title: Single-cell transcriptomics reveals the cellular identity of a novel progenitor population crucial for murine neural tube closure

first: Zihao

last: Deng

state: VIC

address: L6, The Alfred Centre, 99 Commercial Rd

department: Medicine (Alfred Hospital)

summary: Neural tube closure in vertebrates is achieved through a highly dynamic and coordinated series of morphogenic events involving neural plate, surface ectoderm, and neural plate border. Failure of this process in the caudal region causes spina bifida. Grainyhead-like 3 (GRHL3) is an indispensable transcription factor for neural tube closure as constitutive inactivation of which leads to fully penetrant spina bifida. Here, through single-cell transcriptomics we show that at E8.5, the time-point preceding mouse neural tube closure, the co-expression of Grhl3, Tfap2a, and Tfap2c defines a previously unrecognised progenitor population of surface ectoderm. Specific deletion of Grhl3 expression using Tfap2a-Cre recapitulate the spina bifida observed in Grhl3-null animals. Moreover, conditional inactivation of Tfap2c expression in Grhl3-expressing neural plate border cells also causes mild spina bifida. These findings clearly indicate that Grhl3-expressing neural plate border cells cohort is required for the early-stage neurulation.

overall_design: E8.5 wild-type and Grhl3-null embryos were harvested at 3 p.m. on the eighth day post-fertilisation. The caudal half of five wild-type and three Grhl3-null embryos was separated to generate single cell suspension. Cells from five wild-type embryos and three Grhl3-null embryos were then pooled. Two wild-type and two Grhl3-null single cell-RNA sequencing libraries were constructed for analysis.

first: Marina

last: Carpinelli

first: Tariq

last: Butt

first: Graham

last: Magor

first: Peinan

last: Zhao

first: Kevin

last: Gillinder

first: Andrew

middle: C

last: Perkins

first: Stephen

last: Jane


title: IL-12 drives the expression of the inhibitory receptor NKG2A on human tumor-reactive CD8 T cells

status: Public on Oct 08 2024

city: Portland

first: Wesley

last: Rosales

state: OR

address: 4805 NE Glisan St

country: USA

institute: Earle A. Chiles Research Institute

type: Expression profiling by high throughput sequencing

summary: Blockade of NKG2A/HLA-E interaction is a promising strategy to unleash the anti-tumor response. Yet the regulation of NKG2A expression on CD8+ lymphocytes and the T cell anti-tumor response are still poorly understood. Here, by performing CITE-seq on human T cells derived from head and neck squamous cell carcinoma and colorectal cancer, we show that NKG2A expression is induced in tumor-infiltrating  CD8+ T cells differentiating into cytotoxic, CD39+CD103+ double positive (DP) T cells. This developmental trajectory lead to TCR repertoire overlap between the NKG2A– and NKG2A+ DP CD8 T cells, suggesting shared antigen specificities. Mechanistically, IL-12 is essential for the expression of the NKG2A on naïve CD8+ T cells in a CD40/CD40L- dependent manner, in conjunction with TCR stimulation and increased TGF-β levels. Our study thus reveals that NKG2A is induced by IL-12 on human tumor-reactive CD8+ T cells exposed to a TGF-b-rich environment, highlighting an underappreciated immuno-regulatory feedback loop dependent on IL-12 stimulation.

overall_design:
CD3+ T cells were FACS sorted from tumor digest after gating on Live (PI-) CD45+CD3+ cells and analyzed by scRNA-seq

***************************************************************
Submitter states that missing raw files are due to patient privacy concerns.
***************************************************************

first: Olivier

last: Fesneau

first: Thomas

last: Duhen


title: The grass–microbiome-rumen axis in grazing sheep farming intervened by supplementary feeding

city: Beijing

first: hua

middle: xiu

last: ma

address: NO.2 Yuanmingyuan weat road

country: China

institute: China Agriclutural university

summary: This study identifies key microbiome and epithelial cell subtypes involved in grass digestion and VFA metabolism in the rumen. By integrating multi-omic data, we reveal novel links between microbial activity, epithelial cell function, and grassland foraging, providing critical insights into mechanisms underlying grass prevalence and their implications for optimizing ruminant health and productivity. This research enhances our understanding of the grass-microbiome- rumen axis and its role in sustainable grazing systems.

overall_design: we selected six lambs using scRNAseg,  tissues were collected from the ven_x0002_tral sac region of the rumen.


title: Comprehensive profiling of migratory primordial germ cells reveals niche-specific differences in non-canonical Wnt and Nodal-Lefty signaling in anterior vs posterior migrants

city: San Francisco, CA

first: Rebecca

middle: Garrett

last: Jaszczak

state: California

address: 35 Medical Center Way, Room 913D

institute: UCSF

summary: Mammalian primordial germ cells (PGCs) migrate asynchronously through the embryonic hindgut and dorsal mesentery to reach the gonads. To characterize transcriptional heterogeneity of migrating PGCs and their niches, we performed single-cell RNA sequencing of 13,262 mouse PGCs and 7,868 surrounding somatic cells during migration (E9.5, E10.5, E11.5) and in anterior versus posterior locations to enrich for leading and lagging migrants. Analysis of PGCs by position revealed dynamic gene expression changes between faster or earlier migrants in the anterior and slower or later migrants in the posterior at E9.5. At E10.5, we found that anterior PGCs upregulate Nodal transcriptional targets including Lefty1/2 and validated this LEFTY1/2 upregulation via whole-mount immunofluorescence staining. This positional and temporal atlas of mouse PGCs supports the idea that niche interactions along the migratory route elicit changes in proliferation, actin dynamics, pluripotency and epigenetic reprogramming.

overall_design: We pooled sorted GFP+ germ cells and an equal amount of GFP- somatic cells from each timepoint and anatomical location. We loaded counted cells onto a 10X V2 chip to create a single cell emulsion. For E9.5 Anterior, we loaded 3400 cells, and for E9.5 Posterior, 2800 cells. For E10.5 Anterior, we loaded two technical replicates totaling 9300 cells. For E10.5 Posterior, we loaded two technical replicates totaling 11200 cells. For the two E11.5 biological replicates, we loaded 15,000 and 10,000 cells, respectively. Library creation was done in house following 10X ChromiumTM Single Cell 3' Reagent Kits v2 User Guide Rev A. E9.5 and E11.5 libraries were sequenced on HiSeq 4000 (Illumina) with paired-end sequencing parameters: Read1, 98 cycles; Index1, 14 cycles; Index2, 8 cycles; and Read2, 10 cycles. E10.5 libraries were sequenced on Novaseq (Illumina) with paired-end sequencing parameters: Read1, 150 cycles; i7 Index, 8 cycles; i5 index, 0 cycles; and Read2, 150 cycles.

middle: G

first: Jay

last: Zussmann

first: Diana

middle: J

last: Laird


title: Maternal immune activation upregulates the AU020206-IRFs-STAT1 axis in modulating cytokine production in the brain

status: Public on Oct 09 2024

city: Guangzhou

first: Wenjun

last: Yu

address: Nanfang Hospital, Southern Medical University

institute: Center for Genetics and Developmental Systems Biology, Department of Obstetrics and Gynecology

summary: Maternal immune activation (MIA) is reported to increase the risk of psychiatric disorders in the offspring. However, the underlying mechanism remains unclear. Here, we constructed an MIA mouse model by intraperitoneal injection of LPS into pregnant mice and evaluated the behaviors and gene expression profiles in the brains of the female and male offspring, respectively. We found that the MIA female offspring exhibited increased anxiety and a large number of differentially expressed genes (DEGs) in the brain, which were enriched with disease genes of psychiatric disorders and immune functions. In contrast, the MIA male offspring exhibited no significant abnormal behaviors and only a small number of DEGs, which were not enriched with disease genes and immune functions. Therefore, we further pursued the downstream study on the molecular mechanism underlying the increased anxiety in the female offspring. We identified the lncRNA AU020206-IRFs-STAT1-cytokine axis by integrating lncRNA-protein interaction data and TF-promoter interaction data and verified the axis in vitro and in vivo. The female mice with overexpression of AU020206 in the prefrontal cortex exhibited anxiety behaviors. This study illustrates that MIA upregulates the AU020206-IRFs-STAT1 axis in controlling the brain immunity linked to abnormal behaviors, providing a basis for understanding the role of MIA in psychiatric disorders.

overall_design: Comparative gene expression profiling analysis of RNA-seq data on the whole brain of MIA female and male offspring mice.

first: Jing

last: Yang


title: Newly identified ZNHIT3 variants, which cause intrauterine fetal loss, disrupt snoRNP assembly

city: Atlanta

first: Bing

last: Yao

address: 615 Michael Street

department: Department of Human Genetics

institute: Emory University

summary: ZNHIT3 (zinc finger HIT type containing protein 3) is an evolutionarily conserved protein required for ribosome biogenesis by mediating the assembly of small nucleolar RNAs (snoRNAs) of class C/D into ribonucleoprotein complexes (snoRNPs). Missense mutations in the gene encoding ZNHIT3 protein have been previously reported to cause PEHO syndrome, a severe neurodevelopmental disorder typically presenting after birth. We present here the case of two fetuses from a single family who presented with isolated hydrops during the early second trimester of pregnancy, resulting in intrauterine demise. Autopsy revealed no associated malformation. Through whole-genome quartet analysis, we identified two novel variants within the ZNHIT3 gene, both inherited from healthy parents and occurring as compound heterozygotes in both fetuses. The c.40T>C p.Cys14Arg variant originated from the father, while the c.251_254delAAGA r.? p.? variant was of maternal origin. Western capillary analysis of ZNHIT3 protein in lymphoblastoid cell lines of the parents shows that the parental c.40T>C p.Cys14Arg variant causes a significant drop in steady-state ZNHIT3 protein level. Analysis of the variants in human cell culture models reveals that both variants reduce cell growth, albeit to different extents, and impact the protein's stability and function in distinct ways. The c.251_254delAAGA results in production of a stable form of ZNHIT3 that lacks a domain required for mediating snoRNP biogenesis, whereas the c.40T>C p.Cys14Arg variation behaves similarly to the previously described PEHO-associated ZNHIT3 variants that destabilize the protein. Interestingly, both variations lead to a marked decrease in specific box C/D snoRNA levels, reduced rRNA levels and cellular translation. Analysis of rRNA methylation pattern in fetus samples reveals distinct sites of hypo 2’-O-methylation. RNA-seq analysis of undifferentiated and differentiated SHSY5Y cells transfected with the ZNHIT3 variants reveals differential expression of a set of genes, many of which are associated with developmental processes and RNA binding compared to cells expressing wild-type ZNHIT3. In summary, this work adds two novel ZNHIT3 variants to the other human disease-linked variants of this protein and describes the molecular defects caused by these pathogenic variants.

overall_design: The Lenti-viral plasmids of ZNHIT3-WT, ZNHIT3-C14R, and ZNHIT3-∆251-254 were generated from the FUGW plasmid (Addgene #14883) and transfected into SHSY5Y cell.

first: Lutfur

last: Rahman

first: Yangping

last: Li

first: Homa

last: Ghalei


title: Targeting TREX1 induces innate immune response in drug resistant Small Cell Lung Cancer

city: Philadelphia

first: Israel

last: Canadas

state: PA

address: 333 Cottman Ave.

department: Nuclear Dynamics and Cancer Program

institute: Fox Chase Cancer Center

summary: Small cell lung cancer (SCLC) is the most lethal type of lung cancer. Paradoxically, this tumor displays an initial exquisite response to chemotherapy; however, at relapse, the tumor is highly resistant to subsequent available therapies. Here, we report that the expression of three prime repair exonuclease 1 (TREX1) is strongly induced in chemoresistant SCLCs. ATAC-seq and ChIP-seq revealed a significant increase in chromatin accessibility and transcriptional activity of TREX1 gene locus in chemoresistant SCLC. Analyses of human SCLC tumors and patient-derived xenografts (PDXs) also showed increase in TREX1 expression in post-chemotherapy samples. TREX1 depletion caused the activation of cyclic GMP-AMP synthase (cGAS)-stimulator of interferon genes (STING) pathway due to cytoplasmic accumulation of damage-associated double-stranded DNA, inducing immunogenicity and enhancing the sensitivity of drug resistant cells to chemotherapy. These findings suggest TREX1 upregulation may partially contribute to the survival of resistant cells, and its inhibition may represent a promising therapeutic strategy to enhance antitumor immunity and potentiate the efficacy of chemotherapy and/or immunotherapy in chemoresistant SCLCs.

overall_design: To investigate the function of TREX1, we knocked-down the target gene in H69AR cell line.

first: Takahiko

last: Murayama

first: Jun

last: Nakayama

last: Cañadas


title: Correction of Exon 2, Exon 2-9 and Exons 8-9 duplications in DMD patient myogenic cells by a Single CRISPR/Cas9 system

city: EVRY-COURCOURONNES

first: Tao

last: WANG

state: France

address: 1 bis, rue de l’Internationale

department: R & D

country: France

institute: genethon

summary: Duchenne Muscular dystrophy (DMD), a yet-incurable X-linked recessive disorder that results in muscle wasting and loss of ambulation is due to mutations in the dystrophin gene. Exonic duplications of dystrophin gene are a common type of mutations found in DMD patients. In this study, we utilized a single guide RNA CRISPR strategy targeting intronic regions to delete the extra duplicated regions in patient myogenic cells carrying duplication of exon 2, exons 2 to 9, and exons 8 to 9 in the DMD gene. Immunostaining on CRISPR-corrected derived myotubes demonstrated the rescue of dystrophin protein. Subsequent RNA sequencing of the corrected myotubes indicated rescue of dystrophin related molecular pathways. Examination of predicted close-match off-targets evidenced no aberrant gene editing at these loci. Here, we further demonstrate the efficiency of a single guide CRISPR strategy capable of deleting multi-exon duplications in the DMD gene without significant off target effect. This resulted in the restoration of dystrophin expression and reversal of transcriptome pathological signatures in the corrected patient-derived cells. Our study contributes valuable insights into the safety and efficacy about the application of the single guide CRISPR strategy as a potential therapeutic approach for DMD patients with duplications of variable size.

overall_design: To profile the editing effect of DMD gene correction, we did RNAseq on the wt myoblasts C25 differentiated myotubes (8 days), as welll as three DMD duplicated cell lines and the corresponding CRISPR corrected cell lines with three biological repeats.


title: Decapping activators Edc3 and Scd6 act redundantly with Dhh1 in nutrient-replete cells to post-transcriptionally repress starvation-induced pathways [Ribo and RNA-seq]

city: Bethesda

first: Alan

last: Hinnebusch

state: MD

address: 6 Center Drive, Building 6, Room 233

department: NICHD

institute: NIH

summary: Degradation of many yeast mRNAs involves decapping by Dcp1:Dcp2. Previous studies on decapping activators Edc3 and Scd6 suggested limited roles in promoting mRNA decay in yeast cells. RNA-seq analysis of mutants lacking one or both proteins reveals that Scd6 and Edc3 have largely redundant activities in targeting numerous mRNAs for degradation, which are masked in the single mutants. These transcripts are frequently targeted by decapping activators Dhh1 and Pat1 and the evidence suggests that Scd6/Edc3 act interchangeably to recruit Dhh1 to Dcp2 independently of Pat1. Ribosome profiling shows that redundancy between Scd6 and Edc3 and their functional interactions with Dhh1 and Pat1 extends to translational repression of particular transcripts, including a cohort of poorly translated mRNAs displaying interdependent regulation by all four factors. Scd6/Edc3 also participate with Dhh1/Pat1 in post-transcriptional repression of proteins required for respiration and catabolism of non-optimal carbon sources, which are normally expressed only in limiting glucose. Simultaneously eliminating Scd6/Edc3 increases mitochondrial membrane potential and elevates metabolites of the tricarboxylic acid and glyoxylate cycles critical for growth at low glucose levels. Thus, Scd6/Edc3 act redundantly, in parallel with Dhh1 and in cooperation with Pat1, to adjust gene expression to nutrient availability by controlling mRNA decapping and decay.

overall_design: Ribosome profile and RNA-sequencing studies include total 16 samples, 8 for ribosome profiling and 8 for total RNA-sequencing from duplicate cultures of WT, edc3∆, scd6∆, and edc3∆scd6∆ yeast strains. The RNA-Seq samples had ERCC Spike-In added to the samples.  Yeast strains were grown in YPD to OD600 = 0.6~0.8 at 30˚C, then flash frozen.  Ribosome protected or total RNA libraries were prepared from yeast cells and sequenced.

first: Rakesh

last: Kumar

first: Fan

last: Zhang

first: Shreyas

last: Niphadkar

first: Chisom

last: Onu

first: Anil

middle: K

last: Vijjamarri

first: Miriam

middle: L

last: Greenberg

first: Sunil

last: Laxman


title: Prolonged pause during follicle growth contributes to offspring throughout reproductive lifespan

city: San Francisco

first: Bikem

last: Soygur

address: 35 Medical Center Way, Parnassus, RMB, PodB, 913D

department: Obstetrics, Gynecology & Reproductive Sciences

summary: Ovulation results from cyclical recruitment of quiescent oocytes for growth, hence the primordial follicles established during development comprise the lifelong reserve. In mouse fetal oocytes, we lineage traced a subset with earlier onset and slower progression through meiotic prophase using Sycp3CreERT2/+; Rosa26nT-nG/+ females. Initially GFP labeled first meiotic entrants survived at elevated rates and contributed to primordial follicles as well as the first wave of growing follicles in neonates. However, by puberty, only oocytes in the first wave of growing follicles remained. To identify the transcriptional signature of GFP+ oocytes in growing follicles, we performed Bulk RNA sequencing, which revealed remarkable similarity between the transcriptomes of GFP+ (first meiotic entrants) and RFP+ (the rest of the oocytes), with only 12 genes showing differential expression.

overall_design: MII oocytes from Sycp3CreERT2/+; Rosa26nT-nG/+ female mice were collected and categorized into three groups (GFP high, GFP low, and RFP) based on their GFP and RFP fluorescence signals. The oocyte numbers used for the three replicates in each group were as follows: GFP high: 15, 15, 16; GFP low: 49, 52, 57; and RFP: 88, 87, 100. Total RNA isolated from oocytes was enriched for poly-A mRNA followed by library preparation for RNA-seq. Following quality control, libraries were pooled and sequenced on an Illumina platform.

first: Steven

middle: A

last: Cincotta

first: Diana

last: Laird


title: Decapping activators Edc3 and Scd6 act redundantly with Dhh1 in nutrient-replete cells to post-transcriptionally repress starvation-induced pathways [ChIP-seq]

type: Genome binding/occupancy profiling by high throughput sequencing

overall_design: ChIP-Seq study includes 6 samples.  Triplicates of each wild type and edc3∆scd6∆ yeast strain.


title: Expansion of in vivo HDAd-transduced HSPCs by constitutive expression of tHMGA2 without additional treatment allows for therapeutically relevant transgene expression in HSPC progeny cells without clonal dominance.

city: Boston

first: Robert

last: Policastro

state: Massachusetts

address: 451 D St Unit #600

institute: Ensoma Inc.

summary: We developed an in vivo hematopoietic stem and progenitor cell (HSPC) gene therapy approach that does not require HSC cell transplantation. To achieve therapeutically relevant numbers of corrected cells, we tested truncated HMGA2 instead of chemo-selection for enrichment of transduced HSPCs. We constructed HSC-tropic HDAd5/35++ vectors expressing a 3’UTR HMGA2 truncated gene and a GFP reporter gene. A SB100x transposase vector was used to mediate random integration of the tHMGA2/GFP transgene cassette. Mice were mobilized by subcutaneous injections of G-CSF and AMD3100/Plerixafor and intravenously injected with the integrating tHMGA2/GFP vector. This resulted in slow but progressive competitive expansion of GFP+ PBMCs, reaching about 50% by week 44 with further expansion in secondary recipients. Expansion occurred at the level of HSCs as well as at the level of myeloid, lymphoid, and erythroid progenitors within the bone marrow and spleen. Importantly, based on genome-wide integration site analyses, expansion was polyclonal, without any signs of clonal dominance. The results were validated in humanized mice. This is the first demonstration that simple subcutaneous and intravenous injections of mobilization drugs and HDAd vectors can trigger autocatalytic, self-limiting expansion of in vivo transduced HSPCs resulting in marking rates that, theoretically, are curative for hemoglobinopathies.

overall_design: CD46 transgenic mice were mobilized by subcutaneous (SC) injections of human recombinant G-CSF (250 mg/kg/mouse/day, 4 days) followed by an SC injection of AMD3100 (5mg/kg) on day 5. In addition, animals received Dexamethasone (10 mg/kg, IP) 16 h and 2 h before virus injection to blunt innate toxicity associated with intravenous HDAd injection. Forty-five minutes after AMD3100, animals were intravenously injected with HDAd virus vectors through the retro-orbital plexus (4x1010 viral particles per mouse). Group 1 (N=10) were injected with HDAd-mgmt/GFP + HDAd-SB). Group 2 received HDAd-tHMGA2/GFP + HDAd-SB. DNA from bone marrow MNCs of week 44 primary mice with different rates of GFP marking in PBMCs (ranging from 25% to 65% ) were subjected to genome-wide transgene integration site analysis.

first: Hongjie

last: Wang

first: Aphrodite

last: Georgakopoulou

first: Raïssa

last: Eluère

first: Ka

middle: W

last: Mok

first: André

last: Lieber


title: The Distinct Profile of Tumor-Reactive T Cells in RUNX1::RUNX1T1 AML Bone Marrow Unveiled by Single-Cell Analysis

city: Tianjin

first: Yihan

last: Mei

address: Nanjing road

institute: Peking union medical school

summary: Unlike the well-known knowledge of tumor-infiltrating T cells in solid tumors, the overall T cell state and the characteristics of tumor-reactive T cells in RUNX1:RUNX1T1 acute myeloid leukemia remain unclear. In this study, we employed single-cell transcriptome sequencing and single-cell TCR sequencing to elucidate the characteristics of T cells in the bone marrow of RUNX1:RUNX1T1 leukemia patients. Both CD4+ and CD8+ T cells exhibited features of reduced immune reserves and an increase in terminally differentiated T cells, indicative of immune mobilization.

overall_design: scRNA-seq of RUNX1::RUNX1T1 mouse and wild type


title: Monitor the dynamic changes of GBM cell lines under TTFields treatment

city: Los Angeles

first: David

last: Tran

state: CA

address: 1441 Eastlake Ave

department: Department of Neurological Surgery

institute: University of Southern California

summary: We used 3 established GBM cell lines to generate TTFields-resistant cells using Inovitro system.

overall_design: To generate TTFields-resistant GBM cells, we utilized the Inovitro™ system to treat GBM cells continuously with TTFields (200 kHz unless otherwise noted) until resistant cells emerged. To ensure that changes in proliferative rates were not due to culture confluency and to determine temporal changes in gene expression as resistance developed, we maintained a constant growth condition by reseeding cells at the same starting density every seven days (7D) of TTFields treatment followed by a two-day treatment break to allow cells to recover before a new treatment cycle resumed. Numbers of live cells were enumerated on days two, four, and seven of each treatment cycle. At different indicating time points, total RNA was also collected for RNA-sequencing (RNA-seq) from the remaining cells. A maximum of five treatment cycles were performed for each cell line. For control, the same cells were subjected to the identical number of culturing cycles without TTFields treatment (non-TTFields-treated or NT). The libraries are named as {sample_name}_{batch}_{replicate_number}_{technical_replicate}. There is only one library that has a techincal replicate. While analyzing the data, make sure to correct for any batch effect as recorded in the "batch" field.

middle: D

first: Dongjiang

last: Chen

first: Son

middle: B

last: Le

first: Harshit

last: Manektalia


title: Transcriptome profile of Haloferax volcanii H26 strain under sodium bisulfite induction (RNA-seq)

city: Shenzhen

first: Jinquan

last: Li

state: Guangdong

address: 3688 Nanhai Avenue

institute: Shenzhen University

summary: DNA deamination occurs constantly in a cell and causes DNA damage. As this damage can be deleterious, organisms have evolved many systems to eliminate it. Deamination of cytosine, guanine, adenine, and 5-methylcytosine results in the formation of uracil, xanthine, hypoxanthine, and thymine, respectively. Sodium bisulfite is a kind of DNA deaminating agent that can increase the frequency of DNA deamination in cells. This study measures the transcriptome profile of  Haloferax volcanii H26 strain and HVO_RS06830 gene knockout strain, induced with different concentrations of sodium bisulfite.

overall_design: Haloferax volcanii H26 strain and HVO_RS06830 gene knockout strain were grown in Hv-YPC medium at 45°C until OD600 reach 0.5. Subsequently, sodium bisulfite was added to the final concentration of 0.1 or 0.5 mM and continuously cultured for 48 h. Cells were harvested by centrifugation at 12,000 rpm at 4°C for 20 min. After that, total RNA was extracted immediately using Direct-zol RNA MiniPrep Kits. Sequencing libraries were generated using NEBNext Ultra II Directional RNA Library Prep Kit for Illumina following manufacturer’s recommendations, and sequenced by Illumina NovaSeq6000 PE150 platform. The experiment was repeated two times.

first: Runyue

last: Xia

first: Wen-cong

last: Huang

first: Jiazheng

last: Gu

first: Meng


title: Differential usage of shared binding sites drives functional specialization of an essential family of transcription factors [RNA-seq]

first: Aman

last: Husbands

email: aman.husbands@gmail.com, ayh@upenn.edu

state: Pennsylvania

address: 103B Caroline Lynch Bldg, 433 S University Ave

institute: University of Pennsylvania

summary: Throughout evolution, the duplication and functional divergence of transcription factors (TFs) has driven cellular and organismal complexity. Mechanisms by which paralogous TFs functionally diverge are thus of broad interest yet remain poorly understood. One well-established mechanism underlying TF divergence is the occupation and regulation of distinct sets of genes. Here we test for new mechanisms using CORONA (CNA) and PHABULOSA (PHB), two representative members of the CLASS III HOMEODOMAIN LEUCINE ZIPPER (HD-ZIPIII) family of plant TFs. CNA and PHB have largely overlapping binding profiles yet each paralog has hundreds of uniquely regulated targets. Regulation of a given gene thus depends on whether its local binding site is considered primed (inactive) or regulated (active) by CNA or PHB. This decision appears to be controlled, at least in part, by their lipid binding START domain, proposing a model in which HD-ZIPIII TFs use information integrated by their START domain to generate paralog-specific transcriptional outcomes at commonly bound genes. Taken together, our study identifies a new mechanism of TF paralog divergence and proposes the ubiquitously distributed START evolutionary module as a driver of functional divergence.

overall_design: To investigate genes differentially regulated after indiction of CLASS III HOMEOEDOMAIN LEUCINE ZIPPER (HD-ZIPIII) transcription factors. Sequencing was performed on 3 biological replicates of induced and mock induced genotypes of CORONA (CNA) and PHABULOSA (PHB) as well as chimeric CNA constructs.

first: Ashton

last: Holub

first: Sarah

last: Choudury

first: Ekaterina

last: Andrianova

first: Courtney

last: Dresden

first: Igor

last: Zhulin


title: Effects of paternal high cholesterol diet on F1 generation intimal transcriptome

city: Riverside

first: Changcheng

last: Zhou

address: 900 University Avenue

department: Biomedical Sciences

institute: The University of California Riverside

summary: Purpose: To investigate the effects of paternal high cholesterol diet on the F1 intimal transcriptome related to atherosclerosis. Methods:Three-week-old male LDL Receptor-deficient (LDLR-/-) male mice were fed a low-cholesterol diet (LCD) or high-cholesterol diet (HCD) for 8 weeks and then mated with age-matched control LDLR-/- female mice to generate F1 offspring. F1 LDLR-/- mice were fed a LCD for 16 weeks.  Intimal RNA was extracted by injecting Trizol into the aorta and collecting the flow through for further processing. Results: .  Total RNA isolated from the F1 intima were then subjected for RNA-sequencing analysis and results show that paternal hypercholesterolemia led to upregulation of 147 genes in the intimal of F1 female mice, many of which are involved in immune and inflammatory pathways.

overall_design: intimal RNA from F1 mice of high cholesterol or loq cholesterol LDLR-/- mice were process through RNA-seq.

first: Rebecca

last: Hernandez

first: Tong


title: Paternal hypercholesterolemia elicits sex-specific exacerbation of atherosclerosis in adult offspring of LDL receptor-deficient mice

type: Non-coding RNA profiling by high throughput sequencing

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

overall_design: Refer to individual Series


title: Effect of Drp1 inhibitor on gene expression profiles of RAW264.7 cells at 24 hours or 96 hours after LPS and IFN-γ stimulation

city: Kitakyushu

first: Jun-ichiro

last: Koga

state: --- Select One ---

address: 1-1, Iseigaoka, Yahatanishi-ku

country: Japan

institute: University of Occupatinal and Environmental Health, Japan

summary: Effects of Drp1 inhibitor Mdivi-1 on gene expression profiles of LPS and IFN-γ-treated RAW264.7 cells were examined.

overall_design: RAW264.7 cells were treated with 10 ng/mL LPS and 10 ng/mL mouse IFN-γ. Drp1 inhibitor Mdivi-1 was started simultaneously, and mRNAs were extracted 24 or 96 hours later.

first: Yuki

last: Kondo


title: Transcriptional shut-off of MAP kinase signaling enables pluripotency maintenance during diapause [ChIP-Seq]

first: Ryan

middle: Jared

last: Marina

state: MA

address: 45 Shattuck Street

department: Biological Chemistry and Molecular Pharmacology

institute: Harvard Medical School

summary: Exposure of unicellular or multicellular organisms to adverse environmental conditions, including nutrient deprivation, may induce a state of suspended animation or diapause. We here report that broad repression of RNA Pol II-driven gene expression by inhibiting the BET family's bromodomain-containing proteins (BET) triggers diapause in mouse embryonic stem (ES) cells. The diapause ES cells upregulate a functionally linked group of genes encoding negative regulators of MAP kinase signaling (NRMKS), which play a crucial role in ES cell differentiation. The elevated NRMKS expression is a hallmark of cells exposed to distinct diapause-inducing conditions, including mTOR inhibition, and are required for the pluripotency maintenance during diapause. Mechanistically, inhibition of mTORC1/2 leads to rapid decline of the Capicua transcriptional repressor (CIC) at the NRMKS gene promoters, followed by rapid transcriptional NRMKS gene upregulation. The mTOR and BET-dependent transcriptional switch supporting the undifferentiated state of the diapause ES cells suggests a broader usage of this mechanism in maintaining the undifferentiated state of metabolically dormant stem- or stem-like cells in different tissues.

overall_design:
To reveal the mechanism of elevated NRMKS expression in distinct diapause-inducing conditions, we focused on the Capicua transcriptional repressor (CIC) binding patterns at the NRMKS gene promoters.
We performed the CIC chromatin binding profiling analysis using data from CIC ChIP-seq in control and Torin1 treated ES cells for 60min and 2 weeks. Two biological replicates of ChIP samples and one replicate of input sample were included in each condition of the assay. To investigate the acute effects of mTORi on BRD4 and Histone acetylation levels, we also performed the BRD4 and Histone H4 acetylation chromatin binding profiling analysis using data from BRD4 and pan-H4Ac ChIP-seq in control and Torin1 treated ES cells for 15min and 30min.

first: Tuo

last: Zhang

first: Karen

last: Adelman


title: A transcriptome data set for comparing skin, muscle and DRG between acute and chronic postsurgical pain rats.

city: Shanghai

first: xiaoyan

last: Meng

address: Changhai road 225. Shanghai, China

institute: eastern hepatobililary surgical hospital

summary: Chronic postsurgical pain (CPSP), with a high prevalence and rising epidemic of opioids crisis, is typically derived from acute postoperative pain. Revealing the peripheral mechanisms behind the transition from acute to chronic pain after surgery is necessary. We established on two recognized animal models, the Lateral Paw Incision (LPI) model and the Skin/Muscle Incision and Retraction (SMIR) model, in simulation of acute and chronic postsurgical pain. The tissue of incisional skin, muscle and DRG at several time points are harvested and send for next-generation RNA sequencing (RNA-seq). Then the quality of sequencing results is validated. Our data could explain the time-course transcriptomic variation in the tissue of skin, muscle and DRG, and further to identify the potential origin and mechanism of CPSP.

overall_design: We established the Lateral Paw Incision (LPI) model as well as the Skin/Muscle Incision and Retraction (SMIR) model, in simulation of acute and chronic postsurgical pain, respectively. Mechanical allodynia via Von-frey test were applied to verify the quality of animal model. In LPI, at least of 4 qualified animals in the time of 0day, 6-hour, 2-day, 10-day were anesthetized for tissue harvest, while in SMIR, at least of 4 qualified animals in the time of 0day, 6-hour, 2-day, 10-day and 32-day were anesthetized for tissue harvest. The harvested tissues are the skin, muscle and DRG of the incisional area or side. A total of 36 rats, with 108 tissue samples were ultimately send for sequencing.

first: Xiaoyan

first: Lan

last: Bu

first: Ling

last: Shen

first: Kun-ming

last: Tao


title: Comparative gene expression profiling analysis of scRNA-seq data for MyoD-tdTomato high expressing cultured muscle stem cells from MyoD KI mice

city: Tsukuba

first: Ryo

last: Fujita

state: Ibaraki

department: Transborder Medical Research Center

institute: University of Tsukuba

summary: We previosuly generated MyoD-KI mice to distinguish the heterogenous MuSCs populations based on MYOD expression. Using the mice, we performed scRNA-seq to identify the unique gene expression profile in MyoD-tdTomato high expressing MuSCs.

overall_design: MyoD-tdTomato-high MuSCs were sorted using a MoFlo XDP flow cytometer (Beckman Coulter) based on our previous study.

first: Takuto

last: Hayashi

first: Wenchao

last: Gu

first: Takehito

last: Sugasawa


title: Effect of FcεRIα Deficiency on gene expression of gut epithial cell

first: Yue

last: Yin

address: NO.650 Xin Songjiang Road, Shanghai, P.R.China

institute: Department of Laboratory Medicine, Shanghai General Hospital, Shanghai Jiao Tong University School of Medicine

summary: To investigate the effect of FcεRIα knockout on colitis,we established FcεRIα knockout (FC) mice. We then performed gene expression profiling analysis using data obtained fromRNA-seg of gut of wide type mice (n=3)and FC mice (n=3)

overall_design: Comparative gene expression profiling analysis of RNA-seg data for gut of wideype mice(n=3)and FC mice(n=3)

first: Y

last: Yue

first: W

last: Ruilong

first: M

last: Qixiang

first: L

last: Li


title: Transcriptional shut-off of MAP kinase signaling enables pluripotency maintenance during diapause [PRO-seq]

first: Ryan

overall_design:
To generate the consensus gene annotations used for our RNA-seq and TT-seq analyses, we carried out the Precision Run-On Sequencing (PRO-seq) assay and used the 5’ end data for active TSS calling.
We used control wild-type ES cells treated with vehicle (DMSO) for 60min for the assay, with three biological replicates.


title: Transcriptional shut-off of MAP kinase signaling enables pluripotency maintenance during diapause


title: Transcriptional shut-off of MAP kinase signaling enables pluripotency maintenance during diapause [TT-seq]

overall_design:
Our study here based on bulk RNA-seq with time course dependent samples suggested the rapid gene expression changes occurring in ES cells upon mTOR inhibition by Torin1.
To further investigate the rapid gene expression changes in ES cells upon mTOR inhibition by Torin1, we performed TT-seq by using 4-thiouridine (4sU) labeled nascent RNA samples derived from time course dependent assays with Torin1 treatment including 15min, 30min and 60min. Three biological replicates were included in each condition of the assay.

first: Tuo


title: Effects of high cholesterol diet on sperm  small RNA expression profile utilizing two different RNA-seq methods

first: Changcheng

last: Zhou

summary:
Purpose: To  reveal the small RNA profiles in the sperm using traitional and PANDORA-seq methods
Methods: 3 week old LDLR-/- male mice were fed a high cholesterol or low cholesterol diet for 9 weeks. Sperm RNA was extracted via Trizol.
Results: paternal HCD feeding can lead to sperm tsRNA/rsRNA landscape changes that are undetected by traditional RNA-seq

overall_design: Sperm small RNA were sequenced from LDLR-/- male mice after 9 weeks of low cholesterol or high cholesterol feeding

first: Rebecca

last: Hernandez

first: Junchao

last: Shi

first: Tong


title: Aryl Hydrocarbon Receptor Ligands Present In The Environment Impact Human Trophoblast Cell Development

city: Stillwater,

first: Khursheed

last: Iqbal

state: OK

address: 310 N Monroe St,

institute: Oklahoma state university


title: Aryl Hydrocarbon Receptor Ligands Present In The Environment Impact Human Trophoblast Cell Development [EVT]

summary: TCDD do not activate AHR signaling in rat TS cells but do so in human TS cells with different biological outcomes. TCDD activates AHR signaling, stimulates the synthesis of 2 methoxy estradiol (2ME), a biologically active catechol estrogen, without directly affecting human TS cell differentiation. TCDD actions on human TS cells were dependent upon AHR signaling.

overall_design: TCDD was added as a supplement to the standard EVT cell differentiation culture conditions of CT27 human TS cells (Okae et al.). On day eight of differentiation, cells treated with either control or TCDD 10 nM were collected and RNA was isolated.

first: Vinay

last: Shukla

first: Michael

last: Soares


title: Aryl Hydrocarbon Receptor Ligands Present In The Environment Impact Human Trophoblast Cell Development [ST3D]

overall_design: TCDD was added as a supplement to the standard ST3D cell differentiation culture conditions of CT27 human TS cells (Okae et al.). On day six of differentiation, cells treated with either control or TCDD 10 nM were collected and RNA was isolated.


title: Single Cell Droplet-Based Efficacy and Transcriptomic Analysis of a Novel Anti-KLRG1 Antibody for Elimination of Autoreactive T Cells

first: Matthew

middle: Ryan

last: Sullivan

address: 140 Fenway

department: Pharmaceutical Sciences

institute: Northeastern University

summary: Progress in developing improvements in the treatment of autoimmune disease has been gradual, due to challenges presented by the nature of these conditions. Namely, the need to suppress a patient’s immune response while maintaining the essential activity of the immune system in controlling disease. Targeted treatments to eliminate the autoreactive immune cells driving disease symptoms present a promising new option for major improvements in treatment efficacy and side effect management. Monoclonal antibody therapies can be applied to target autoreactive immune cells if the cells possess unique surface marker expression patterns. Killer cell lectin like receptor G1 (KLRG1) expression on autoreactive T cells presents an optimal target for this type of cell depleting antibody therapy. In this study, we apply a variety of in vitro screening methods to determine the efficacy of a novel anti-KLRG1 antibody at mediating specific natural killer (NK) cell mediated antibody-dependent cellular cytotoxicity (ADCC). The methods include single-cell droplet microfluidic techniques, allowing timelapse imaging and sorting based on cellular interactions. Included in this study is the development of a novel method of sorting cells using a droplet-sorting platform and a fluorescent calcium dye to separate cells based on CD16 recognition of cell-bound antibody. We applied this novel sorting method to visualize transcriptomic variation between NK cells that are or are not activated by binding the anti-KLRG1 antibody using RNA sequencing. The data in this study reveals a reliable and target-specific cytotoxicity of the cell depleting anti-KLRG1 antibody, and supports our droplet-sorting calcium assay as a novel method of sorting cells based on receptor activation.

overall_design: Primary NK and transfected KRLG1+ CHO-K1 cells were co-encapsulated in droplets with anti-KLRG1 antibody. Droplets were sorted based on NK cell calcium release after CD16 binding of a-KLRG1 antibody, and the collected cell populations were submited to Novogene for sequencing.

middle: R

first: Aldo

last: Rozzo

first: Antonio

last: Ward

first: Patricia

middle: E

last: Rao

first: Dulce

last: Soler-Ferran

first: Steven

last: Greenberg

first: Tania

last: Konry


title: Environmental cadmium-induced circRNA-miRNA-mRNA network regulatory mechanism in human normal liver cell model [mRNA]

city: Zhengzhou

first: Panpan

last: Wang

address: 郑州市高新区科学大道100号郑州大学

department: College of Public Health

institute: 郑州大学

summary: Cadmium is a naturally existing heavy metal and a typical environmental pollutant which is a significant threat to human health and can lead to acute and chronic damage to various organs. At present, hundreds of lncRNAs and miRNAs have been confirmed to be related to the toxicity of Cd. However, the role of circRNA in the toxicity of Cd and the involved regulatory mechanism has not been clarified. In this work, human normal liver cell (L-02) was selected as model to identify the changes in the expression level of circRNA after exposure to Cd. Total RNA of each sample was extracted by Trizol method, the original data of circRNA, miRNA and mRNA expression levels of each sample were obtained by microarray hybridization and scanning were standardized and differentially expressed. Differential circRNAs, miRNAs, and mRNAs associated with the toxic effects of Cd were identified. Construct a ceRNA network by screening the predicted circRNA, miRNA and mRNA and predict the main biological functions and metabolic pathways of the network. A total of 266 different circRNAs, 223 different miRNAs and 519 different mRNAs were identified by differential expression analysis. After screening, seven circRNAs, 10 miRNAs and 97 mRNAs were constructed into ceRNA network. After GO enrichment and KEGG pathway analysis of 97 mRNAs in circRNA-miRNA-mRNA network, indicated that circRNA in the ceRNA network may regulate cell proliferation, apoptosis, oxidative stress and inflammatory response under the toxic effect of Cd to damage L-02, and it has obvious dose-response effect and time effect.

first: Zhi

last: Qu

first: Lugang

last: Deng

first: Chunqian

last: Feng

first: Peisen

last: Guo

first: Peixi

last: Liu


title: Environmental cadmium-induced circRNA-miRNA-mRNA network regulatory mechanism in human normal liver cell model [circRNA]

address: 郑州市高新区科学大道100号郑州大学

overall_design: In this work, human normal liver cell (L-02) was selected as model to identify the changes in the expression level of circRNA after exposure to Cd. Total RNA of each sample was extracted by Trizol method, the original data of circRNA, miRNA and mRNA expression levels of each sample were obtained by microarray hybridization and scanning were standardized and differentially expressed. Differential circRNAs, miRNAs, and mRNAs associated with the toxic effects of Cd were identified. Construct a ceRNA network by screening the predicted circRNA, miRNA and mRNA and predict the main biological functions and metabolic pathways of the network.


title: Influence of the WDR4 c.251G>A (p.R84H) variant on the transcriptome in human fibroblasts

city: Jena

first: Marco

last: Groth

address: Beutenbergstraße 11

department: Core Facility - Next Generation Sequencing

country: Germany

institute: Leibniz Institute on Aging - FLI

summary: The WD repeat-containing protein 4 (WDR4) has repeatedly been associated with primary microcephaly, a condition of impaired brain and skull growth, which may be due to faulty cilia. Here, we show using a combination of approaches in human fibroblasts, zebrafish embryos and patient-derived cells that WDR4 facilitates cilium formation. Molecularly, we associated WDR4 loss-of-function with increased protein synthesis and concomitant upregulation of proteasomal activity, while ubiquitin precursor pools are reduced. Inhibition of proteasomal activity as well as supplementation with free ubiquitin restored normal ciliogenesis. Proteasome inhibition ameliorated microcephaly phenotypes. Thus, we propose that WDR4 loss-of-function impairs head growth and neurogenesis via aberrant cilia formation, initially caused by disturbed protein and ubiquitin homeostasis.

overall_design:
RNA sequencing of human fibroblast cell line samples: 8 samples: 4 control samples (1BR3) and 4 WDR4-R84H samples

Submitter states that missing raw files are due to patient privacy concerns for the 4 WDR4-R84H samples.

first: Martin

last: Burkhalter

first: Melanie

last: Philipp


title: Transcriptional shut-off of MAP kinase signaling enables pluripotency maintenance during diapause [RNA-seq]

first: Ryan

last: Marina

overall_design:
The exposure of ES cells to incrementally increased BET protein inhibitor (I-BET) concentration results in the generation of diapause ES cells, hereafter defined as I-BET resistant (I-BETR) ES cells.
To investigate the gene expression patterns of I-BETR diapause ES cells, we performed the gene expression profiling analysis using data from bulk RNA-seq of 4 samples of control and 4 samples of I-BETR ES cells.
To understand the similarity of gene expression patterns in the I-BETR ES cells and other models of diapause ES cells, we also carried out the gene expression profiling analysis using data from bulk RNA-seq of 3 samples EBSS starvation induced diapause ES cells, 3 samples of Leucine and serum withdraw induced diapause ES cells and 3 samples of mTOR inhibitor Torin1 induced diapause ES cells comparing with 3 samples of their own Control ES cells.
Lastly, to understand the quick response in terms of gene expression changes upon mTOR inhibition in ES cells, we also performed gene expression profiling analysis using time course dependent samples in control and Torin1 treated ES cells including 30min, 60min, 120min and 360min.

first: Tuo

last: Zhang

first: Karen

last: Adelman


title: Aryl Hydrocarbon Receptor Ligands Present In The Environment Impact Human Trophoblast Cell Development [STEM]

first: Khursheed

last: Iqbal

overall_design: TCDD was added as a supplement to the standard STEM cell differentiation culture conditions of CT27 human TS cells (Okae et al.). Cells treated with either control or TCDD 10 nM (24 hr) were collected and RNA was isolated.

first: Vinay

last: Shukla

first: Michael

last: Soares


title: Environmental cadmium-induced circRNA-miRNA-mRNA network regulatory mechanism in human normal liver cell model

address: 郑州市高新区科学大道100号郑州大学

type:
Non-coding RNA profiling by array
Expression profiling by array


title: Environmental cadmium-induced circRNA-miRNA-mRNA network regulatory mechanism in human normal liver cell model [miRNA]

address: 郑州市高新区科学大道100号郑州大学


title: Spermidine-eIF5A axis is essential for muscle stem cell activation via translational control (RNA-Seq)

city: Beijing

first: Yong

address: 5 Dong Dan San Tiao

institute: School of Basic Medicine, Peking Union Medical College

summary:
Adult skeletal muscle stem cells, also known satellite cells (SCs), are quiescent and activate in response to injury. However, the activation mechanisms of quiescent SCs (QSCs) remain largely unknown. Here, we investigated the metabolic regulation of SC activation by identifying regulatory metabolites that promote SC activation. Using targeted metabolomics, we found that spermidine acts as a regulatory metabolite to promote SC activation and muscle regeneration in mice. Mechanistically, spermidine activates SCs via generating hypusinated eIF5A. Using SC-specific eIF5A-knockout (KO) and Myod-KO mice, we further found that eIF5A is required for spermidine-mediated SC activation by controlling MyoD translation. More significantly, depletion of eIF5A in SCs results in impaired muscle regeneration in mice. Together, the findings of our study define a novel mechanism that is essential for SC activation and acts via spermidine-eIF5A-mediated MyoD translation. Our findings suggest that the spermidine-eIF5A axis represents a promising pharmacological target in efforts to activate endogenous SCs for the treatment of muscular disease.
We then performed gene expression profiling analysis using data obtained from RNA-seq of primary myoblast from total muscle of WT and eIF5A KO mice.

overall_design: Primary myoblasts were isolated from the hind-limb muscles of adult (3-week-old) mice by digesting the muscle chops with 2 U/ml dispase and 0.2% collagenase for 45 min. The cells were filtered through 70-µm cell strainers and washed in PBS. Then the cells were resuspended in culture medium. The cells were collected after 2 days and the total RNA were extracted.

first: Qianying

first: Hu

last: Li

first: Dahai

last: Zhu


title: Spermidine-eIF5A axis is essential for muscle stem cell activation via translational control (Ribo-Seq)

summary: Adult skeletal muscle stem cells, also known satellite cells (SCs), are quiescent and activate in response to injury. However, the activation mechanisms of quiescent SCs (QSCs) remain largely unknown. Here, we investigated the metabolic regulation of SC activation by identifying regulatory metabolites that promote SC activation. Using targeted metabolomics, we found that spermidine acts as a regulatory metabolite to promote SC activation and muscle regeneration in mice. Mechanistically, spermidine activates SCs via generating hypusinated eIF5A. Using SC-specific eIF5A-knockout (KO) and Myod-KO mice, we further found that eIF5A is required for spermidine-mediated SC activation by controlling MyoD translation. More significantly, depletion of eIF5A in SCs results in impaired muscle regeneration in mice. Together, the findings of our study define a novel mechanism that is essential for SC activation and acts via spermidine-eIF5A-mediated MyoD translation. Our findings suggest that the spermidine-eIF5A axis represents a promising pharmacological target in efforts to activate endogenous SCs for the treatment of muscular disease.

overall_design: Primary myoblasts were isolated from the hind-limb muscles of adult (3-week-old) mice by digesting the muscle chops with 2 U/ml dispase and 0.2% collagenase for 45 min. The cells were filtered through 70-µm cell strainers and washed in PBS. Then the cells were resuspended in culture medium. Cells were treated CHX at 100 μg/ml for 1 min and collected, and 5 x 10 6 cells were lysed in lysis buffer (5 mM MgCl2, 10% Triton X-100, Tris-HCl pH 7.5, and 150 mM NaCl) supplemented with 1 U/μl DNase Ⅰ, 50 μg/ml CHX, and 100 mM dithiothreitol (DTT) for 30 minutes on ice. Cell lysates were  centrifuged at 18,000 x g at 4 ℃ for 15 min and the supernatant was digested with RNase Ⅰ.Ribosome-protected RNA fragments (RPF) were obtained using MicroSpin S-400 column resin; the was column enriched for ribosome-mRNA complexes and the targeted RNA fragments were purified by PAGE. The purified RNA fragments were end-repaired and A-tailed, and 5’ and 3’ joints were directly added to the ends. RNA reverse transcriptase was used to synthesize cDNA, and additional steps of PCR enrichment and PAGE-based screening of target fragments were used to obtain a library containing target fragments. The library was verified by quality control screening and sequencing.


title: Spermidine-eIF5A axis is essential for muscle stem cell activation via translational control (scRNA-Seq)

overall_design: QSCs were freshly FACS (Fluorescence-activated cell sorting)-isolated from uninjured TA muscles of 8-week-old Pax7 CreERT2/+ ;Rosa26 mTmG/+ mice,  ASCs were FACS-isolated from injured TA muscles on day 1.5 after CTX injection. QSCs and ASCs were soreted and analyed using scRNA-seq.


title: Spermidine-eIF5A axis is essential for muscle stem cell activation via translational control

overall_design: Refer to individual Series


title: Single-cell transcriptome profiling implicates the psychological stress-induced disruption of spermatogenesis

city: Xi'an, Shaanxi Province

first: Yungang

last: Xu

state: Shaanxi

address: Xi'an, Shaanxi Province

institute: Xi'an Jiaotong University

summary: Male infertility has become a global problem. And psychological stress has been proven to be an important cause of male infertility. In this work, we constructed a terrified-sound stress model in SD rats and performed single-cell RNA sequencing of the testicular tissues from the post-stressed and normal rats. Through the comparative analysis of the gene expression patterns of the 11,744 rat testicular cells, we revealed the potential mechanism by which terrified-sound induced psychological stress affects spermatogenesis and ultimately leads to reproductive impairment in male rats. This will provide a theoretical basis for the study of psychological stress on male reproductive impairments for other animals and humans, and further find the therapeutic targets for male infertility caused by psychological stress.

overall_design: We constructed a terrified-sound stress model in SD rats and performed single-cell RNA sequencing of the testicular tissues from 3 post-stressed rats and 3 normal rats.

first: Rufeng

first: Yuefeng

last: Du

first: Kang

first: Xiaofan

last: Xiong

first: Lingyu

first: Chen

first: Shanfeng

last: Gao

first: Yufei

last: Yao

first: Juan

last: Yang


title: Molecular subtypes are associated with clinical benefit in cisplatin-treated metastatic urothelial cancer patients

city: Lund

first: Gottfrid

last: Sjodahl

address: Medicon Village

department: Department of Translational Medicine

country: Sweden

institute: Lund Univeristy

summary:
Cisplatin-based combination chemotherapy (CHT) is standard of care in locally advanced and metastatic urothelial cancer (mUC). No predictive biological biomarkers are available for clinical use. Here, we investigate the impact of molecular subtypes in relation to response and survival in mUC patients treated with first-line CHT. Molecular subtype classification according to Lund taxonomy was performed by tumour transcriptomic profiling and immunostaining in a retrospective cohort with mUC patients treated with CHT. We analyze overall response rate (ORR) and secondary endpoint overall survival (OS) in comparison to prespecified molecular subtypes. Differential gene expression and association to treatment response was explored.
Ninety-five patients with mUC were classified into urothelial (Uro, 43%), genomically unstable (GU, 26%), basal/squamous (Ba/Sq, 20%), mesenchymal like (Mes-like, 8%) and small-cell/neuroendocrine-like (Sc/NE, 3%) subtypes. Gene expression data is available for 86 patients. Patients with Mes-like tumours had significantly lower ORR (14%), compared to Uro (70%), GU (77%), Ba/Sq (75%) and Sc/NE, 67%), p = 0.020. Further, patients with the Mes-like subtype had significantly shorter mOS (HR 3.13, 95% CI: 1.37-7.13, p = 0.007), with Uro as reference. An enrichment of down-regulated stromal- and immune-related genes was seen in responders. Down-regulation of interferon-induced transmembrane protein 2 (IFITM2), an inflammation related gene, was associated with increased ORR and improved OS. In patients treated with CHT for mUC, the Mes-like subtype showed significantly lower response rate and shorter overall survival. Down-regulation of IFITM2 was associated with clinical benefit, suggesting the presence of IFITM2 to be a possible unfavourable prognostic marker in mUC.

overall_design: Retrospective cohort study. Note that the majority but not all samples are re-analysis of samples present in previous GEO-submissions (see description).

first: Karin

last: Holmsten

last: Sjödahl

first: Johan

last: Abrahamsson

first: Pontus

last: Eriksson

first: Carina

last: Bernardo

first: Mattias

last: Höglund

first: Fredrik

last: Liedberg

first: Anders

last: Ullén


title: Spermidine-eIF5A axis is essential for muscle stem cell activation via translational control (Polysome-Seq)

first: Yong

last: Zhang

first: Qianying

first: Hu

last: Li

first: Dahai


title: Transcriptome analysis of ZCCHC7  wildtype(WT) and overexpression (OE) in colorectal cancer

city: nanjing

first: xian

last: luo

address: xianlin

institute: NJUCM

summary: Transcriptome sequencing was used to analyze changes in related signaling pathways and gene expression in colon cancer cells when ZCCHC7 was overexpressed .

overall_design: Examination of  two cell types

first: Xian

last: Luo

first: Mengjie

last: Guo


title: Neuronal stimulation changes U1 snRNP binding and intronic polyadenylation

city: Baltimore

first: Lisa

middle: Nicole

last: Learman

address: 725 N. Wolfe St., PCTB 917

department: Neuroscience

institute: Johns Hopkins University School of Medicine

summary: Synaptic activity induces well-known changes in enhancer-promoter driven gene expression but also induces changes in splicing and polyadenylation that are understudied. Here, we investigate the mechanism of expression for alternative polyadenylation isoform Homer1a, an immediate early gene essential to synaptic plasticity. We report that neuronal activation, in neuronal cultures and in adult mouse brain, depletes the splice factor U1 snRNP from Homer1 pre-mRNA and that this causes shifted utilization of a cryptic polyadenylation signal within intron 5 resulting in Homer1a expression. Because U1 snRNP is a ubiquitous splice factor, we tested the generality of activity-driven U1 snRNP depletion as a mechanism for gene expression using RNA immunoprecipitation sequencing. Analysis reveals that neuronal activity changes U1 snRNP binding to ~2000 transcripts and for a subset of transcripts, a reduction in U1 snRNP binding was accompanied by utilization of a cryptic intronic polyadenylation site. This subset is enriched for transcripts encoding synaptic proteins involved in excitability control. Genes demonstrating activity-dependent reduced U1 snRNP binding often encode a binding motif for Sam68, a neuronal alternative polyadenylation factor. Findings reveal that activity-driven changes in intron utilization for transcript termination serves an important role in synaptic plasticity.

overall_design: RNAseq and U1 snRNP CLIPseq profiles from brain of mice with and without neuronal stimulation

middle: N

first: Raozhou

last: Lin

first: Paul

middle: F

last: Worley


title: Effect of inhibiting Neurl1b in repopulating hepatocytes in Fah-/-;dCas9 liver injury and repopulation model

status: Public on Oct 07 2024

city: Rochester

first: Dingzi

last: Yin

state: Minnesota

address: 222 3rd Ave SW

institute: Mayo Clinic

summary: To study the mechanism of NEURL1B in regulating hepatocyte mitosis, we profiled the ribosome bounded mRNA of control and Neurl1b-inhibited repopulating hepatocytes by using TRAP-seq (translating ribosome affinity purification followed by high-throughput RNA sequencing)

overall_design: To inverstigate the effect of inhibiting Neurl1b on repopulating hepatocytes, we expressed FAH, GFP-L10a fusion protein and sgRNA agasint Neurl1b in Fah-/-;dCas9 hepatocytes, we then profiled ribosome bounding mRNAs by TRAP-seq (translating ribosome affinity purification followed by high-throughput RNA sequencing)

first: Kirk

last: Wangensteen


title: MH-seq data of bread wheat

city: Nanjing

first: dongyang

last: Zheng

state: Jiang su

address: NJAU Wei gang No.1

department: college of agriculture

institute: Nanjing Agricultural University

summary: Identification of open chromatin in wheat spikes and seedlings using MH-seq

overall_design: MH-seq (MNase hypersensitivity sequencing) of seedling and spikelet in bread wheat

first: Dongyang

first: Wenli

first: Yijing


title: Selumetinib in adults with NF1 and inoperable plexiform neurofibroma

city: bethesda

first: Jack

last: Shern

address: 10 Center drive

institute: NCI

summary: The MEK inhibitor selumetinib induces objective responses and provides clinical benefit in children with neurofibromatosis type 1 (NF1) and inoperable plexiform neurofibromas (PNs).  To evaluate whether similar outcomes were possible in adult patients, in whom PN growth is generally slower than in pediatric patients, an open-label phase 2 study of selumetinib in adults with NF1 PNs was conducted. Correlative analysis included extraction of RNA followed by sequencing. Samples include paired specimens collected before and on treatment.

overall_design: RNA-seq was performed for gene expression profiling analysis in tumors collected before and on treatment with the MEK inhibitor Selumetinib.

first: Brigitte

last: Widemann

first: Steve

last: Angus

first: Haiyan

last: Lei


title: Effects of running for 1h on gene expression in Achilles tendon cells of SD rats

city: Zhuhai

first: Dantian

last: Zhu

address: No. 52, Meihua East Road, Meihua Street, Xiangzhou District, Zhuhai City, Guangdong Province, China

institute: Department of Radiology, The Fifth Affiliated Hospital of Sun Yat-sen University, Zhuhai

summary: The Achilles tendon is the thickest tendon in the human body, and Achilles tendinopathy is its most prevalent disorder, often considered a consequence of overuse. While disruption to the collagen fibers represents a significant manifestation of Achilles tendinopathy, strikingly little is known about the mechanisms by which healthy tendon accumulates damage in vivo.As existing studies of tendon biomechanics and mechanobiology predominantly relied on in vitro or ex vivo experiments on isolated tissues, it is still largely unknown whether and how disruptions occur to the collagen molecules in healthy Achilles tendons following physiological activities. We reported the first RNA-seq analysis reflecting transcriptome changes in healthy rat Achilles tendons following running, providing a resource for future investigations in tendon mechanobiology and sports medicine.

overall_design: Rats in the running group (0 h p.r. group)  were sacrificed immediately after 1 h of treadmill running. Fresh Achilles tendon tissues from the right legs of the running and naïve control  (NC group) rats were immediately dissected and placed in RNALater™ to prevent RNA degradation. The total RNA was extracted after the tissues were mechanically pulverized in TRIzol. After verification of the RNA integrity by Qsep100 (RNA integrity number > 7.0), polyA mRNAs was purified from total RNA using mRNA capture beads. Then the mRNA was reverse-transcribed to create a cDNA library. VAHTS Stranded mRNA-seq Library Prep Kit for Illumina V2 was used for library preparation. RNA sequencing was performed using the Illumina NovaSeq 6000 sequencer. We then performed gene expression profiling analysis using data obtained from RNA-seq of 2 different tissues at two groups.Comparative gene expression profiling analysis of RNA-sea data for 0 h p.r. group and NC group.

first: Yijie

last: Fang

first: Man

first: Shaolin

first: Yang


title: Gene expression in Achilles tendon cells of SD rats after 1 h of running followed by 12 h of rest

summary: Achilles tendinopathy is often thought to be a consequence of overuse of the Cells within the Achilles tendon of healthy rats undergo a series of changes following physiologic levels of mechanical stimulation after running, and we further explored the transcriptome changes in Achilles tendon cells during post-exercise recovery. Our current experiment reveals RNA-seq analysis of the transcriptome of the rat Achilles tendon after 12 hours of rest following running.

overall_design: Rats in the running group (12 h p.r. group) sacrificed 12 h after 1 h of treadmill running. Control rats rested in cages and were sacrificed at the same time as the running group. Fresh Achilles tendon tissues from the right legs of the running and naïve control rats were immediately dissected and placed in RNALater™ to prevent RNA degradation. The total RNA was extracted after the tissues were mechanically pulverized in TRIzol. After verification of the RNA integrity by Qsep100 (RNA integrity number > 7.0), polyA mRNAs was purified from total RNA using mRNA capture beads. Then the mRNA was reverse-transcribed to create a cDNA library. VAHTS Stranded mRNA-seq Library Prep Kit for Illumina V2 was used for library preparation. RNA sequencing was performed using the Illumina NovaSeq 6000 sequencer. We then performed gene expression profiling analysis using data obtained from RNA-seq of 2 different tissues at two time points. Comparative gene expression profiling analysis of RNA-sea data for 0 h p.r. group and NC


title: Single-cell RNA sequencing of immune cells from murine GBM tumors

city: Daejeon

first: Juhee

last: Lim

address: 291 Daehak-ro

country: South Korea

institute: KAIST

summary: We analyzed effects of IFNAR deficiency on immune cells of the GBM TME and also analyzed how anti-PD1 affected these immune cells in absence of type I IFN signaling. We also analyzed the effects of anti-PD1 treatment on the immune cell populations in absence of  IFN production.

overall_design: CD45.2+ tumor-infiltrating immune cells were sorted from brains of WT and IFNaR-/- mice 20 days following Gl261 intracranial injection. In addition, in order to analyze the effects of ICB therapy on immune cells in absence of IFN signaling, anti-PD1 treatment was intraperitoneally injected at a dose of 200 ug/mouse on days  7,11,15,19 following Gl261 injection. CD45.2+ immune cells were sorted from the brains of anti-PD1 treated WT and IFNaR-/- mice 25 days following Gl261 injection.

first: Jeongwoo

last: La

first: Hyeon

middle: C

last: Kim

first: In

last: Kang

first: Byeong

middle: H

first: Keung

last: Ku

first: Yumin

first: Myoung

middle: S

last: Kwon

first: Heung

last: Lee


title: INO80 regulates chromatin accessibility to facilitate suppression of sex-linked gene expression during mouse spermatogenesis

city: Chapel Hill

first: Prabuddha

last: Chakraborty

state: NC

address: 120 Mason Farm Road5023 B, Genetic Medicine Building, CB#7264

department: Genetics

institute: University of North Carolina at Chapel Hill

summary: The INO80 protein is the main catalytic subunit of the INO80-chromatin remodeling complex, which is critical for DNA repair and transcription regulation in murine spermatocytes. In this study, we explored the role of INO80 in silencing genes on meiotic sex chromosomes in male mice. INO80 immunolocalization at the XY body in pachytene spermatocytes suggested a role for INO80 in the meiotic sex body. Subsequent deletion of Ino80 resulted in high expression of sex-linked genes. Furthermore, the active form of RNA polymerase II at the sex body of Ino80-null pachytene spermatocytes indicates incomplete inactivation of sex-linked genes.  A reduction in the recruitment of initiators of meiotic sex chromosome inhibition (MSCI) argues for INO80-facilitated recruitment of DNA repair factors required for silencing sex-linked genes. This role of INO80 is independent of a common INO80 target H2A.Z. Instead, in the absence of INO80, a reduction in chromatin accessibility at DNA repair sites occurs on the sex chromosomes. These data suggest a role for INO80 in DNA repair factor localization, thereby facilitating the silencing of sex-linked genes during the onset of pachynema.

overall_design: CUT&RUN was performed for MDC1 from spermatogenic cells obtained from synchronized Ino80WT and Ino80cKO (testis specific Ino80-deletion generated with Stra8-Cre) testes at pachynema stage.

first: Terry

last: Magnuson


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

city: guangzhou

first: Yongbo

last: Pan

address: 106 Zhongshan 2 Lu, Guangzhou

department: Guangdong Provincial People's Hospital

institute: Guangdong Academy of Medical Sciences

summary: To reveal the potential m6A modification gene  in  Adenocarcinoma of the esophagogastric junction, we performed methylated RNA immunoprecipitation and sequencing  (meRIP-seq) in OE-19 cells.

overall_design: OE-19 cells lysate was Immunoprecipitation by m6A antibody, then subjected to RNA-sequencing analysis


title: Difference analysis of microRNAs in peripheral blood of  asthmatic children by high-throughput sequencing

city: Shenyang

first: Bing

last: Wei

address: No.83 Wenhua Road

institute: General Hospital of Northern Theater Command

summary: Asthma is the most common chronic pulmonary disease in children. The pathogenesis of asthma is not fully understood,and  existing treatments have little effect on some children.MicroRNAs(miRNAs) play a regulatory role in the occurrence and development of asthma,and miRNA therapy is a promising treatment for asthma.The purpose of this study was to explore the differential expression of miRNAs in peripheral blood of children with asthma and to find a miRNA that can alleviate  asthma inflammation.We used high-throughput sequencing to analyze miRNA differences in peripheral blood between children with acute asthma attacks and healthy children,target gene prediction and functional enrichment analysis were performed for them..Overall,51 differentially expressed miRNAs were identified,with 9 being upregulated and 42 downregulated.

overall_design: To explore the differences in miRNA in peripheral blood of children with asthma and healthy children, we collected venous blood samples of children with asthma and healthy children, extracted total RNA, and analyzed them using high-throughput sequencing.


title: Differential usage of shared binding sites drives functional specialization of an essential family of transcription factors

city: Philadelphia

first: Aman

last: Husbands

address: 103B Caroline Lynch Bldg, 433 S University Ave

institute: University of Pennsylvania

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


title: NSD2 Overexpression in Myeloma Enhances Dependency on Mitochondrial High Energy Phosphate Transport Facilitated by Adenylate Kinase 2.

city: Gainesville

first: Alberto

last: Riva

state: FL

address: 2033 Mowry Rd

department: ICBR

institute: University of Florida


title: NSD2 Overexpression in Myeloma Enhances Dependency on Mitochondrial High Energy Phosphate Transport Facilitated by Adenylate Kinase 2 [AS_RS]

summary: Chromosomal translocation t(4;14), a driver of NSD2 overexpression, remains a negative prognostic factor in multiple myeloma (MM). A genome-wide CRISPR screen in MM cells identified adenylate kinase 2 (AK2) as an NSD2-driven vulnerability. AK2 depletion in t(4;14) MM cells inhibited cell growth in vitro and in vivo, impaired protein folding in the ER  and increased sensitivity to proteasome inhibition, consistent with a defect in shuttling ATP from the mitochondria. Moreover, AK2 suppression decreased intracellular NADP(H) critical for thioredoxin reduction, which is key for deoxyribonucleotides (dNTP) synthesis, leading to DNA replication stress and apoptosis. The increased dependence on AK2 was attributed to insufficient creatine production due to NSD2-mediated diversion of one-carbon metabolism to the epigenome and away from S-adenosylmethionine dependent creatine synthesis. Correspondingly, supplementation with creatine restored NADP(H) levels and rescued AK2-deficient cells from apoptosis. These findings revealed a novel metabolic susceptibility and promising therapeutic strategy for targeting t(4;14) MM.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for KMS11 NTKO and TKO cells wit or without AK2 knockdown (shAK2 vs. shSc)

first: Amin

last: Sobh

first: Elena

last: Encinass

first: Alisha

last: Patel

first: Greeshma

last: Surapaneni

first: Charlotte

last: Kaestner

first: Janai

last: Poullard

first: Monica

last: Clerio

first: Karthik

last: Vasan

first: Tzipporah

last: Freeman

first: Dongwen

last: Lv

first: Benjamin

middle: G

last: Barwick

first: Daohong

last: Zhou

first: Lawrance

last: Boise

first: Constantine

last: Mitsiades

first: Baek

first: Richard

last: Bennett

first: Navdeep

last: Chandel

first: Jonathan

last: Licht


title: Effect of overexpression of MEG3 on gene expression in prostate cancer lncap cells

city: Wuhan

first: Gang

last: Luo

state: Hubei

address: 26 Shengli Street

institute: Huazhong University of Science and Technology

summary: Long non-coding RNA MEG3 is widely involved in tumor development. However, its role in prostate cancer is not very clear. We overexpressed MEG3 in prostate cancer lncap cells and characterized the effect of MEG3 on gene expression profiles by RNA sequencing

overall_design: MEG3 overexpression vector was transfected into lncap cells and RNA sequencing identified changes in gene expression profiles.The experiment was replicated three times


title: Determining the Injury-Associated Microbial Nutrients that Facilitate Secondary Bacterial Infection in Acutely Injured Lungs

status: Public on Oct 06 2024

city: Ann Arbor

first: Piyush

last: Ranjan

state: Michigan

address: 1150 W Medical Center Dr

department: Internal Medicine - Pulmonary and Critical Care

institute: University of Michigan

summary: The goal of this experiment was to determine the changes in the transcriptome of Pseudomonas aeruginosa when cultured in bronchoalveolar lavage fluid (BALF) from mice with oxygen-induced lung injury relative to healthy controls.

overall_design: Adult C57BL/6 mice were sacrificed following exposure to hyperoxia (FiO2 95% for 72 hours) or room air (FiO2 21% for 72 hours), BALF was collected and centrifuged to remove host and microbial cells, and stored at -80 C. Upon thawing, BALF was pooled within exposure group, divided into 1 mL aliquots, inoculated with Pseudomonas aeruginosa strain PAO1, and incubated at 37 C shaking to mid-log phase (10 hours). Lennox Broth was used as a positive control for the experiment. Bacterial cell pellets were collected for RNA isolation and bulk RNA-sequencing.


title: Effect of overexpression of LINC01518 on gene expression in prostate cancer lncap cells

summary: LINC01518 is differentially expressed in prostate cancer, but the exact role and potential mechanisms are unknown. We overexpressed LINC01518 in prostate cancer lncap cells and examined the effect of LINC01518 on gene expression profiles by RNA sequencing


title: Single-cell transcriptomics of female fetal mouse ovarian cells

city: Guangzhou

first: manman

last: cui

address: Baiyun District

institute: Southern Medical University

summary: In mammal, female reproductive health is largely related to fetal ovarian development, along with tight coordination of transcriptional and metabolic regulations, as well as cell-cell communications. Mice is an ideal modal for the understanding of network mechanism in this process, thus providing essential information on the diagnosis and therapy of female reproductive diseases, as well as on oogenesis in vitro. To plot the landscape of mouse fetal ovarian development, STRT-seq was applied for ovarian cells isolated from E11.5 to E18.5 female gonads using Blimp1-mVenus and Stella-ECFP (BVSC) mice. Based on the expression patterns of well-known marker genes, 9 germ cell clusters and 8 somatic cell clusters were annotated, presenting a continuous distribution across fetal ovarian development. To summarize, we constructed a systematic cell-fate transition of mouse female fetal ovarian cells at single-cell resolution, including stage-specifically activated TFs, cellular metabolism pathway, and cell–cell communication network in ovarian microenvironment.

overall_design: A single cell transcriptome profile of mouse ovarian cells from E11.5 to E18.5 were generated by next generation sequencing using Illumina HiSeq X-Ten.

first: Manman

last: Cui

first: Ping

last: Lu


first: Prabuddha

last: Chakraborty

summary: The INO80 protein is the main catalytic subunit of the INO80-chromatin remodeling complex, which is critical for DNA repair and transcription regulation in murine spermatocytes. In this study, we explored the role of INO80 in silencing genes on meiotic sex chromosomes in male mice. INO80 immunolocalization at the XY body in pachytene spermatocytes suggested a role for INO80 in the meiotic sex body. Subsequent deletion of Ino80 resulted in high expression of sex-linked genes. Furthermore, the active form of RNA polymerase II at the sex chromosomes of Ino80-null pachytene spermatocytes indicates incomplete inactivation of sex-linked genes. A reduction in the recruitment of initiators of meiotic sex chromosome inhibition (MSCI) argues for INO80-facilitated recruitment of DNA repair factors required for silencing sex-linked genes. This role of INO80 is independent of a common INO80 target, H2A.Z. Instead, in the absence of INO80, a reduction in chromatin accessibility at DNA repair sites occurs on the sex chromosomes. These data suggest a role for INO80 in DNA repair factor localization, thereby facilitating the silencing of sex-linked genes during the onset of pachynema.

overall_design: ATAC-seq was performed on spermatogenic cells obtained from Ino80WT testes.

first: Terry

last: Magnuson


title: TREX1 is required for microglial cholesterol homeostasis and subsequent oligodendrocyte maturation during brain development [scRNA-seq]

city: Carlsbad

first: Tom

last: Chen

address: 6155 Corte Del Cedro

department: R&D

institute: Universal Sequencing Technology Corp.

summary: Three Prime Repair Exonuclease 1 (TREX1) gene mutations have been associated with Aicardi-Goutières Syndrome (AGS) - a rare, severe pediatric autoimmune disorder that primarily affects the brain and has a poorly understood etiology. Microglia are brain-resident macrophages indispensable for brain development and implicated in multiple neuroinflammatory diseases. However, the role of TREX1 - a DNase that cleaves cytosolic nucleic acids, preventing viral- and autoimmune-related inflammatory responses - in microglia biology remains to be elucidated. Here, we leverage a model of human embryonic stem cell (hESC)-derived engineered microglia-like cells, bulk, and single-cell transcriptomics, optical and transmission electron microscopy, and three-month-old assembloids composed of microglia and oligodendrocyte-containing organoids to interrogate TREX1 functions in human microglia. Our analyses suggest that TREX1 influences cholesterol metabolism, leading to an active microglial morphology with increased phagocytosis in the absence of TREX1. Notably, regulating cholesterol metabolism with an HMG-CoA reductase inhibitor, FDA-approved atorvastatin, rescues these microglial phenotypes. Functionally, TREX1 in microglia is necessary for the transition from gliogenic intermediate progenitors known as pre-oligodendrocyte precursor cells (pre-OPCs) to precursors of the oligodendrocyte lineage known as OPCs, impairing oligodendrogenesis in favor of astrogliogenesis in human assembloids. Together, these results suggest routes for therapeutic intervention in pathologies such as AGS based on microglia-specific molecular and cellular mechanisms.

overall_design: Samples represent five independent preps of 100-day-old assembloids of microglia co-cultured with oligodendrocyte-containing cortical neural organoids. Microglia and organoids were generated from the H9 human embryonic stem cell (hESC) line. Two  microglia genotypes were tested: control (WT) and CRISPR-edited TREX1 mutant (KO) microglia. Three independent assembloid preps were generated (1, 2, and 3). We used AmpliDrop single-cell RNA-seq (scRNA-seq) technology to characterize these assembloids and interrogate the role of human TREX1 variants in early brain development.

first: Gabriela

last: Goldberg

first: Alysson

last: Muotri

first: Ivan

last: Garcia-Bassets


title: Specific features of the transcriptomic response to nitrogen starvation in methylotrophic yeast Komagataella phaffi

city: Saint Petersburg

first: Anton

middle: V.

last: Sidorin

address: 7/9 Universitetskaya nab., St. Petersburg, Russia

department: Faculty of Biology

country: Russia

institute: St. Petersburg State University

summary: Non-conventional methylotrophic yeast Komagataella phaffii is an important production host in biotechnology and an emerging model organism. In this work, we studied K. phaffii response to nitrogen starvation during cultivation in media with methanol as the sole carbon source. The results were compared with well-established model yeast Saccharomyces cerevisiae. Some of the observed effects of nitrogen starvation in K. phaffii were similar to those in S. cerevisiae, despite this yeast does not have metabolic pathway for methanol utilization. The effects include activation of autophagy, transport and catabolism of nitrogen-containing compounds, interconversions of amino acids, and biosynthesis of fatty acids. K. phaffii cells also demonstrated specific response to nitrogen starvation including suppression of genes involved in methanol metabolism and other peroxisomal processes and activation of purine catabolism genes.

overall_design: To investigate the impact of nitrogen starvation on K. phaffii we analyzed the transcriptome from three control replicates grown on the methanol-containing medium with a nitrogen source (ammonium sulfate) and three experimental replicates grown on the methanol-containing medium without a nitrogen source.

first: Anastasiya

last: Makeeva

middle: V

first: Valeria

last: Ishtuganova

first: Elena

last: Sambuk

first: Marina

last: Padkina

first: Andrey

middle: M

last: Rumyantsev


title: NSD2 Overexpression in Myeloma Enhances Dependency on Mitochondrial High Energy Phosphate Transport Facilitated by Adenylate Kinase 2 [AS_CR]

first: Alberto

last: Riva

overall_design: CRISPR Screens/comparison of relative sgRNA abundance between initial and final timepoints in KMS11 NTKO and TKO cells

first: Amin

last: Sobh

last: Encinass

first: Alisha

last: Patel

first: Greeshma

last: Surapaneni

first: Charlotte

last: Kaestner

first: Janai

last: Poullard

first: Monica

last: Clerio

first: Karthik

last: Vasan

first: Tzipporah

last: Freeman

first: Dongwen

last: Lv

first: Benjamin

last: Barwick

first: Daohong

last: Zhou

first: Lawrance

last: Boise

first: Constantine

last: Mitsiades

first: Baek

last: Kim

first: Richard

middle: L

last: bennett

first: Navdeep

last: Chandel

first: Jonathan

last: Licht


title: Vitamin D signaling orchestrates skeletal muscle metabolic flexibility by modulating its fuel choice

city: New Delhi

first: G Aneeshkumar

last: Arimbasseri

state: Delhi

address: Aruna Asaf Ali Marg

department: Molecular Genetics Laboratory

country: India

institute: National Institute of Immunology

summary: To understand the transcriptome wide changes in the skeletal muscles of mice lacking vitamin D receptor (VDR), we isolated quad muscles from these mice and their WT littermates. Total RNAseq was performed to check the changes in transcriptome. We show that the muscles of vdr-/- mice exhibit upregulation of mRNAs coding for fatty acid oxidation.

overall_design: WT and vdr-/- QUAD muscles were isolated from 7 weeks old mice. Total RNA was isolated and subjected to library preparation and sequencing using TruSeq total RNA sequencing kit from Illumina. Differential expression was performed followed by GO analysis and GSEA analysis.

first: Das

last: Anamica

first: Arimbasseri

last: Aneeshkumar


title: Ileal Crohn’s disease exhibits reduced activity of phospholipase C-β3 (PLC-β3)-dependent Wnt/b-catenin signaling pathway [ATAC-Seq]

city: La Jolla

first: Ikuo

last: Takazawa

address: 9420 Athena Cir

institute: La Jolla Institute for Immunology

summary: Crohn’s disease is a chronic, debilitating inflammatory bowel disease. Here we report a  critical role for phospholipase C-β3 (PLC-β3) in intestinal homeostasis. In PLC-β3- deficient mice, exposure to oral dextran sodium sulfate induced lethality and severe  inflammation in the small intestine. The lethality was due to PLC-3 deficiency in multiple  non-hematopoietic cell types. PLC-β3 deficiency resulted in reduced Wnt/-catenin signaling, which is essential for homeostasis and regeneration of the intestinal epithelium.  PLC-β3 regulated the Wnt/β-catenin pathway in small intestinal epithelial cells (IECs) at  transcriptional, epigenetic, and potentially protein-protein interaction levels. PLC-β3- deficient IECs were unable to respond to stimulation by R-spondin 1, an enhancer of  Wnt/β-catenin signaling. Reduced expression of PLC-β3 and its signature genes was found  in biopsies of ileal Crohn’s disease patients. PLC-β regulation of Wnt signaling was  evolutionally conserved in Drosophila. Our data indicate that reduction of PLC-β3- mediated Wnt/β-catenin signaling contributes to the pathogenesis of ileal Crohn’s disease.

overall_design: To investigate changes of the chromatin accessibility caused by the loss of PLC-b3, we performed ATAC-seq expreiment using PLC-β3 deficient (KO) or PLC-β3 sufficient scramble sgRNA treated (Scr) mouse colorectal cancer cell line (CMT-93)

first: Tomoaki

last: Ando

first: Zach

last: Spencer

first: Ryoji

last: Ito

first: Yoshiaki

last: Tomimori

first: Zbigniew

last: Mikulski

first: Kenji

last: Matsumoto

first: Tohru

last: Ishitami

first: Lee

middle: A

last: Denson

first: Yu

last: Kawakami

first: Yuko

first: Jiro

last: Kitaura

first: Yashi

last: Ahmed

first: Toshiaki


title: Ileal Crohn’s disease exhibits reduced activity of phospholipase C-β3 (PLC-β3)-dependent Wnt/b-catenin signaling pathway [16S rRNA]

overall_design: To investigate changes of the microbiome caused by the loss of PLC-β3, we corrected feces from wild type (WT), PLC-β3-deficient or fostered WT mice with or without DSS treatment and performed 16s ribosomal RNA (rRNA) sequencing.


title: Ileal Crohn’s disease exhibits reduced activity of phospholipase C-β3 (PLC-β3)-dependent Wnt/b-catenin signaling pathway

sample_organism:
Mus musculus
feces metagenome

platform_organism:
Mus musculus
feces metagenome

overall_design: Refer to individual Series


title: PHF8 Demethylates YY1 to Regulate Electron Transfer Chain Gene Transcription and License Mitochondrial Reactive Oxygen Species Production to Promote Cancer Cell Growth

city: Xiamen

first: Jiancheng

last: Ding

state: Fujian

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

institute: Xiamen University

summary: Emerging evidence suggest that mitochondrial reactive oxygen species (mROS) production is tightly controlled by electron transport chain (ETC), and increased levels of ROS production is linked to cancer development. The ETC therefore represents a promising way of cancer treatment. However, the epigenetic mechanisms governing the transcription of ETC genes to drive mROS production and cell proliferation remain to be fully characterized in cancer cells. Here, we reported that, while well-known to use histone substrates to activate transcription, PHF8, a PHD Finger and Jumonji C (JmjC) domain-containing histone demethylase, also functions as a transcriptional co-repressor for a large set of ETC genes, driving mROS production and promoting growth of cancer cells. PHF8 repression of ETC gene transcription is mediated through its demethylation of the transcription factor Yin and Yang 1 (YY1) at a highly conserved lysine (K) residue, K258. Using a structure-based virtual screening combined with demethylase activity assays, we identified a potent and specific small molecule inhibitor, iPHF8, which targets the catalytic core of PHF8 and is effective in regulating the transcription of ETC genes and mROS production in multiple types of cancer cells. Furthermore, iPHF8 is potent in suppressing cancer cell growth in vitro and tumor growth in cell line- and patient-derived xenografts in mice. Taken together, our data uncover a key epigenetic mechanism underlying transcriptional regulation of ETC genes and mROS production, which involves PHF8 demethylation of its non-histone substrate YY1, and demonstrated that targeting PHF8 has great potential to treat cancers.

overall_design: ChIP-seq performed in this study was designed to understand the molecular mechanisms underlying chromatin binding of YY1 regulated by PHF8

first: Wen

last: Liu

first: Xiao-Nan

last: Wu

first: Jian-Chen


title: Encoding of odors by mammalian olfactory receptors

city: Durham

first: Aashutosh

last: Vihani

address: 264 CARL Building

institute: Duke University

summary: Deorphanization of mouse olfactory receptors using pS6-IP-Seq.

overall_design: Phosphorylated S6 ribosomal subunit capture and associated mRNA profiling (pS6-IP-Seq) on whole olfactory mucosa.

first: Hiroaki

last: Matsunami


title: LNCRNA expression in pancreatic cancer tissues

status: Public on Oct 05 2024

city: hang zhou

first: de yang

last: mu

address: 杭州医学院临安校区

institute: hang zhou medical college

type:
Expression profiling by array
Non-coding RNA profiling by array

summary: LNCRNA plays an important role in the development of pancreatic cancer, and in order to investigate the biological function of LNCRNA in pancreatic cancer, we have measured its expression

overall_design: Divided into control and experimental groups, the control group was normal pancreatic tissue and the experimental group was pancreatic cancer tissue, n=3

first: JunGang

last: Zhang

first: DeYang

last: Mu


title: Exploring Origin-dependent Susceptibility of Smooth Muscle Cells to Aortic Diseases via Intersectional Genetics

city: Shanghai

first: Yi

last: Li

address: 320 Yueyang Road

institute: Center for Excellence in Molecular Cell Science, Chinese Academy of Sciences

summary: The developmental heterogeneity of smooth muscle cells (SMCs) plays a crucial role in the prevalence of segment-specific aortic diseases. Traditional genetic tools, however, are insufficient for in vivo analysis of disease susceptibility associated with cellular origin. To overcome this challenge, we engineered a state-of-the-art dual recombinase-mediated intersectional genetic system, tailored to precisely target SMCs from distinct developmental origins. Employing this system, we selectively knocked out Tgfbr2 in the ascending aorta and Smad4 in the aortic arch. Our results distinctly demonstrate the indispensable roles these genes play in sustaining SMC integrity across different aortic segments. This advanced genetic system not only deepens our understanding of aortopathies but also significantly enhances the potential for modeling and elucidating the mechanisms driving segment-specific aortic diseases.

overall_design: Aortic cells were harvested from adult mice each of CNC-SMC-Cre;R26-tdT and CNC-SMC-Cre;Smad4flox/flox;R26-tdT genotyped and analyzed using scRNA-seq.

first: Ximeng

last: Han

first: Enci

last: Wang

first: Huan

last: Zhu

first: Xiuzhen

last: Huang

first: Wenjuan

last: Pu

first: Mingjun

first: Kuo

last: Zhao

first: Zixin

first: Ben

last: He

first: Bin

last: Zhou


title: Phase separation of chimeric antigen receptor promotes immunological synapse maturation and persistent cytotoxicity

first: Haochen

last: Yang

state: Shanghai

address: 320 Yue-yang Road

institute: Shanghai Institute of Biochemistry and Cell Biology

summary: Major challenges of CAR-T cell therapy include poor antigen sensitivity and cell persistence. Here we report a solution to these issues by exploiting CAR phase separation. We found that incorporation of an engineered CD3e motif, EB6I, to the conventional 28Z or BBZ CAR induced self-phase separation through cation-π interactions. EB6I CAR can form a mature immunological synapse with CD2 corolla to transduce efficient antigen and costimulatory signaling, while its tonic signaling remains at low levels. Functionally, EB6I CAR-T cells exhibited improved signaling and cytoxicity against low-antigen tumor and persistent tumor killing function. In multiple primary and relapsed murine tumor models, EB6I CAR-T cells exerted better antitumor functions than conventional CAR-T cells against blood and solid cancers. This study thus unveils a new CAR engineering strategy to improve CAR-T cell immunity by leveraging molecular condensation and signaling integration.

overall_design: To assess the long-term antritumor functions of the CD19.28Z/E28Z/EB6I28Z CAR-T cells, we continuously challenged CAR-T cells with multiple rounds of tumor cells. CAR-T cells with/without tumor cell challenge were sorted for RNA-seq.

first: Xinyi

last: Xu

first: Zhengxu

last: Ren

first: Chenqi


title: Ileal Crohn’s disease exhibits reduced activity of phospholipase C-β3 (PLC-β3)-dependent Wnt/b-catenin signaling pathway [microarray]

first: Ikuo

last: Takazawa

state: CA

summary: Crohn’s disease is a chronic, debilitating inflammatory bowel disease. Here we report a   critical role for phospholipase C-β3 (PLC-β3) in intestinal homeostasis. In PLC-β3-  deficient mice, exposure to oral dextran sodium sulfate induced lethality and severe   inflammation in the small intestine. The lethality was due to PLC-β3 deficiency in multiple   non-hematopoietic cell types. PLC-β3 deficiency resulted in reduced Wnt/β-catenin  signaling, which is essential for homeostasis and regeneration of the intestinal epithelium.   PLC-β3 regulated the Wnt/β-catenin pathway in small intestinal epithelial cells (IECs) at   transcriptional, epigenetic, and potentially protein-protein interaction levels. PLC-β3-  deficient IECs were unable to respond to stimulation by R-spondin 1, an enhancer of   Wnt/β-catenin signaling. Reduced expression of PLC-β3 and its signature genes was found   in biopsies of ileal Crohn’s disease patients. PLC-β regulation of Wnt signaling was   evolutionally conserved in Drosophila. Our data indicate that reduction of PLC-β3-  mediated Wnt/β-catenin signaling contributes to the pathogenesis of ileal Crohn’s disease.

overall_design: To gain insight into the molecular mechanisms by which Plcb3 deficiency increases DSS   susceptibility, gene expression in isolated small intestinal IECs, whole jejunum or whole ileum from WT, Plcb3_KO, Plcb3_Flox or Plcb3_deltaIEC mice with or without 3% DSS treatment was analyzed using DNA microarray.

first: Tomoaki

last: Ando

first: Zach

last: Spencer

first: Ryoji

last: Ito

first: Yoshiaki

last: Tomimori

first: Zbigniew

last: Mikulski

first: Kenji

last: Matsumoto

first: Tohru

last: Ishitami

first: Lee

last: Denson

first: Yu

last: Kawakami

first: Yuko

first: Jiro

last: Kitaura

first: Yashi

last: Ahmed

first: Toshiaki


title: N-Palmitoylglycine activates transient receptor potential channel 5 and increases the risk of Brugada syndrome

city: Beijing

first: Hongxuan

address: No.8 Xishiku Street

department: Cardiology

institute: Peking university first hospital

summary: Brugada syndrome (BrS) is an arrhythmic disorder associated with an increased risk of sudden cardiac death; however, current treatment options are limited due to their side effects and variable efficacy. In this study, we employed Mendelian randomization analysis utilizing proteomic, transcriptomic, and metabolomic data to identify potential therapeutic targets for BrS. Our findings indicate that N-palmitoylglycine (PalGly) is linked to an increased risk of BrS and interacts with BrS-associated proteins, demonstrating moderate binding affinities for proteins such as DCC, CR1, CTSB, NAAA, DEFB1, EPHA1, IGF1/IGFBP3/ALS, and LTA. Although PalGly does not interact with Nav1.5, it enhances calcium sparks in ventricular cardiomyocytes. We determined that the calcium-modulating effect of PalGly is mediated by its binding to and activation of the transient receptor potential channel 5 (TRPC5) channel. Furthermore, PalGly was found to shorten the QT interval and action potential duration in Langendorff-perfused rabbit hearts and ventricular cardiomyocytes. Transcriptomic and lipidomic analyses of PalGly-treated neonatal rat cardiomyocytes revealed significant modulation of immune pathways, akin to the effects observed with TRPC5 agonists. Collectively, our study underscores the involvement of PalGly and TRPC5 in the pathophysiology of BrS.

overall_design: To investigate the cardiac effect of N-palmitoylglycine, we extracted neonatal rat cardiomyocytes and stimulated these cells with 1 μM N-palmitoylglycine 24h hours.

first: Bingxun

last: Li

first: Lin

last: Wu