title: Identification of a miRNA panel as a promising biomarker for diagnosis of glioblastoma patients using next-generation RNA sequencing

status: Public on Oct 31 2024

city: Debrecen

first: Zsuzsanna

last: Birkó

address: Egyetem tér 1.

department: Humangenetics

country: Hungary

institute: University of Debrecen

type: Non-coding RNA profiling by high throughput sequencing

summary: Applying Next Generation Sequencing technique we compared the miRNA expression pattern of tumor tissue sample of 6 GPs and peritumoral region of 6 lower grade (I-II) Glioma patients, serving as control group. To determine the difference on miRNA expresion level between GBM and control cases, we performed cluster analysis on the NGS dataset of 6 replicates for each of the two goups of samples with iDEP 96 software. In order to characterize the extent of up- or downregulation, log2FC values were calculated using the iDEP.96 web tool applying the DESeq2 algorithm. On the base of that 117 known miRNAs were identified to be differentially expressed using a threshold of false discovery rate (FDR) <0.05 and fold-change> 2 during the analysis. Among them, 35 miRNAs were upregulated (log2FC > 2) and 82 miRNAs were downregulated (log2FC < -2) with biological revelance in tissue samples comparing with the control samples. To validate our results obtained by NGS, five upregulated miRNAs: hsa-miR-196a-5p, hsa-miR-21-3p, hsa-miR-92b-5p, hsa-miR-10b-3p, hsa-miR-503-5p and three downregulated miRNAs: hsa-mir-383-5p, hsa-mir-490-3p, hsa-mir-1224-3p were chosen for RT-qPCR analysis. As the result of that hsa-miR-196a-5p, hsa-miR-21-3p, and hsa-miR-10b-3p was significantly upregulated while hsa-mir-383-5p  and hsa-mir-490-3p was significantly downregulated, compared with those in the control samples. The other three miRNAs: hsa-miR-1224-3p, hsa-miR-92b-5p, hsa-miR-503-5p  did not show significant difference between the control group and GPs.

overall_design: Tissue samples were gathered from peritumoral region of six individuals diagnosed with lower grade (Grade I-II.) glioma, serving as control group, and from six patients with GBM. All samples in both group were confirmed histopathologically. These intraoperative quick-frozen tissue samples were stored at -80⁰C until further processing. Total RNA was isolated from  the tissue samples of GBM and control group as well. Global transcriptomic profile was studied by RNA-sequencing applying the Illumina NextSeq500 platform including microRNA sequencing. In the case of all samples 3 replicates were included. In order to identify up-, or down-regulated microRNAs the GBM samples were compared to the  control samples.

first: Dóra

last: Géczi

first: Álmos

last: Klekner

first: István

last: Balogh

first: András

last: Penyige

first: Melinda

last: Szilágyi

first: József

last: Virga

first: Andrea

last: Bakó

first: Bálint

last: Nagy

first: Bernadett

last: Torner


title: An ancient competition for the conserved branchpoint sequence influences physiological and evolutionary outcomes in splicing

status: Public on Nov 22 2024

city: Galveston

first: Haiping

last: Hao

state: Texas

address: 301 University Blvd

country: USA

institute: UTMB NGS Core

type: Expression profiling by high throughput sequencing

summary: Branchpoint (bp) recognition is an early event that is required for spliceosome recruitment to a pre-mRNA substrate. The bp sequence can range from nearly invariant in yeast (UACUAAC) to quite degenerate in higher eukaryotes (YUNAY in humans), which correlates with the frequency of alternative splicing observed in the latter. The closely related KH-type RNA binding proteins (RBPs) SF1 and Quaking (QKI) can bind to a similar bp-like sequence, but the extent to which competition for the bp sequence between these two proteins might influence gene expression regulatory mechanisms is unknown. Here we find genomewide evidence for competition between these two RBPs for a subset of alternatively spliced exons that have ACUAAY-like bp sequences. Using a combination of molecular genetic and biochemical techniques with a model intron containing two bp-like sequences, we find that either can be used as a bp but high affinity binding and strong splicing repression by QKI requires both sites. This directly prevents SF1 binding and the subsequent recruitment of spliceosome-associated factors. Finally the ectopic expression of QKI in S. cerevisiae (which lacks QKI) is lethal, due at least in part to widespread splicing repression. In conclusion, QKI can function as a splicing repressor by directly competing with SF1/BBP for a subset of bp sequences that closely mirror its high affinity binding site. This suggests that QKI and degenerate bp sequences may have co-evolved which allowed for expansion of the alternative splicing repertoire in metazoans, plants, and animals.

overall_design: We inserted a galactose-inducible cDNA encoding either EGFP, mutant K120A;R124A Qki5, or wildtype Qki5 into BY4741

first: Karen

middle: L

last: de Castro

first: Robert

middle: E

last: Johnson

first: Jose

last: Abril

first: Liao

last: Kuo-Chieh

first: Steven

middle: G

last: Widen

first: John

middle: P

last: Donohue

first: William

middle: K

last: Russell

first: W

middle: S

last: Fagg


title: α4 Blockade Reduces CD8 TEM Cells in the Brain and Elevates CNS Viral Loads

status: Public on Nov 01 2024

city: Pittsburgh

first: Smita

last: Iyer

state: Pennsylvania

address: 3550 Terrace Street

department: Pathology

institute: University of Pittsburgh School of Medicine

summary: Background. The blood-brain barrier (BBB) regulates immune cell entry into the CNS, and various strategies have been developed to target this barrier in efforts to alleviate neuroinflammation in neurodegenerative diseases. α4β1 is crucial for T cell influx into the CNS. In the context of HIV, CD4 T cells drive viral infiltration, while CD8 T cells are essential for viral clearance. This dual role complicates strategies aimed at targeting T cell entry during HIV infection, yet it remains an area that has not been thoroughly explored. Methods. We investigated the impact of disrupting α4-mediated T cell entry into the CNS in SIVmac251-infected rhesus macaques treated with an α4-blocking antibody (n=4) versus an isotype control (n=4). Our comprehensive approach included single-cell analysis of CD45+ cells in the brain and spleen, spatial transcriptomics of the hippocampus, viral load measurements, and flow cytometry on brain tissue. Results. Single-cell analysis revealed a marked reduction in the brain CD8 TEM cluster, particularly in cells expressing Granulysin, CCL5, and STAT4, while CD4 T cell clusters remained unchanged, and two monocyte clusters were enriched. This shift was accompanied by an increase in brain SIV RNA+ CD45+ CD4+ T cells in the α4-treated group (vRNA+ cells = 60) compared to controls (n=11). Spatial transcriptomics of SIV vRNA+ versus SIV vRNA- regions in the hippocampus of α4-treated macaques showed enrichment of CD4 TCM and monocyte gene signatures. Correspondingly, quantitative PCR revealed higher vRNA levels in the gray matter of the PFC, STS, and Hp. These findings suggest that α4 blockade selectively disrupts CD8 TEM populations while sparing CD4 T cells, leading to elevated viral loads and heightened neuroinflammation. The increase in SIV RNA+ CD4+ T cells and viral loads in the gray matter furthermore underscores the compartmentalized effect of α4-mediated immune modulation on viral dynamics across distinct brain regions Conclusion. These results underscore the critical need to evaluate immune-targeting therapies, with implications for strategies aimed at controlling T cell influx in the context of viral infections in the brain.

sample_organism:
Macaca mulatta
blank sample

platform_organism:
Macaca mulatta
blank sample

overall_design: Sections from the prefrontal cortex and hippocampus of six rhesus macaques were collected 3 weeks post-infection with SIVmac251. Animals were treated with either a monoclonal antibody targeting α4 integrin (n=3) or an isotype control (n=3). Morphological markers, including CD45, CD3, and NeuN, were used to define regions of interest. RNAscope was employed to further delineate SIV-positive and SIV-negative regions for transcriptomic analysis.

first: Petkov

last: Alex

first: Elizaldi

last: Sonny

first: Newhouse

last: Daniel

first: Iyer

last: Smita


title: Polycomb Repressive Complexes occupancy reveals PRC2- independent PRC1 critical role in the control of limb development [ChIP-seq]

status: Public on Oct 21 2024

city: Montreal

first: marie

last: Kmita

state: Quebec

address: 110 Pine avenue west

country: Canada

institute: IRCM

type: Genome binding/occupancy profiling by high throughput sequencing

summary: The Polycomb Repressive Complexes (PRC) are key players in the regulation of tissuespecific gene expression through their ability to epigenetically silence developmental genes. They are subdivided into two multicomponent complexes, PRC1 and PRC2, functioning through posttranslational modifications of histone tails. A large body of work revealed functional interactions between PRC1 and PRC2, whereby trimethylation of lysine 27 on histone H3 (H3K27me3) by PRC2 contributes to the recruitment of canonical PRC1 (cPRC1). In parallel, a PRC2-independent binding of PRC1 has been uncovered and referred to as non-canonical PRC1 or variant PRC1 (vPRC1), in which PRC1-dependent ubiquitination of lysine 119 on histone H2A is involved in recruiting PRC2/propagating PRC2-dependent H3K27 trimethylation. While it was initially assumed that cPRC1 and vPRC1 bind distinct targets, subsequent evidence pointed to cPRC1 and vPRC1 sharing a significant subset of targets. How the functional interplay between PRC2, cPRC1 and vPRC1 contributes to gene regulation remains partially understood. Here, we show that, in the developing limb, PRC2 inactivation barely affects PRC1 occupancy, as the majority of PRC2- bound loci are bound by vPRC1 (RYBP-PRC1), both in wild type and PRC2 mutant limbs. Consistent with this, we found that loci bound by CBX2, a PRC1 subunit involved in the recognition of H3K27me3 and thereby recruitment of cPRC1, are, for the vast majority, also bound by vPRC1. Intriguingly, analysis of PRC2 mutant limbs revealed that while a large part of CBX2 occupancy is lost in absence of PRC2 function, as expected from the absence of H3K27me3, there is a significant number of genes retaining CBX2 occupancy as well as a few genes with apparent gain of CBX2 binding. Importantly, among these genes, 56 of them correspond to developmental genes known for playing a key role in limb morphogenesis. Based on the importance of vPRC1 in gene silencing, our findings emphasize the primary role of PRC2-independent PCR1 function in regulating developmental genes and questions the role of PRC2/cPRC1 in controlling developmental programs.

overall_design: ChIP analysis of SUZ12, H3K27me3, RING1B, H2AK119ub, RYBP, CBX2, CBX8, H3K27ac in proximal and distal limb tissue at E12.5

first: Fanny

last: GUERARD-MILLET

first: Claudia

last: GENTILE

first: Racheal

last: PAUL

first: Christian

last: POITRAS


title: Effects of Denervation on Muscle Function

city: Nanjing

first: Zhiqiang

last: Huang

state: Jiangsu

address: Hankou 22

department: Medical School

country: China

institute: Nanjing University

summary: Innervation of skeletal muscle fibers plays a crucial role in the maintenance of muscle tone and normal functioning, but little is known to date about denervated muscle atrophy and its underlying mechanisms. To this end, we performed RNA sequencing of skeletal muscle from sciatic nerve-excised C57BL/6 and BALB/c mice to investigate the underlying mechanisms of denervated skeletal muscle atrophy.

overall_design: We investigated the important role of the muscle membrane protein α7nAChR in the maintenance of muscle function by denervation.

first: Xiaolu

last: Jin

first: Hongwei

last: Wang


title: eQTL in diseased colon tissue identifies novel target genes associated with IBD

status: Public on Oct 16 2024

city: Chapel Hill

first: Terrence

last: Furey

state: NC

address: 120 Mason Farm Rd

institute: University of North Carolina at Chapel Hill

summary: Genome-wide association studies (GWAS) have identified over 300 loci associated with the inflammatory bowel diseases (IBD), but putative causal genes for most are unknown. We conducted the largest disease-focused expression quantitative trait loci (eQTL) analysis using colon tissue from 252 IBD patients to determine genetic effects on gene expression and potential contribution to IBD. Combined with two non-IBD colon eQTL studies, we identified 194 potential target genes for 108 GWAS loci. eQTL in IBD tissue were enriched for IBD GWAS loci colocalizations, provided novel evidence for IBD-associated genes such as ABO and TNFRSF14, and identified additional target genes compared to non-IBD tissue eQTL. IBD-associated eQTL unique to diseased tissue had distinct regulatory and functional characteristics with increased effect sizes. Together, these highlight the importance of eQTL studies in diseased tissue for understanding functional consequences of genetic variants, and elucidating molecular mechanisms and regulation of key genes involved in IBD.

overall_design:
RNA-seq of colon tissue from IBD patients for eQTL mapping

***************************************************************
Submitter states that missing raw data are being made available for controlled access in dbGaP.
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first: Shehzad

middle: Z

last: Sheikh

first: Nina

middle: C

last: Nishiyama


title: Polycomb Repressive Complexes occupancy reveals PRC2- independent PRC1 critical role in the control of limb development [RNA-seq]

overall_design:
RNAseq analysis in proximal and distal limb tissue at E12.5

UPDATE: [June 06, 2025] The titles of Samples GSM6693221-GSM6693232 were updated.


title: TNFSF13 insuffiency causes epithelial and immune disruption in the human colon I

status: Public on Nov 05 2024

city: Philadelphia

first: Noor

last: Dawany

address: 421 Curie Blvd

institute: CHOP

summary: Objective: Cytokines produced by epithelial and immune cells can act upon one another to promote mucosal healing- a process that goes awry with chronic inflammation as in inflammatory bowel disease. TNFSF13 is a cytokine important for B cell function, but its role in epithelial cell behavior and contribution to inflammatory bowel disease is poorly understood.  Design: We performed histological and functional assays, single cell transcriptomics, and imaging mass cytometry on tissue-derived colonoids or biopsies from a patient with very early onset inflammatory bowel disease (VEO-IBD) harboring a novel TNFSF13 variant. To confirm causal, variant-driven phenotypes, parallel experiments were performed in induced pluripotent stem cell (iPSC)- derived colon organoids engineered with the same TNFSF13 variant.  Results: TNFSF13 variant colonoids exhibited reduced TNFSF13 expression associated with increased proliferation and reduced apoptosis, which was confirmed in iPSC-derived colon organoids. Single cell RNA-sequencing and flow cytometry defined FAS as the predominant colonic epithelial receptor for TNFSF13. TNFSF13 variant colon biopsies demonstrated a shift in tissue B cell populations consistent with altered differentiation of memory B cells. Finally, imaging mass cytometry revealed an increase in epithelial-associated B cells in TNFSF13 variant colon tissue sections compared to healthy and non-monogenic VEO-IBD controls.  Conclusions: The current study defines a previously unknown role for the cytokine TNFSF13 as a regulator of colonic epithelial proliferation and apoptosis. Furthermore, this study suggests that variant TNFSF13 can contribute to aberrant epithelial- B cell crosstalk in the human colon.

overall_design:
scRNA-seq analysis of colonoids from 2 controls, 2 patients with VEO-IBD and 1 patient with VEO-IBD and variant in TNFSF13 (APRIL).  Samples included untreated colonoids as well as colonoisd treated with TNFa, IFNg or TRAIL. Cell hashing was used to allow for multiplexing of untreated and treated colonoids from the same sample ID.
HTO barcode sequence hastagged sample
TotalSeq-B0251 anti-human Hashtag 1 GTCAACTCTTTAGCG Untreated
TotalSeq-B0252 anti-human Hashtag 2 TGATGGCCTATTGGG TNFa-treated
TotalSeq-B0253 anti-human Hashtag 3 TTCCGCCTCTCTTTG IFNg-treated
TotalSeq-B0254 anti-human Hashtag 4 AGTAAGTTCAGCGTA TRAIL-treated

first: Kathryn

last: Hamilton

first: Judith

middle: R

last: Kelsen


title: Effect of GCN5 depletion in hepatocytes in NAFLD

status: Public on Oct 23 2024

city: 南京市

first: Jing

state: 江苏省

address: 南京市江宁区龙眠大道639号中国药科大学

institute: 中国药科大学

summary: Non-alcoholic fatty liver disease (NAFLD) is a burden of global health, yet the mechanism of the disease is not fully elucidated. General control non-repressed protein 5 (GCN5) is histone acetyltransferases. We invesgated the effect of GCN5 deletion on gene expression in a NAFLD cell model.

overall_design: To invesgate the effect of GCN5 depeltion on gene expression in hepatocytes. GCN5 knockout and wild-type HL-7702 cells were separately seeded into 6-well plates, treated with PAOA for 24 h, and harvested for RNA-seq.

first: Zu-Guo

last: Zheng

first: Ping

last: Li


title: TNFSF13 insuffiency causes epithelial and immune disruption in the human colon II

overall_design: scRNA-seq analysis of epithelial and stromal cells from colon biopsies of a patient with VEO-IBD and a healthy control


title: The RNA exosome maintains cellular RNA homeostasis by controlling transcript abundance in the brain

city: Los Angeles

first: Lauryn

middle: Alexis

last: Higginson

state: CA

address: 1050 Childs Way Room 201B

department: Biological Sciences

institute: University of Southern California

summary: Intracellular ribonucleases (RNases) are essential in all aspects of RNA metabolism, including maintaining accurate RNA levels. Inherited mutations in genes encoding ubiquitous RNases are associated with human diseases, primarily affecting the nervous system. Recessive mutations in genes encoding an evolutionarily conserved RNase complex, the RNA exosome, lead to syndromic neurodevelopmental disorders characterized by progressive neurodegeneration, such as Pontocerebellar Hypoplasia Type 1b (PCH1b). We establish a CRISPR/Cas9-engineered Drosophila model of PCH1b to study cell-type-specific post-transcriptional regulatory functions of the nuclear RNA exosome complex within fly head tissue. Here, we report that pathogenic RNA exosome mutations alter activity of the complex, causing widespread dysregulation of brain-enriched cellular transcriptomes, including rRNA processing defects—resulting in tissue-specific, progressive neurodegenerative effects in flies. These findings provide a comprehensive understanding of RNA exosome function within a developed animal brain and underscore the critical role of post-transcriptional regulatory machinery in maintaining cellular RNA homeostasis within the brain.

overall_design: Two biological replicates of newly eclosed adult female flies were prepared for each biological replicate (Rrp40WT, Rrp40G11A, and Rrp40G146C). For each sample, 20 fly heads were pooled together, and nuclei were dissociated. Nuclei quantity and and quality were determined using propidium iodide. Single-cell libraries were prepared using Chromium Next GEM Single Cell 3' GEM, Library & Gel Bead Kit v3.1. Sequencing reads were mapped to the Drosophila melanogaster genome and CellRanger was used to create the reference.

middle: A

first: Derrick

middle: J

last: Morton


title: Mouse uterine gene expressions during peri-implantation period

status: Public on Oct 08 2024

city: Bunkyo-ku

first: Shizu

last: Aikawa

state: Tokyo

address: 7-3-1 Hongo

country: Japan

institute: The University of Tokyo

summary: The mouse uterine gene expressions during peri-implantation period at pregnancy day 4 and 6 was investigated by Visium spatial transcriptome.

overall_design: The day when the vaginal plug was detected was considered day 1 of pregnancy. Pregnant mice were euthanized by cervical dislocation on designated days of pregnancy for the evaluation of pregnancy phenotypes and sample collection. On day 4, one uterine horn was flushed with saline to confirm the presence of blastocysts. Embryo attachment sites were observed as blue bands soon after intravenous injection of 1% solution of Chicago blue dye (Sigma-Aldrich) in saline on day6. Collected tissues were snap-frozen. Frozen sections (10 μm) were mounted on gene expression slides and sent to KOTAI Biotechnologies for subsequent procedures. After reaction with Proteinase K for 40 min, the sections were hybridized with spatial tags on the slides and reverse-transcribed in situ. The synthesized cDNAs were subjected to RNA-sequencing using DNBseq (MGI), producing 300 million reads/sample.

first: Yasushi

last: Hirota


title: RNA-Seq experiment of wild-type mouse livers with mLMRs knockdown and hLMRs rescue

status: Public on Oct 15 2024

city: Bethesda

first: Haiming

last: Cao

state: MD

address: 9000 Rockville Pike

department: NHLBI

institute: NIH

summary:
Rescue experiment in wild-type mice offers a valuable mean to verify the functional similarity of lncRNAs. By suppressing mLMRs in mice and observing the impact on gene regulation, followed by overexpression of hLMRs, we can determine the ability of hLMRs to rescue the loss-of-function effects of mLMRs. Should hLMRs exhibit a reversal of the loss-of-function on transcriptome regulations, this would suggest that they have similar functions to mLMRs.
To do this, we used adenovirus-delivered shRNAs to suppress the expression of mLMRs in the livers of wild-type mice, while overexpressing corresponding hLMRs as a rescue group.

overall_design: Wild-type mice (C57BL6, 12 weeks age) were injected with three groups of viruses, including control viruses for knockdown and overexpression (Lac+pAdv5, control group), mLMR knockdown viruses and control viruses for overexpression (mouse-lncRNA KD+pAdv5, mLMR knockdown group), and mLMR knockdown viruses and hLMR overexpression group (mouse-lncRNA KD+human-lncRNA OE, hLMR rescue group). The adenoviruses were intravenously delivered to  mice at a dose of 5 × 10^8 PFU/mouse for overexpression virus or 1 × 10^9 PFU/mouse for knockdown virus. After a 5-hour fast (~9:00 am to 2:00 pm), the livers  were harvested for RNA-Seq analysis after 7 days.

first: Chengfei

last: Jiang


title: TNFSF13 insuffiency causes epithelial and immune disruption in the human colon

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

overall_design: Refer to individual Series


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

city: Boston

first: Ryan

middle: Jared

last: Marina

state: MA

address: 45 Shattuck Street

department: Biological Chemistry and Molecular Pharmacology

institute: Harvard Medical School


title: Discovering Genetic Modulators of the Protein Homeostasis System through Multilevel Analysis

status: Public on Oct 03 2024

city: Amherst

first: Peter

last: Chien

address: 240 Thatcher Rd, Life Science Laboratories Room N325

department: Biochemistry and Molecular Biology

institute: University of Massachusetts Amherst

summary: Every protein progresses through a natural lifecycle from birth to maturation to death; this process is coordinated by the protein homeostasis system. Environmental or physiological conditions trigger pathways that maintain the homeostasis of the proteome. An open question is how these pathways are modulated to respond to the many stresses that an organism encounters during its lifetime. To address this question, we tested how the fitness landscape changes in response to environmental and genetic perturbations using directed and massively parallel transposon mutagenesis in Caulobacter crescentus. We developed a general computational pipeline for the analysis of gene-by-environment interactions in transposon mutagenesis experiments. This pipeline uses a combination of general linear models (GLMs), statistical knockoffs, and a nonparametric Bayesian statistical model to identify essential genetic network components that are shared across environmental perturbations. This analysis allows us to quantify the similarity of proteotoxic environmental perturbations from the perspective of the fitness landscape. We find that essential genes vary more by genetic background than by environmental conditions, with limited overlap among mutant strains targeting different facets of the protein homeostasis system. We also identified 146 unique fitness determinants across different strains, with 19 genes common to at least two strains, showing varying resilience to proteotoxic stresses. Experiments exposing cells to a combination of genetic perturbations and dual environmental stressors show that perturbations that are quantitatively dissimilar from the perspective of the fitness landscape are likely to have a synergistic effect on the growth defect.

overall_design:
To investigate the fitness landscapes of Caulobacter crescentus under proteotoxic stresses, we have generated transposon mutagenesis libraries from wild-type, ∆lon, ∆clpA, ∆clpB, and dnaK-NI strains.
We then subjected these libraries to heat, oxidative, and l-canavanine stress at three different levels. (low, medium, high)
Each condition contains 4 replicates. There are 2 control conditions performed with each condition. T0 control obtained from the overnight recovery after thawing. T24 control obtained by growing the libraries for 24 hours in 30C shaker after the overnight recovery.
We then performed transposon sequencing to quantify number of total and unique transposon insertions in each sample/condition.
Sequencing is utilized to quantify the frequency of transposon-induced mutations and identify a set of conditionally essential, beneficial, or detrimental genes for each environment by applying a regularized negative binomial regression combined with local False Discovery Rate (FDR) testing within a general linear model (GLM) framework.

first: Vishal

last: Sarsani

first: Berent

last: Aldikacti

first: Tingting

last: Zhao

first: Shai

last: He

first: Patrick

last: Flaherty


title: A massively parallel reporter assay library to screen short synthetic promoters in mammalian cells

city: Salt Lake City

first: Justin

last: English

state: UT

address: 15 N Medical Drive East, Rm 4100

department: Biochemistry

institute: University of Utah

summary: Cellular transcription enables cells to adapt to various stimuli and maintain homeostasis. Transcription factors bind to transcription response elements (TREs) in gene promoters, initiating transcription. Synthetic promoters, derived from natural TREs, can be engineered to control exogenous gene expression using endogenous transcription machinery. This technology has found extensive use in biological research for applications including reporter gene assays, biomarker development, and programming synthetic circuits in living cells. However, a reliable and precise method for selecting minimally-sized synthetic promoters with desired background, amplitude, and stimulation response profiles has been elusive. In this study, we introduce a massively parallel reporter assay library containing 6184 synthetic promoters, each less than 250 bp in length. This comprehensive library allows for rapid identification of promoters with optimal transcriptional output parameters across multiple cell lines and stimuli. We showcase this library’s utility to identify promoters activated in unique cell types, and in response to metabolites, mitogens, cellular toxins, and agonism of both aminergic and non-aminergic GPCRs. We further show these promoters can be used in luciferase reporter assays, eliciting 50-100 fold dynamic ranges in response to stimuli. Our platform is effective, easily implemented, and provides a solution for selecting short-length promoters with precise performance for a multitude of applications.

sample_organism:
Homo sapiens
Mesocricetus auratus
Mus musculus
synthetic construct

overall_design: Multiple mammalian cell lines were transfected with the TRE-MPRA library for 24 hours and were then treated with growth factors or specific compounds for six hours. For some experiments, cells were co-transfected with specific G protein-coupled receptors and treated with receptor agonsists for six hours. Some treatments contained multiple technical replicates, while others were performed once. RNA and DNA barcode abundance was determined via Illumina next-generation sequencing.

first: Adam

middle: M

last: Zahm


title: Canonical and variant PRC1 functionally converge on PRC2 targets to regulate transcriptional programs during mouse limb development

first: marie

last: Kmita


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

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: Ribosome collisions trigger subunit splitting in E. coli

status: Public on Oct 10 2024

city: Baltimore

first: Allen

last: Buskirk

address: 725 N. Wolfe St

department: Molecular Biology and Genetics

institute: Johns Hopkins University School of Medicine

summary: Although many clinically important antibiotics inhibit bacterial ribosomes, the mechanisms by which bacterial cells rescue ribosomes stalled by antibiotics remain poorly understood. Ribosome stalling leads to collisions that recruit ribosome quality control (RQC) factors that recycle the ribosome subunits and target nascent proteins for degradation. Surprisingly, loss of known RQC factors in E. coli does not lead to significant antibiotic sensitivity, even though antibiotics stall ribosomes and induce collisions, suggesting the existence of additional, uncharacterized RQC mechanisms. Here we report a novel mechanism for ribosome quality control (RQC) in bacteria in which the DExH-box ATPase HrpA splits stalled ribosomes into subunits. HrpA selectively acts on collided ribosomes and its activity is dependent on ATP hydrolysis. The cryo-EM structure of HrpA bound to collided ribosomes reveals insight into its selectivity and mechanism: the C-terminal domain of HrpA senses the collision and its helicase domain bind mRNA downstream of the ribosomes, where it likely exerts a pulling force that destabilizes the stalled ribosome. These studies highlight the importance of ribosome splitting as a highly conserved RQC mechanism across all three domains of life and identify an important pathway in proteobacteria that allows proteobacteria to tolerate ribosome-targeting antibiotics.

sample_organism: Escherichia coli str. K-12 substr. MG1655

overall_design: Ribosome profiling with two replicates for three strains (wild-type MG1655, delta-hrpA, and the delta-hrpA delta-smrB double) containing the plasmid "IRAGP_reporter".


title: Mast Cells Proliferate in the Peri-Hippocampal Space During Early Development and Modulate Local and Peripheral Immune Cells

status: Public on Sep 22 2024

first: Margaret

last: McCarthy

state: Maryland

address: 670 W Baltimore St

institute: University of Maryland, Baltimore

summary: Brain development is a non-linear process of regionally specific epochs occurring during windows of sensitivity to endogenous and exogenous stimuli. We have discovered an epoch in the neonatal rat brain defined by a transient population of peri-hippocampal mast cells (phMCs) that are abundant from birth through 2-weeks postnatal but absent thereafter. The phMCs are maintained by proliferation and harbor a unique transcriptome compared to mast cells residing in the skin, bone marrow or other brain regions. Pharmacological activation of this population broadly increases blood-brain-barrier permeability, recruits peripheral immune cells and stunts local microglia proliferation. Examination of the post-mortem human brain revealed mast cells in the peri-hippocampal region of a newborn but not an older child, suggesting a similar developmental period exists in humans. Mast cells specifically and early life inflammation generally, have been linked to heightened risk for neurodevelopmental disorders and these results reveal a plausible source of that risk.

overall_design: We perfused PN2 and PN7 neonates and dissected the hippocampus, hypothalamus, bone marrow and skin. Each replicate contains cells pooled from 6 animals. We sorted mast cells using flow cytometry for CD45 high, CD11b- and FcERI+ cells. We performed bulk gene RNA-seq of each sample using SMART-Seq v4 UltraLow Input RNA Sequencing.

first: Alexa

last: Blanchard

first: Anna

last: Maximova

first: Taylor

last: Phillips-Jones

first: Matthew

last: Bruce

first: Pavlos

last: Anastasiadis

first: Christie

middle: V

last: Dionisos

first: Kaliroi

last: Engel

first: Erin

last: Reinl

first: Aidan

last: Pham

first: Sonia

last: Malaiya

first: Nevil

last: Singh

first: Seth

last: Ament


title: Immunotherapies that repolarize macrophages and regulatory T cells enhance the effect of chemotherapy in high-grade serous ovarian cancer (Human)

status: Public on Oct 11 2024

city: London

first: Samar

last: Elorbany

address: Barbican

department: Tumour Microenvironment

country: United Kingdom

institute: Queen Mary University of London

summary:
We conducted single-cell RNA sequencing (scRNAseq) of tumor-infiltrating immune cells in high-grade serous ovarian cancer (HGSOC) omental biopsies to identify potential targets that could enhance response to neo-adjuvant chemotherapy (NACT). Analysis of 64,097 cells identified NACT-induced overexpression of stabilin-1 (clever-1) on macrophages and FOXP3 in Tregs and these were validated at the protein level. STAB1 inhibition in vitro induced anti-tumor macrophages. FOXP3 anti-sense oligonucleotide (FOXP3-ASO), repolarised Tregs to an effector T cell phenotype.
ScRNAseq on 69,781 cells from an HGSOC syngeneic mouse model recapitulated the patients’ data. Combining chemotherapy with anti-stabilin1 antibody and/or Foxp3-ASO significantly increased survival of mice with established peritoneal disease in two HGSOC syngeneic models and progression-free survival in a third model. Long-term survivors (300 days+) were resistant to tumor rechallenge. Anti-stabilin1 antibody enriched the tumors with CXCL9+ macrophages and Foxp3-ASO increased TBET cell infiltration. Our results suggest targeting these molecules may improve chemotherapy response in patients.

overall_design: Tumours were taken either during primary debulking surgery pre-NACT (n=3) or three week after chemotherapy in the post-NACT(n=4).

first: Chiara

last: Berlato

first: Larissa

last: Carnevalli

first: Eleni

last: Maniati

first: Jun

last: Wang

first: Simon

middle: T

last: Barry

first: Ranjit

last: Manchanda

first: Julia

last: Kzhyshkowska

first: Frances

last: Balkwill


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

first: Ryan

last: Marina

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.

first: Tuo

last: Zhang

first: Karen

last: Adelman


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.


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

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.


title: Triptolide sensitizes cancer cells to nucleoside DNA methyltransferase inhibitors through inhibition of DCTPP1 mediated cell-intrinsic resistance

status: Public on Oct 01 2024

first: Jianyong

last: Liu

address: 1550 Orleans St

department: Oncology

institute: Johns Hopkins University

type: Methylation profiling by genome tiling array

first: Ajay

last: Vaghasia


title: Kidney injury induced by sodium oxalate

status: Public on Sep 21 2024

city: Fuzhou

first: Zhimin

last: Chen

address: Chazhong Road NO.20

institute: the First Affiliated Hospital, Fujian Medical University, Fuzhou

summary: In order to reveal the molecular physiological damage mechanism of kidney induced by oxalic acid, single cell sequencing was performed on the kidney of mice with control group and oxalic acid induced acute kidney injury.

overall_design: Harvested cells at the required densities were combined with gel beads  containing the barcoded information along with a mixture of cells and  enzymes. Oil surfactant droplets in a microfluidic double-cross system were used for encapsulation to generate Gel Beads-In-Emulsions (GEMs). GEMs were passed through a reservoir and collected while the gel beads were lysed to release the barcode sequence. The cDNA fragment was then reverse transcribed before the sample was labeled. The gel beads and the oil droplets are ruptured and PCR amplification was performed using the generated cDNA as a template. The products of all GEMs were mixed and a standard sequencing library is constructed. The cDNA was first enzymatically digested into fragments of about 200300 base pairs (bp), together with the library building process of traditional secondgeneration sequencing such as sequencing junction and primers. Finally, the DNA library was generated by PCR amplification

first: Yanfang

last: Xu


title: Proton pump inhibitors modulate the barrier dysfunction caused by IL-13 in the epithelium in eosinophilic esophagitis

status: Public on Sep 13 2024

first: Megha

last: Lal

state: PA

address: 3615 Civic Center Blvd

department: Pediatrics

institute: Children's Hospital of Philadelphia

summary: Eosinophilic esophagitis (EoE) is a chronic immune-mediated disorder triggered by specific food antigens, characterized by eosinophil-rich multicellular inflammation and structural changes in the epithelium. Treatment options for EoE include dietary therapy, pharmacological therapy, or a combination of both, chosen based on the patient's preference and clinical progression. Pharmacological options include proton pump inhibitors (PPIs), corticosteroids, and biologic therapy. Although PPI therapy is used for EoE management, its underlying mechanism of action remains unclear. To investigate the effects of omeprazole, an orally bioavailable PPI commonly used for EoE therapy, on the dynamics of esophageal epithelial barrier function and inflammation, we employed air-liquid interface (ALI) culture. We examined how omeprazole affects gene expression changes caused by IL-13 treatment. ALI cultures were treated with 100 ng/ml of IL-13 and/or 50 µM of acid-activated omeprazole for 96 hours, and bulk RNA sequencing was performed to analyze the epithelial-specific transcriptomes of IL-13 and/or omeprazole-treated ALI. Our findings suggest potential gene interactions where omeprazole may mitigate transcriptional changes induced by IL-13, indicating that omeprazole may attenuate IL-13 mediated epithelial dysfunction relevant to EoE pathophysiology by modulating pathways associated with inflammation, tissue repair, and cell-cell communication.

overall_design: Effect of Omeprazole on gene expression profiling of air-liquid interface (ALI) derived from telomerase-immortalized human esophageal keratinocyte cell line, EPC2-hTERT.

first: Ravi

last: Gautam

first: Melanie

last: Ruffner


title: Blood-brain barrier dysfunction in response to Alzheimer's disease mutations and aged serum in a tissue-engineered microvascular model

status: Public on Sep 01 2024

first: Peter

last: Searson

address: 3400 North Charles Street

summary: Alzheimer’s disease is a disease of neurodegeneration and aging that affects millions of Americans, and is expected to impact millions more without significant advancement in our understanding of the disease. An emerging focus in Alzheimer’s study is the role of the cerebrovasculature in the initiation, progression, and exacerbation of symptomatic disease.  Disruption of the blood-brain barrier, which tightly controls any exchange between systemic circulation and brain tissue, has manifested in post-mortem and in vivo studies of late-stage Alzheimer’s disease as microbleeds, dysfunctional glucose transport, and impaired efflux of toxins; additional animal studies have indicated that some vascular dysfunction precedes neuronal degeneration in the progression of the disease.  Thus, to understand the drivers and progression of Alzheimer’s disease in hopes of identifying therapeutic breakpoints, the role of blood-brain barrier dysfunction must be investigated. Here, we utilize a tissue-engineered model of the blood-brain barrier with high spatiotemporal resolution to assess its dysfunction with cell-intrinsic mutations associated with Alzheimer’s (APP(Swe) and PSEN1(M146V)) and the systemic influence of aged blood components (exposure to aged vs. young human serum).

overall_design: total samples collected n = 27; n = 3 for each condition across three cell lines (APP mutant, PSEN1 mutant, WT control) and three serum perfusion conditions (standard media, young human serum supplementation, aged human serum supplementation)

first: Tracy

middle: D

last: Chung


title: Droplet Hi-C enables scalable, single-cell profiling of chromatin architecture in heterogeneous tissues

status: Public on Sep 04 2024

city: La Jolla

first: Yang

last: Xie

address: Rm.2071, 9500 Gilman Dr

department: Department of Cellular and Molecular Medicine

institute: UC San Diego

type: Methylation profiling by high throughput sequencing

summary: Current methods for analyzing chromatin architecture are not readily scalable to heterogeneous tissues. Here we introduce Droplet Hi-C, which uses a commercial microfluidic device for high-throughput, single-cell chromatin conformation profiling in droplets. Using Droplet Hi-C, we mapped the chromatin architecture of the mouse cortex and analyzed gene regulatory programs in major cortical cell types. In addition, we used this technique to detect copy number variations, structural variations and extrachromosomal DNA in human glioblastoma, colorectal and blood cancer cells, revealing clonal dynamics and other oncogenic events during treatment. We refined the technique to allow joint profiling of chromatin architecture and transcriptome in single cells, facilitating exploration of the links between chromatin architecture and gene expression in both normal tissues and tumors. Thus, Droplet Hi-C both addresses critical gaps in chromatin analysis of heterogeneous tissues and enhances understanding of gene regulation.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design:
Commercial microfluidic device based high-throughput, single-cell chromatin conformation profiling in droplets

>>>Submitter states: Raw data for GSM8521393, GSM8521394, or GSM8521395 will be submitted to dbGaP due to patient privacy concerns.<<<

first: Lei

last: Chang

first: Bing

last: Ren


title: Scaled and efficient derivation of loss-of-function alleles in risk genes for neurodevelopmental and psychiatric disorders in human iPSCs

status: Public on Sep 17 2024

city: Evanston

first: Jubao

last: Duan

state: IL

address: 1001 University Place

department: Center for Psychiatric Genetics

institute: NorthShore University HealthSystem

summary: Translating genetic findings for neurodevelopmental and psychiatric disorders (NPD) into actionable disease biology would benefit from large-scale and unbiased functional studies of NPD genes. Leveraging the cytosine base editing (CBE) system, here we developed a pipeline for clonal loss-of-function (LoF) allele mutagenesis in human induced pluripotent stem cells (hiPSCs) by introducing premature stop-codons (iSTOP) that lead to mRNA nonsense-mediated-decay (NMD) or protein truncation. We tested the pipeline for 23 NPD genes on 3 hiPSC lines and achieved highly reproducible, efficient iSTOP editing in 22 NPD genes. Using RNAseq, we confirmed their pluripotency, absence of chromosomal abnormalities, and NMD. Interestingly, for three schizophrenia risk genes (SETD1A, TRIO, CUL1), despite the high efficiency of base editing, we only obtained heterozygous LoF alleles, suggesting their essential roles for cell growth. We replicated the reported neural phenotypes of SHANK3-haploinsufficiency and found CUL1-LoF reduced neurite branches and synaptic puncta density. This iSTOP pipeline enables a scaled and efficient LoF mutagenesis of NPD genes, yielding an invaluable shareable resource.

overall_design: As part of the SSPsyGene Consortium, our Assay and Data Generation Center (ADGC) for the Model of iPSC-derived Neurons for NPD (MiNND) aims to employ the CBE editor-based iSTOP approach to generate isogenic hiPSC lines carrying LoF alleles for about 150-200 NPD genes on multiple donor genetic backgrounds.  Here, leveraging an improved reporter gene editing enrichment system that can substantially increase the CBE iSTOP editing efficiency in hiPSC, we established a semi-automated pipeline for parallel and efficient clonal LoF mutagenesis of a large number of genes. We tested the workflow on 23 NPD genes with 3 donor hiPSC lines (KOLF2.2J, CW20107, MGS_CD14). We obtained high and reproducible iSTOP editing efficiency across all three hiPSC lines. We systematically characterized the engineered isogenic iSTOP hiPSC lines for pluripotency, karyotyping, neuron differentiation capacity, and the expected NMD and LoF.

first: Siwei

first: Zhiping

last: Pang


title: Longitudinal transcriptome analysis of high risk cattle to determine host expressional changes associated with BRD development

status: Public on Sep 18 2024

city: Canyon

first: Matthew

last: Scott

state: TX

address: 3201 Russell Long Blvd

institute: Texas A&M University

summary: Whole blood samples from 36 cattle, across two independent populations, were sequenced via a Illumina NovaSeq 6000 system. Samples were bioinformatically processed in a HISAT2/StringTie pipeline, and stratified based on treatment frequency for BRD (healthy, treated once, treated twice or more and/or died). Differential expression analysis was conducted in R, utilizing an additive, multifactor generalized linear model and blocking for paired samples and year. Genes were considered differentially expressed with an FDR cutoff of 0.10. The objective of this study was to identify differentially expressed genes and mechanisms within and across treatment cohorts overtime, and determine host expressional changes influenced by BRD within the first 28 days of facility arrival.

overall_design: 73 whole blood mRNA samples from 36 individual cattle, taken at days 0, 28, and end of study (~63).


title: Cancer-associated DNA Hypermethylation of Polycomb Targets Requires DNMT3A Dual Recognition of histone H2AK119 Ubiquitination and the Nucleosome Acidic Patch [CUT7Run]

status: Public on Aug 31 2024

city: New York City

first: Kristjan

middle: H.

last: Gretarsson

state: New York

institute: Columbia University

summary: During tumor development, promoter CpG islands (CGIs) that are normally silenced by Polycomb repressive complexes (PRCs) become DNA hypermethylated. The molecular mechanism by which de novo DNA methyltransferase(s) catalyze CpG methylation at PRC-regulated regions remains unclear. Here we report a cryo-EM structure of DNMT3A long isoform (DNMT3A1) N-terminal region in complex with nucleosome carrying PRC1-mediated histone H2A lysine 119 mono-ubiquitination (H2AK119Ub). We identify regions within the DNMT3A1 N-terminus that bind H2AK119Ub and the nucleosome acidic patch. This bidentate interaction is specifically required for effective DNMT3A1 engagement with H2AK119Ub-modified chromatin in cells. Furthermore, aberrant redistribution of DNMT3A1 to Polycomb target genes inhibits their transcriptional derepression during cell differentiation and recapitulates the cancer-associated DNA hypermethylation signature, an effect rescued by disruption of DNMT3A1-acidic patch interaction. Together, our analyses reveal a binding interface critical for countering promoter CGI DNA hypermethylation, a major molecular hallmark of cancer.

overall_design: Cut&Run of IgG, H3K36me2, H3K27me3, H2AK119Ub and H3K4me3 in 10T cells and QKO mESC

first: Danielle

last: Maryanski

first: Jamie

last: McCuiston

first: Michael-Christopher

last: Keogh


title: KIAA1429 promotes cell proliferation by regulating MYC messenger RNA in clear cell renal cell carcinoma

status: Public on Sep 10 2024

city: Nantong

first: Zhan

last: Chen

address: No. 666 Shengli Road

department: Urology Surgery

institute: The Second Affiliated Hospital of Nantong University

summary: KIAA1429 is an integral component of the m6A methyltransferase complex, essential for facilitating the interaction between the catalytic core components METTL3 and METTL14. However, the mechanisms by which KIAA1429 influences clear cell Renal cell carcinoma (ccRCC) remain incompletely elucidated. Analysis of the TCGA and GEO database confirmed that KIAA1429 was significantly upregulated in ccRCC samples, which correlated with poor survival. Additionally, qRT-PCR, Western Blot and IHC detection revealed significantly upregulated levels of KIAA1429 in ccRCC tissues. Moreover, multivariable Cox regression analysis showed that patients with ccRCC with high KIAA1429 expression had a poor clinical prognosis. Notably, silencing KIAA1429 significantly inhibited the proliferation, migration and invasion of ccRCC cells. Similarly, KIAA1429 knockdown also significantly inhibited the growth of ccRCC cells in nude mice. Mechanistically, KIAA1429 silencing significantly reduced the overall level of m6A in, the stability of MYC mRNA and the mRNA and protein expressions of MYC in ccRCC cells. Furthermore, RIP-qRT-PCR experiments showed that KIAA1429 could directly bind to MYC mRNA. MeRIP assay revealed that KIAA1429 knockdown resulted in a significant decrease in the m6A level of MYC mRNA in ccRCC cells. In conclusion, KIAA1429 regulates the proliferation, migration and invasion of ccRCC cells by stabilizing MYC mRNA in an m6A-dependent manner. Therefore, targeting the KIAA1429-MYC signalling pathway emerges as a potential strategy for the treatment of renal cell carcinoma.

overall_design: KIAA1429 knockout ACHN cells and Negative Control ACHN cells


title: Single-cell transcriptomic atlas of enteroendocrine cells along the murine gastrointestinal tract.

city: Cambridge

first: Fiona

last: Gribble

state: Cambridgeshire

address: Box 289 Addenbrooke's Hospital, Hills Road

department: Medical Research Laboratories

institute: University of Cambridge

summary: Enteroendocrine cells (EECs) produce over 20 gut hormones which contribute to intestinal physiology, nutrient metabolism and the regulation of food intake. The objective of this study was to generate a comprehensive transcriptomic map of mouse EECs from the stomach to the rectum.

overall_design:
EECs were purified by flow-cytometry from the stomach, upper small intestine, lower small intestine, caecum and large intestine of NeuroD1-Cre mice, and analysed by single cell RNA sequencing.
Neurod1-Cre, large intestine, scRNA-seq; re-analysis of GSE137572

first: Christopher

last: Smith


title: Cancer-associated DNA Hypermethylation of Polycomb Targets Requires DNMT3A Dual Recognition of histone H2AK119 Ubiquitination and the Nucleosome Acidic Patch [RRBS]

overall_design: DNA methylation analysis using RRBS of 10T cells, TKO mESC, and Normal and SCC Esophogus Organoids with different DNMT3A1 mutations

middle: H

first: Chao

last: Lu


title: Epigenetic reader ZMYND11 noncanonical function restricts HNRNPA1-mediated stress granule formation and oncogenic activity

status: Public on Aug 26 2024

city: shanghai

first: cheng

last: Lian

address: dongan road 131

institute: Fudan university

summary: To investigate the function of ZMYND11 in  prostate cancer, we knockdown ZMYND11 in 22Rv1 cells and do RNA-seq.

overall_design: We then performed gene expression profilling analysis using data obtained from RNA-seq of 22Rv1 cells.

first: Gong-Hong

last: Wei

first: Cheng

first: Chunyi

last: Zhang

first: Zixian

last: Wang


title: Isolation and identification of patient-derived liver cancer stem cells and development of personalized treatment strategies

status: Public on Aug 21 2024

city: Guang Zhou

first: Tingting

last: Guo

state: Guang Dong

address: University Town Campus (South Campus): 382 Zhonghuan Road East, Panyu District, Guangzhou Higher Education Mega Centre,

institute: South China University of Technology

summary: Liver cancer is a devastating disease characterized by its molecular complexity and the insidious behavior of liver cancer stem cells (LCSCs), which drive the metastasis and recurrence. However, the heterogeneity of molecular markers of LCSCs has hindered the development of effective methods to isolate them. This study introduced a pioneering approach to isolate and culture LCSCs from primary tumor tissues, leveraging mouse embryonic fibroblasts (MEFs) as a feeder cell to provide the niche in conjunction with using defined medium, which stands in stark contrast to traditional techniques, and promised to revolutionize our understanding of LCSCs. Total seventeen cell strains were directly isolated from 24 patients’ tissues without sorting, seven of which could be maintained long-term culture as colony growth on MEFs, which is unique characteristics of stem cells. Cloned cells formed a round packed morphology on MEFs and could be cryopreserved and re-cultured. Even 10 of cloned cells could form xenografts of human liver cancers in immunodeficient mice, indicating high tumorigenicity. Thus, cloned cells could be designated as liver cancer stem cells. Our further investigation found that reported putative liver cancer stem cell markers were all expressed in cloned cells, and stemness of cancer stem cells could well maintained under our developed culture conditions when compared to those under traditional suspension conditions. Furthermore, our findings revealed a distinct molecular landscape within hepatocellular carcinomas (HCCs), pinpointing a C2 subtype with profound implications for personalized medicine and the development of targeted therapies. By illuminating the unique responses of this subtype to chemotherapeutic agents, our research not only advances the frontiers of liver cancer biology but also charts a course for more effective, patient-specific treatment strategies.

overall_design: ALDH-positive HCC2 CSCs were cultured in three different ways: on MEFs, in suspension conditions, and in differentiation conditions using DMEM medium containing 10% FBS.

first: Yuyou


title: Comparative analysis between single-cell RNA-seq and single-molecule RNA FISH indicates that the pyrimidine nucleobase idoxuridine (IdU) globally amplifies transcriptional noise

status: Public on Aug 28 2024

city: Miami

first: Binyamin

last: Zuckerman

state: Florida

department: Department of Cell and Systems Biology

institute: University of Miami

summary: Stochastic fluctuations (noise) in transcription generate substantial cell-to-cell variability, but the physiological roles of noise have remained difficult to determine in the absence of generalized noise-modulation approaches.  Previous single-cell RNA-sequencing (scRNA-seq) suggested that the pyrimidine-base analog (5′-iodo-2′-deoxyuridine, IdU) could generally amplify noise without substantially altering mean-expression levels but scRNA-seq technical drawbacks potentially obscured the penetrance of IdU-induced transcriptional noise amplification.  Here we quantify global-vs.-partial penetrance of IdU-induced noise amplification by assessing scRNA-seq data using numerous normalization algorithms and complement our previous results with data from Human Jurkat T Lymphocytes, including two biological replicates to allow rigorous statistical analysis.  Collectively, this analysis indicates which scRNA-seq algorithms are appropriate for quantifying noise and argues that IdU is a globally penetrant noise-enhancer molecule that could enable investigations of the physiological impacts of transcriptional noise in various cell types and organisms.

overall_design: Jurkat cells were treated with either DMSO or 20 uM IdU for 48h. Two biological replicates were performed, resulting in total four samples of scRNA-seq.

first: Leor

last: Weinberger


title: Cancer-associated DNA Hypermethylation of Polycomb Targets Requires DNMT3A Dual Recognition of histone H2AK119 Ubiquitination and the Nucleosome Acidic Patch [EM-Seq]

first: Kristjan

overall_design: DNA methylation analysed in DNMT1/3A/3B/3L (QKO) mESC expressing different DNMT3A1 mutants using  EnzymaticMethyl-seq (EM-seq)


title: Cancer-associated DNA Hypermethylation of Polycomb Targets Requires DNMT3A Dual Recognition of histone H2AK119 Ubiquitination and the Nucleosome Acidic Patch [Cut&Tag]

overall_design: Cut&Tag of H3K27me3, H2AK119Ub and H3K4me3 in Normal and SCC Esophogus Organoids


title: Defining the nature of human pluripotent stem cell-derived interneurons via single cell analysis

status: Public on Aug 09 2024

city: Santa Clara

first: Marcos

last: Otero-Garcia

address: 2255 Martin Ave suite D

institute: Rebus Biosystems

summary: sc-RNA-Seq profiling of human brain nuclei

overall_design: Human brain cortex nuclei were isolated and subjected to sc-RNA-seq

first: William

last: Lowry

first: Inma

last: Cobos

first: Kathrin

last: Plath

last: Otero

first: Tom

last: Allison


title: Fetal hypoplastic lungs have multilineage inflammation that is reversed by amniotic fluid stem cell extracellular vesicle treatment

city: Toronto

first: Lina

last: Antounians

state: Ontario

address: 686 Bay St

institute: The Hospital for Sick Children

summary: We performed single nucleus RNA-seq on rat fetal lungs from: control (normal), hypoplastic (injury), and hypoplastic+ amniotic fluid stem cell extracellular vesicles treated (injury+treatment).

overall_design: Single nucleus RNA-seq on rat fetal lungs at E21.5. Conditions: 1) control (normal), 2) hypoplastic (injury), and 3) hypoplastic+ amniotic fluid stem cell extracellular vesicles treated (injury+treatment)


title: The proteomic landscape and temporal dynamics of human and mouse gastruloid development

status: Public on Sep 06 2024

city: Seattle

first: Riddhiman

last: Garge

state: WA

address: Foege Building S-210, 3720 15th Ave NE

department: Genome Sciences

institute: University of Washington

summary: Gastrulation is the highly coordinated process by which the early embryo breaks symmetry, establishes germ layers and a body plan, and sets the stage for organogenesis. As early mammalian development is challenging to study in vivo, stem cell-derived models have emerged as powerful surrogates, e.g. human and mouse gastruloids. However, although single cell RNA-seq (scRNA-seq) and high-resolution imaging have been extensively applied to characterize such in vitro embryo models, a paucity of measurements of protein dynamics and regulation leaves a major gap in our understanding. Here we sought to address this by applying quantitative proteomics to human and mouse gastruloids at four key stages of their differentiation (naïve ESCs, primed ESCs, early gastruloids, late gastruloids). To the resulting data, we perform network analysis to map the dynamics of expression of macromolecular protein complexes and biochemical pathways, including identifying cooperative proteins that associate with them. With matched RNA-seq and phosphosite data from these same stages, we investigate pathway-, stage- and species-specific aspects of translational and post-translational regulation, e.g. finding peri-gastrulation stages of human vs. mice to be discordant with respect to the mitochondrial transcriptome vs. proteome, and nominating novel kinase-substrate relationships based on phosphosite dynamics. Finally, we leverage correlated dynamics to identify conserved protein networks centered around congenital disease genes. Altogether, our study showcases the potential of intersecting proteomics and in vitro embryo models to advance our understanding of early mammalian development in ways that may not be possible through transcriptomics alone.

overall_design: To map the temporal changes underlying gastruloid development, we performed multi-omic phenotyping on 4 stages of gastruloid differentiation namely Naive ESCs, Primed ESCs, Early gastruloids and Late gastruloids.

first: Valerie

last: Lynch

first: Rose

last: Fields

first: Silvia

last: Casadei

first: Sabrina

last: Best

first: Jeremy

last: Stone

first: Matthew

last: Snyder

first: Connor

last: Kubo

first: Arata

last: Wakimoto

first: Zukai

last: Liu

first: Chris

last: McGann

first: Jay

last: Shendure

first: Lea

last: Starita

first: Nobuhiko

last: Hamazaki

first: Devin

last: Schweppe


title: Cancer-associated DNA Hypermethylation of Polycomb Targets Requires DNMT3A Dual Recognition of histone H2AK119 Ubiquitination and the Nucleosome Acidic Patch [ChIP-seq]

first: Kristjan

last: Gretarsson

overall_design: Comparative gene expression analysis at different timepoints during adipocyte differentiation for all DNMT3A1 mutants.

first: Chao

last: Lu


title: A developing HPA axis permits neonatal immune sensitivity to common allergens [scRNA-Seq]

status: Public on Jul 30 2024

city: New York

first: Dr. Shruti

last: Naik

state: NY

address: 1425 Madison Ave

department: Immunology and Immunotherapy

institute: Icahn School of Medicine at Mount Sinai

summary: Atopic and allergic diseases predominantly manifest in early life, a period of profound growth and maturation. Yet, there is a dearth of information on the impact of developmental parameters on the neonatal immune reaction to allergens. Here we demonstrate that common allergens including house dust mite (HDM) elicited potent type 17 inflammatory responses from neonatal but not adult mice. HDM-induced inflammation was abolished in Rorgt-, gd T cell-, or epithelial IL-17RC- deficient animals and surprisingly did not require nociceptors or lymph node engagement. Instead, neonatal CD301b+ type 2 conventional dendritic cells (cDC2s) abundantly engulfed allergen and locally activated gd T cells in the skin. Despite uptaking equivalent antigen to neonates, adult cDC2s did not promiscuously activate in the skin. Mechanistically, this discrepancy was not attributable to cell-intrinsic differences arising from the distinct developmental origins of neonatal and adult cDC2s. Instead, neonatal cDC2 hyperactivation could be traced to a lower systemic glucocorticoid level, a consequence of an immature hypothalamic-pituitary-adrenal (HPA) axis in early life, and could be reversed by cortisol administration at the time of allergen challenge. These findings bring to fore the influence of organismal developmental factors in shaping immune sensitivity to allergens in early life.

overall_design: To dissect the baseline composition of  antigen presenting cell populations  we isolated immune cells from neonate and adult dorsal skin and performed single cell RNA sequencing. To characterize the activation of dendritic cells in house dust mite (HDM) treated neonate skin, we performed single cell RNA sequencing with neonate skin cells isolated 9 hours after PBS and HDM inoculation.

first: Yue

last: Xing

first: Ikjot

last: Sidhu

first: Aleksandr

last: Prystupa

first: Shruti


title: Cancer-associated DNA Hypermethylation of Polycomb Targets Requires DNMT3A Dual Recognition of histone H2AK119 Ubiquitination and the Nucleosome Acidic Patch [RNA-Seq]

overall_design: To investigate the role of DNMT3A1 induced hypermethylation in differentiation we generated 10T cell lines expressing different DNMT3A1 mutants. Comparative gene expression analysis at different timepoints during adipocyte differentiation for all DNMT3A1 mutants.


title: Distance-depending transcriptome changes of pancreatic stellate cells in paracrine pancreatic ductal adenocarcinoma co-culture models

status: Public on Aug 14 2024

city: San Diego

first: Anais

middle: Aurelia

last: Zourelidis

address: 3855 Health Sciences Dr

institute: University of California San Diego

summary: Pancreatic stellate cells (PSC) are one source of cancer-associated fibroblasts (CAF) and play, therefore, an essential role in pancreatic ductal adenocarcinoma (PDA). Paracrine signalling between PDA cancer cells and CAF has been widely studied, yet external influences on paracrine crosstalk are poorly understood. This study aimed to gain a deeper insight into the communication of PSC and cancer cells under different co-culture conditions via analysis of PSC gene expression profiles. Two contactless co-culture models with tumor cells from the p48-Cre; lox-stop-lox-KrasG12D/+; lox-stop-lox-Trp53R172H/+ mouse model (KPC) and murine PSC separated through a microporous membrane and grown in different compartments (standard co-culture) or on different sides of the same membrane (inverse co-culture), were established. RNA-Sequencing analysis of PSC mRNA was performed 24 h and 72 h after co-culture with KPC cells. For selected genes, results were confirmed by quantitative RT-PCR and immunocytochemistry. Standard co-culture displayed 19 differentially expressed genes (DEG) at 24 h and 52 DEG at 72 h. In inverse co-culture, 800 DEG at 24 h and 2213 DEG at 72 h were enriched. PSC showed great heterogeneity in their gene expression profiles; however, mutually regulated genes of both co-cultures, such as VCAN and CHST11, could be identified. VCAN-protein-protein interaction-network analysis revealed several shared genes between co-culture models, such as SDC4 and FN1. In conclusion, PSC show a varying susceptibility to cancer cell signals depending on the co-culture method, with intensified transcriptome changes with closer proximity.

overall_design: To investigate PSC gene expression under different conditions, we developed two contactless, paracrine co-culture models: in standard-co culture, PSC grew on top of a microporous membrane with 1 μm pores, with KPC cells growing in the lower compartment, in inverse co-culture PSC and KPC cells were grown on different sides of the same microporous membrane. RNA-Seq was performed from PSC at two different time points, 24 h and 72 h.

first: Bogusz

last: Trojanowicz

first: Yoshiaki

last: Sunami

first: Gerd

last: Hause

first: David

last: Vieweg

first: Joerg

last: Kleeff


title: ZFP57 is a regulator of postnatal growth and life-long health

status: Public on Jul 31 2024

first: Kevin

middle: Richard

last: Costello

address: Genetics Building Downing St.

department: Department of Genetics

institute: Univerity of Cambridge

summary: Early-life factors, including nutrition, shape long-term health outcomes. Despite the essential role of lactation in maternal nutritional support, the influence of epigenetic factors on lactation and postnatal growth remains poorly understood. Zinc-finger protein 57 (ZFP57), is an epigenetic regulator of genomic imprinting, a process that directs gene expression based on parental origin, playing a vital role in mammalian prenatal growth.   Here, we identify a novel function of ZFP57 in the mammary glandI, where it serves as a key modulator of postnatal resource control, independently of imprinted genes.   ZFP57 regulates multiple aspects of mammary gland functions, including ductal branching and cellular homeostasis. Its absence leads to significant differential gene expression, related to alveologenesis, lactogenesis and milk synthesis, associated with delayed lactation and altered milk composition. This results in life-long impacts on offspring including the development of metabolic syndrome.  Cross-fostering reveals intricate dynamics between mother and offspring during lactation.  Pups raised by a dam of a different genotype than their birth mother exhibit exacerbated metabolic features in adulthood, providing additional novel insight into the programming of offspring long-term health by maternal context.  This study deepens our understanding of the interplay between epigenetic factors, lactation, and postnatal resource control.

overall_design: Mammary tissue from Wild type and ZFP57 KO mice were collected at various stages during pregancy and sorted into descreet cell population. RNA from each of these cell types were the harvested and sequenced to profile in impact ZFP57 KO has on transcriptional regulation of mammary cells during pregancy

first: Geula

last: Hanin

first: Boshra

last: Alsulaiti

first: Hugo

last: Tavares

first: Nozomi

last: Takahashi

first: Liudmila

last: Mikheeva

first: Anjuli

last: Freeman

first: Shrina

last: Patel

first: Albert

last: Koulman

first: Anne

last: Ferguson-Smith


title: Role of the STAR proteins in early mammalian development

status: Public on Aug 02 2024

city: Barcelona

first: Elias

last: Bechara

address: Dr Aiguader 88

country: Spain

institute: CRG

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

overall_design: Refer to individual Series


title: Cancer-associated DNA Hypermethylation of Polycomb Targets Requires DNMT3A Dual Recognition of histone H2AK119 Ubiquitination and the Nucleosome Acidic Patch

first: Kristjan

last: Gretarsson


title: The transcriptional signatures of Caenorhabditis elegans dauer larvae

status: Public on Aug 15 2024

city: Buffalo

first: Johnny

last: Cruz Corchado

address: Elm & Carlton Streets Buffalo,L3-L125

department: Cell Stress

institute: Roswell Park Comprehensive Cancer Institue

summary: Both plasticity and robustness are pervasive features of developmental programs.  The dauer in Caenorhabditis elegans is an alternative to the third larval stage of the nematode and is an example of phenotypic plasticity. The dauer is an arrested, hypometabolic state that undergoes dramatic changes in gene expression compared to conspecifics that continue development, and can be induced by several adverse environments or genetic mutations that act as independent and parallel inputs into the larval developmental program. However, given the different genetic or environmental triggers that can induce dauer, gene expression in dauer larvae could be invariant or vary depending on the larvae’s route into dauer entry; this question has not been examined. Here we use RNA-sequencing to characterize gene expression in dauer larvae induced to arrest development in response to different stimuli.  By assessing the variance in the expression levels of all genes and computing the Spearman's rank-order correlation of gene expression within several Gene Ontologies (GO) and gene networks, we find that the expression patterns of most genes, except for those that act in specific defense and metabolic pathways, are strongly correlated between the different dauer larvae, suggestive of transcriptional robustness.  We speculate that the transcriptional robustness of core dauer pathways allows for the buffering of variation in the expression of genes involved in their response to the environment, allowing the different dauers to be better suited to survive in and exploit different niches.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for  C. elegans dauer of the following gennotypeswild-type dauers (N2; Bristol) ;daf-2 (e1370) III,daf-7(e1372) III ;ilc-17.1 (syb5296) X  ;cep-1 (cep-1oe) [unc-119 (ed4); gtIs1 [CEP-1::GFP + unc-119 (+)]

last: Cruz

first: Abhishiktha

last: Godthi

first: Veena

last: Prahlad


title: Transcriptome analysis of high-risk cattle supplemented with inorganic or organic Co, Cu, Mn, and Zn

status: Public on Jul 26 2024

first: Matthew

last: Scott

summary: This objective of this study was to identify gene expression and biological mechanisms related to trace mineral metabolism, central to inflammatory mediation and disease development, through the evaluation of the whole blood transcriptome in post‐weaned cattle receiving different mineral treatments. Here, 56 high‐risk cross-bred beef cattle (~250 kg) were assigned to receive ad libitum access to supplements containing 1 of 3 treatments over a 60-day period: 1) sulfate sources of Cu, Co, Mn, and Zn in feed (INR; custom blend; n= 40); 2) organic complexed source of Cu, Mn, Co, and Zn in feed (AAC; Availa4; n = 40); or 3) organic complexed source of Cu, Mn, Co, and Zn in feed and administration of a drench product [ProFusion] upon arrival (COMBO; n = 40). Whole blood RNA from 26 cattle (~4 calves/pen) were matched and utilized to assess the host transcriptome at days 0, 13, 28, 45, and 60 for the assessment of host transcriptome responses over time.

overall_design: Whole blood from 26 randomly selected and matched individuals per treatment group (n=3;INR, AAC, COMBO) were used for gene expression profiling and differential analysis across five time points (Day 0, 13, 28, 45, 60).

first: Kelsey

last: Harvey

first: Brandi

last: Karisch

first: Amelia

last: Woolums


title: ROS1 regulates blueberry fruit ripening in a negatively ABA manner

status: Public on Jul 19 2024

city: Beijing

first: Zejia

last: Wang

address: Yanyuan Street

institute: Peking University

relation:
SuperSeries of: GSE229139
SuperSeries of: GSE229140


title: Epigenetic heterogeneity hotspots in human liver disease progression [MethylationEPIC Beadchip]

city: Rochester

first: Keith

middle: Darrell

last: Robertson

state: MN

address: 200 First Street SW, Stabile 12-70

department: Pharmacology

institute: Mayo Clinic

summary: Disruption of the epigenome is a hallmark of human disease including liver cirrhosis and hepatocellular carcinoma (HCC). While genetic heterogeneity is an established effector of pathologic phenotypes, epigenetic heterogeneity is less well understood. Environmental exposures including hepatitis C viral (HCV) infection and metabolic syndrome alter the liver-specific DNA methylation landscape and influence the onset of liver cancer. Given that currently available treatments are unable to target the most frequently mutated genes in HCC, there is an unmet need for novel therapeutic strategies to prevent or reverse hepatic tumorigenesis, which the epigenome may provide.  We performed genome-wide profiling of DNA methylation, copy number, and gene expression landscapes from multiple liver regions from 31 patients with liver disease to examine their crosstalk and define the individual and combinatorial contribution of these processes to liver disease progression.  We identify epigenetic heterogeneity hotspots that are conserved across patients. Elevated epigenetic heterogeneity associates with increases in gene expression heterogeneity. Cirrhotic regions comprise two distinct cohorts, one exclusively epigenetic, and a second where epigenetic and copy number variations collaborate. Epigenetic heterogeneity hotspots are enriched for genes central to liver function (e.g., HNF1A) and known tumor suppressors (e.g., RASSF1A). These hotspots encompass genes including ACSL1, ACSL5, MAT1A, and ELFN1, which have phenotypic effects in cell line functional screens, supporting their relevance to hepatocarcinogenesis.   Substantial epigenetic heterogeneity arises early in liver disease development, targeting key pathways in the progression and initiation of both cirrhosis and HCC. Integration of epigenetic and transcriptional heterogeneity unveils putative epigenetic regulators of hepatocarcinogenesis.

overall_design: Genome-wide DNA methylation from multi-region sampling in normal and liver tissues

first: Ryan

last: Hlady


title: Aromatase in adipose tissue exerts osteoprotective function in male mice via phosphorus regulation

status: Public on Jul 22 2024

city: Toon

first: Yuuki

last: Imai

state: Ehime

address: Shituskawa

department: Proteo-Science Center

institute: Ehime University

summary: Aromatase contributes to the maintenance of bone mass because male patients with loss-of-function mutations of CYP19A1 exhibit bone loss. Furthermore, treatment with aromatase inhibitor also causes bone loss in men as well as post-menopausal women, in whom serum estrogen level is very low, suggesting that a part of the anabolic effects of testosterone in men is dependent on estradiol (E2) biosynthesized by aromatase in non-gonadal tissues. However, it remains unclear how locally biosynthesized E2 contributes to the maintenance of bone mass. We examined the function of aromatase in local tissues rather than gonad using cell type specific aromatase KO mice. Because osteoblast-specific aromatase KO mice exhibited no bone phenotype, we focused on adipose tissue, which is known as reservoir of steroid hormones, and analyzed the bone phenotypes of adipose tissue-specific aromatase KO mice (AroΔaP2). Sixteen-week-old male AroΔaP2 mice exhibit significantly lower bone mineral density in tibia and femur, especially in trabecular bone compared to the control. Bone histomorphometry showed that AroΔaP2 mice exhibited an osteomalacia-like bone phenotype with increased osteoid volume and width, and decreased osteoclast area and numbers. Moreover, serum PI and renal phosphorus reabsorption were significantly decreased but FGF23 level was decreased in AroΔaP2, suggesting that an osteomalacia-like phenotype in AroΔaP2 was not caused by abnormal FGF23 level. Finally, we analyzed NaPi2a and NaPi2c, a phosphate transporter localized in the kidney, and found that protein levels in renal brush border membrane vesicles were decreased in AroΔaP2. These results indicate that locally biosynthesized estrogens by aromatase in adipocytes can play a significant role in bone mass maintenance via regulate phosphorus reabsorption in kidney by NaPi2.

overall_design: To clarify target gene (s) of Cyp19a1 in adipose tissue that regulate phosphorus metabolism, we performed RNA-seq analysis using total RNA from eWAT in AroΔaP2 and Aroflox/flox.

first: Aoi

last: Ikedo


title: Col2a1-Cre;Idh1LSL/+ embryonic growth plate chondrocytes - single cell RNA-sequencing

status: Public on Jun 28 2024

city: Madison

first: Jianhong

last: Ou

state: Wisconsin

address: 330 N Orchard Street

institute: Morgridge Institute For Research

summary: To determine differences between Idh1 mutant and control growth plates, we used scRNA sequencing. Three E18.5 growth plates from our Idh1 mutant mice and three control mice were studied. Differential gene expression analysis revealed genes that were expressed in the distinct clusters.

overall_design:
In order to assess the Idh1 mutant expressing chondrocytes, we utilized Collagen2a1-Cre males crossed with Idh1 LSL/+ females. Growth plate chondrocytes from E18.5 distal part of femur and proximal part of tibia were collected and chondrocytes were harvested using Pronase I digestion followed by Collagenase IV. Live cells were sorted and GEM were made using 10xgenomics kit. After sequencing and downstream analysis using Seurat, we observed distinct clusters of cells with Idh1 mutation
10X Genomics Single cell RNA-sequencing of growth plate chondrocytes

first: Vijitha

last: Puviindran

first: Ze

middle: Y

last: Huang

first: Yu

last: Xiang

first: Hongyuan

last: Zhang

first: Ga

middle: I

last: Ban

first: Xiaolin

last: Wei

first: Makoto

last: Nakagawa

first: Xinyi

last: Ma

first: John

last: Martin

first: Yarui

last: Diao

first: Benjamin

last: Alman


title: Enrichment of various epigenetic histone marks in human myoblasts

status: Public on Jun 25 2024

first: Jeffrey

last: Dilworth

email: jdilworth@ohri.ca, fdilworth@wisc.edu

state: WI

address: 1111 Highland Ave - WIMR II 4518

department: Cell and Regenerative Biology

institute: University of Wisconsin - Madison

summary: WS234 cells are an immortalized human cell line. Here we have explored the distribution of histone marks (H3K27me3, H3K27ac, H3K4me3, H3K4me1, and  H3K9me3) across the genome in proliferative conditions.

overall_design: WS234 cells were grown in culture media for 48 hours and cells were trypsinized, counted and used for CUT&Tag studies. Two biological replicates per groups (total 12 samples) were subjected to CUT&Tag analysis using H3K27me3, H3K27ac, H3K4me3, H3K4me1, and  H3K9me3 or IgG antibodies. Accumulation of modified histones across the genome was analyzed.

first: Sarah

last: Hachmer

first: Matthew

last: Wong

first: Eric

last: Arezza

first: Lynn

last: Megeney

first: F


title: Epigenetic and 3D genome reprogramming during the aging of human hippocampus

status: Public on Dec 05 2024

city: La Jolla

first: Bing

last: Ren

state: CA

address: 9500 Gilman Drive

institute: UCSD

summary: Age-related cognitive decline in humans is associated with altered physiology of the hippocampus, a region critical for memory consolidation and spatial navigation. While changes in gene expression have been observed in aging brain cells, our understanding of the regulatory mechanisms underlying these changes and their connection to chromatin structure remains limited. To unravel these complexities, we have applied a comprehensive multi-omics approach, integrating single-nucleus gene expression, chromatin accessibility, DNA methylation, and 3D chromatin architecture data obtained from the hippocampi of 40 neurotypical human donors spanning the adult lifespan. We observed a striking loss of astrocytes, oligodendrocyte precursor cells, and endothelial cells during aging. In particular, we observed a substantial decline in the subset of astrocytes that play a role in maintaining and regulating synaptic transmission. In addition, microglia undergo a dramatic switch from a predominantly homeostatic state to an epigenetically primed inflammatory state that coincides with reprogramming of their DNA methylomes and activation of endogenous retrotransposons. In the aged cells, we detected erosion of the 3D genome architecture, where local chromatin domains are diminished and trans-chromosomal interactions are strengthened. Importantly, age-related changes in chromatin folding have high correspondence with the transcriptome and epigenome in multiple cell types. Our data identifies age-associated changes in cell types/states and gene regulatory features that provide insight into the loss of synapses and cognitive decline that occurs in the human brain during aging.

overall_design: Analysis of single nucleus multiome ATAC-seq + RNA-seq (10x Genomics) of hippocampus from human

first: Nathan

last: Zemke

first: Seoyeon

last: Lee


title: SMCHD1 activates the expression of genes required for the expansion of human myoblasts (RNA-seq)

summary: In human muscle, SMCHD1 mutations are associated with the onset of FSHD2, but the mechanism driving the disease onset remains unclear. A commonly accepted explanation is the loss of SMCHD1 binding to the D4Z4 locus activates the expression of DUX4 in FSHD2 muscle. In this study, we used human myoblasts having DUX4 non-permissive 4qB alleles as a model to study DUX4-independent functions of SMCHD1 on myoblast cell growth. Surprisingly, depletion of SMCHD1 in these cells resulted in a cell proliferation defect. Despite the absence of DUX4 target genes’ activation, these cells showed a repression of PAX7 target genes (a hallmark of FSHD) and similar changes in expression profile compared to FSHD myoblasts. Interestingly, downregulation of cell proliferation-related genes and dysregulation of fibroblasts-specific genes were observed in SMCHD1 knockdown myoblasts and FSHD2 myoblasts but not FSHD1 myoblasts. Additionally, we identified LAP2 as direct targets of SMCHD1. Depletion of LAP2 leads to cell proliferation defect similar to the effect after SMCHD1 knockdown. These data imply that DUX4 is not the only driver for the onset of FSHD, and SMCHD1 has DUX4-independent functions in muscle growth and development.

overall_design: Transcriptome profiling of proliferating human myoblast cell line WS234 (donor 15VBic) upon lentiviral transduction of non-silencing scramble shRNA or shRNA targeting SMCHD1


title: Control of innate olfactory valence by segregated cortical amygdala circuits

status: Public on Jul 17 2024

first: Cory

middle: Matthew

last: Root

address: 3119 Pacific Hall, 9500 Gilman Drive

department: Biological Sciences

institute: University of California, San Diego

summary: Animals perform innate behaviors that are stereotyped responses to specific evolutionarily relevant stimuli in the absence of prior learning or experience. These behaviors can be reduced to an axis of valence, whereby specific odors evoke approach and avoidance. The cortical amygdala (plCoA) mediates innate attraction and aversion to odor. However, little is known about how this brain area gives rise to behaviors of opposing motivational valence. Here, we sought to define the circuit features of plCoA that give rise to innate olfactory behaviors of valence. We characterized the physiology, gene expression, and projections of this structure, identifying a divergent, topographic organization that selectively controls innate attraction and avoidance to odor. First, we examined odor-evoked responses in these areas and found sparse encoding of odor identity, but not valence. We next considered a topographic organization and found that optogenetic stimulation of the anterior and posterior domains of plCoA elicits attraction and avoidance, respectively, suggesting a functional axis for valence. Using single cell and spatial RNA sequencing, we identified the molecular cell types in plCoA, revealing an anteroposterior gradient in cell types, whereby anterior glutamatergic neurons preferentially express Slc17a6 and posterior neurons express Slc17a7. Activation of these respective cell types recapitulates appetitive and aversive valence behaviors, and chemogenetic inhibition reveals partial necessity for valence responses to innate appetitive or aversive odors. Finally, we identified topographically organized circuits defined by projections, whereby anterior neurons preferentially project to medial amygdala, and posterior neurons preferentially project to nucleus accumbens, which are respectively sufficient and necessary for innate negative and positive olfactory valence. Together, these data advance our understanding of how the olfactory system generates stereotypic, hardwired attraction and avoidance, and supports a model whereby distinct, topographically distributed plCoA populations direct innate olfactory valence responses by signaling to divergent valence-specific targets, linking upstream olfactory identity to downstream valence behaviors, through a population code.  This represents a novel circuit motif in which valence encoding is represented not by the firing properties of individual neurons, but by population level identity encoding that is routed through divergent targets to mediate distinct valence.

overall_design: Comparative single-cell gene expression profiling of pooled anterior and posterior posterolateral cortical amygdala samples using snRNA-seq

first: James

last: Howe


title: Effects of envelope- or membrane-protein segments of SARS-CoV-2 on gene expression of HUVEC cells

status: Public on Jul 01 2024

city: Suzhou

first: Yiqiang

last: Wang

state: Jiangsu

address: 111 Renai Road

institute: Xi'an Jiaotong-Liverpool University

summary: Exterior segments of E-proteins bound onto and modulated gene expression in human vascular endothelial cells in vitro The finding that EC38 segment laid out of SARS-CoV-2 virus envelope prompted us to wonder if this segment would bind onto and impact the biology of host cells.   Bearing the viremia during severe COVID-19 in mind, more attention was paid to the circulation system.   Cultured human umbilical vein endothelial cells (HUVECs) were used to test the binding of EC38 and MN19 peptides and the potential consequence in terms of gene expression upon such binding.  To check if the binding of interest peptides onto HUVEC concurred functional consequences in the cells, HUVECs were treated with EC38 and MN19 peptides (5μM) for 24 hours, and their transcriptome was measured via RNA sequencing.

overall_design: To check if the interested peptides corresponding to segments of envelope (E-) or membrane (M-) proteins of SARS-CoV-2 would stimulate changes of transcriptome in HUVEC cells. HUVECs were treated with EC38 and MN19 peptides (5μM) for 24 hours, and their transcriptome was measured via RNA sequencing.

first: Jie

last: Xu


title: In vivo CRISPR base editing for treatment of Huntington’s disease

status: Public on Jul 12 2024

city: Urbana

first: Pablo

last: Perez-Pinera

state: il

address: 1406 West Green Street

institute: University of Illinois Urbana-Champaign

summary: Huntington's disease (HD) is an inherited and ultimately fatal neurodegenerative disorder caused by an expanded polyglutamine-encoding CAG repeat within exon 1 of the huntingtin (HTT) gene, which produces a mutant protein that destroys striatal and cortical neurons. A potentially critical event in the pathogenesis of HD is the proteolytic cleavage of the mutant HTT protein, which leads to fragments of the N-terminal domain of the protein that form toxic aggregates. Given the role that proteolysis of the mutant HTT protein plays in HD, strategies for preventing this process hold potential for treating the disorder. By screening >1401 CRISPR base editors targeting splice elements in the HTT gene, we identify platforms capable of producing HTT protein isoforms resistant to proteolysis via the disruption of the splice acceptor sequence for exon 13. When delivered to the striatum of an HD rodent model, these base editors induced efficient exon skipping and decreased the formation of the HTT protein fragments, which in turn reduced HTT protein aggregation, improved functional deficits and attenuated striatal and cortical atrophy. Collectively, these results illustrate the potential for CRISPR base editing to decrease the toxicity of the mutant HTT protein for HD.

overall_design:
Mammalian cell transfections with CRISPR base editors to modify DNA bases in HTT exon and influence RNA splicing to investigate its impact on caspase-6 induced proteolysis. Stereotaxic injections using AAV delivery into mice brains to study DNA editing and exon skipping using base editing tools

Off-Target analysis for base editing constructs (ABE or CBE) at multiple sites (ie OT 4) compared to GFP transfected controls (ie OT 4C) at the same sites. Also editing rates of genomic DNA in YAC128 mouse striatal tissue between experimental HTT constructs and Rosa controls.

NOTE: The GSM8393900-GSM8393938 samples records have been deleted and new samples (GSM9644802-GSM9644940) have been added on April 3, 2026.

first: Shraddha

last: Shirguppe

first: Michael

last: Gapinske

first: Devyani

last: Swami

first: Kyrollos

last: Shenouda

first: Thomas

last: Gaj


title: Time course of ethylene and ACC response in ETR1 receptor mutants in roots

status: Public on Jun 20 2024

city: Richmond

first: Wright Center for Clinical and Translational Research

last: Bioinformatics Core

state: VA

address: 203 E Cary St

institute: Virginia Commonwealth University

summary: ETR1 receptor mutants etr1-3 (loss-of-function) and etr1-7 (gain-of-function) were treated with either ethylene or its biosynthetic precursor, ACC (1-aminocyclopropane-1-carboxylate) for 0, 1, 4, and 24 hours.

overall_design: WT (Col-0), etr1-3, and etr1-7 seeds were germinated on MS media (with mesh) and grown for 5 days. Then, seedlings were transferred to new MS media. For ACC treatment, new plates contained 0.75uM ACC. For ethylene treatment, seedlings were transferred to control plates and placed into chambers where they were treated with 0.3ppm ethylene. After indicated times (0hr, 1hr, 4hr, 24hr), roots were cut from seedlings and frozen on liquid nitrogen before RNA extraction.

first: Gloria

last: Muday

first: Alexandria

last: Harkey

first: Joëlle

last: Mühlemann

first: Amy

middle: L

last: Olex

first: Maleana

last: Khoury


title: Col2a1-Cre;Idh1LSL/+ embryonic growth plate  - Bulk RNA-sequencing

first: Jianhong

last: Ou

summary: As another method to determine differences between Idh1 mutant and control growth plates, we used Bulk RNA sequencing.  Three E18.5 growth plates from our Idh1 mutant mice and three control mice were studied.  Differential gene expression analysis revealed genes that were  expressed in the distinct clusters, further supporting our single cell RNA data.

overall_design:
In order to assess the Idh1 mutant expressing chondrocytes, we utilized Collagen2a1-Cre males crossed with Idh1 LSL/+ females. Growth plate chondrocytes from E18.5 distal part of femur and proximal part of tibia were collected and snap frozen. Total RNA was isolated and used for RNA-seq analysis.  After sequencing and downstream analysis using Seurat, we observed distinct clusters of cells with Idh1 mutation
Total RNA was isolated using Single Cell RNA Purification Kit and submitted to core facitly for library generation and sequencing

first: Vijitha

last: Puviindran

first: Ze

last: Huang

first: Yu

last: Xiang

first: Hongyuan

last: Zhang

first: Ga

last: Ban

first: Xiaolin

last: Wei

first: Yarui

last: Diao

first: Benjamin

last: Alman


title: A HDAC4-HDAC2 complex composes an epigenetic sensor that links the DNA damage response to the senescent program by erasing H2BK120 acetylation

status: Public on Jun 18 2024

city: Udine

first: Emiliano

last: Dalla

state: UD

address: Piazzale Kolbe 4

department: Department of Medicine

country: Italy

institute: Università degli Studi di Udine