design: PBMCs of four male healthy volunteers were plated individually in complete RPMI-1640 (RPMI-1640 with 10% FCS, 1% L-glutamate, 1% penicillin/streptomycin, and 55 μM 2-mercaptoethanol), and stimulated with 100 nM TLRstimulation for 16 hrs. Then, DCs (Lin- CD123- CD11c+ HLA-DR+) were FACS-sorted and pooled for scRNA-seq library preparation.


title: Transcriptional analysis of dendritic cells upon TLR stimulation reveals an immunosuppressive profile [pLN scRNA-seq]

overall_design: Mice were immunized subcutaneously (s.c.) in the rear footpads with or without 10 µg TLR stimulation in 40 µL PBS. Popliteal lymph node cells were collected 48 hrs after TLR stimulation  for cDCs (live MHCII+ CD11c+) FACS-sorting for scRNA-seq library preparation.


title: Zinc finger homeobox-3 (ZFHX3) exerts genome-wide regulation of dailygene expression in the suprachiasmatic nucleus [ChIP-seq]

overall_design: Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for ZFHX3 using the SCN tissues from WT C57BL/6J mice.


title: Transcriptional analysis of dendritic cells upon TLR stimulation reveals an immunosuppressive profile [Spl scRNA-seq]

overall_design: Mice were immunized intraperitoneally (i.p.) with or without 60 µg TLR stimulation in 100 µL PBS. Splenic cells were collected 18 hrs after  injection for cDCs (live MHCII+ CD11c+) FACS-sorting for scRNA-seq library preparation.


title: Identification and characterisation of potential targets of N6-methyladenosine (m6A) modification in the process of lumbar disc degeneration

city: shanghai

last: huan

address: shanghaisongjiang

institute: cloudseq

summary: In this study, to explore the potential involvement of m6A modification in intervertebral disc degeneration, we conducted RNA m6A modification profiling in 3 human control nucleus pulposus tissue and 3 degenerative nucleus pulposus tissue by m6A-RIP-sequencing.

overall_design: Biological replicates: 3

first: Jianlin

last: Shen

first: Qiang

last: Zhang

first: Yujian

last: Lan

first: Qingping

last: Peng

first: Ziyu

last: Ji

first: Yanjiao

last: Wu

first: Huan


title: Effect of high glucose on Schwann cells‘ exosomes

city: Shanghai

first: Yingying

state: Shanghai

address: No.639 Zhizaoju Road

institute: Shanghai Ninth People’s Hospital, Shanghai Jiao Tong University School of Medicine

summary: Study on the effect of high glucose on miRNA carried by exosomes of Schwann cells

overall_design: Prepare a miRNA sequencing library using the total RNA of each sample, detect the miRNA expression level of each sample through Illumina Novaseq 6000, and screen for differentially expressed miRNAs.

first: Ximeng

last: Cao

first: Yifan

first: Chun

last: Xu


title: RNA-seq Analysis of Acute myeloid leukemia (AML) primary samples

city: Jena

first: Faezeh

last: Ghazvini Zadegan

address: Hans-Knoll-Strase 2

department: Department of Hematology/Oncology, Clinic of Internal Medicine II

country: Germany

institute: Institute of Molecular Cell Biology, Center for Molecular Biomedicine Jena (CMB)

summary: In this study, we examined gene expression in 69 primary samples from patients with Acute Myeloid Leukemia (AML), a complex hematologic malignancy, aiming to uncover potential avenues for novel treatment strategies.

overall_design: Primary human AML samples were isolated from peripheral blood mononuclear cells utilizing StemSep columns (STEMCELL Technologies, Canada). These samples were subsequently cultured in Stemline II Medium (Sigma, Hamburg, Germany) prior to harvesting for gene expression experimentation.


title: Effects of Candida spp. colonization in the gut for germ-free mice on colonic gene expression

city: Taipei

first: Hao-Sen

last: Chiang

address: No.1, Roosevelt Road

department: Department of Life Science

country: Taiwan

institute: National Taiwan University

summary: To investigate the changes in transcriptomic profiles of colonic samples by the colonization of two Candida spp. in the gut. Both species upregulate the pathways related to protein processing in the endoplasmic reticulum, antigen processing, and presentation. In addition, C. tropicalis enhances cell proliferation and cytokine-related genes.

overall_design: C57BL/6 female mice (6 weeks old) were housed in gnotobiotic isolators for the experiment. Mice were colonized via oral gavage with C. albicans SC5314 and C. tropicalis MYA-3404 (10^8 CFU) for 30 days. We then collected colon for RNA-seq run.

first: Yi-Chun

last: Chou

first: Ha

last: Doan


title: Gene expression profiling of early events of DSF elicitor based defense signalling in Arabidopsis thaliana

city: Hyderabad

first: Subhadeep

last: Chatterjee

address: Uppal, Hyderabad

country: India

institute: Center for DNA Fingerprinting and Diagnostics, Hyderabad

summary: To study the early events of priming response leading to changes in gene expression in Arabidopsis thaliana Col0 wild type plant on infiltration with the potent elicitor molecule Diffusible Signal Factor (DSF). In the present experimentation we have employed total RNA microarray expression profiling as a discovery platform to identify differentially expressing genes that are upregulated and down regulated in the various pathways involved in priming the defense responses by the elicitor molecule DSF on infiltration in the host plant Arabidopsis thaliana. This would help understand the possible defense pathways elicited by the elicitor molecule during interaction. The rosette leaves of Arabidopsis thaliana Col0 plants of 4 weeks old were infiltrated with 100uM DSF and the total RNA samples at 4hpi, 8hpi and 16hpi were extracted along with methanol treatment serving as a control  in two biological replicates 1A and 2A. Further, these RNA samples were checked for the quality and subjected to microarray  analysis. The differentially expressing candidate genes were identified specific to DSF treatment and were quantified by real-time PCR post DSF treatments.

overall_design: Agilent Arabidopsis Gene Expression 8x60k Microarray: AMADID: 48015

first: Durga

last: Bhavani