title: transcriptome of purified PC and non-PC fractions: GR-WT & GR-KO

city: Gent

first: Steven

last: Timmermans

address: Technologiepark 71

country: Belgium

institute: VIB-UGent

summary: Paneth Cells (PCs) are secretory cells located in the crypts of Lieberkühn of the small intestine. They produce antimicrobial peptides (AMPs) and are thought to keep the microbiome under control. The cytokine TNF, which functions mainly via its TNF receptor 1 (TNFR1 or P55), is a driving force in many inflammatory bowel disease (IBD) patients, but its impact on PCs is poorly known. We have generated intestinal glucocorticoid receptor knockout mice (GR villin KO) and investigated the diffrences between WT and KO paneth and non-paneth cell fractionf from the cell sorting.

overall_design: cells were isolated form naive mice that were either GRKO or GR WT. Both the PC and non-PC crypt cells were used for sequencing for a total of 4 conditons with 3 replicates each

first: Charlotte

last: Wallaeys

first: Natalia

last: Garcia-Gonzalez

first: Jolien

last: Vandewalle

first: Tineke

last: Vanderhaeghen

first: Hester

last: Dufoor

first: Melanie

last: Eggermont

first: Riet

last: De Rycke

first: Serge

last: Verbanck

first: George

last: Kollias

last: Souffriau

first: Richard

last: Blumberg

first: Takao

last: Iwawaki

first: Claude

last: Libert


title: Engineering Scalable Vascularized Kidney Organoids for in vivo Glomerular Filtration with Human Endothelial Integration

status: Public on Jul 08 2024

city: Charlestown

first: Ryuji

last: Morizane

state: MA

address: 149 13th Street

country: USA

institute: Massachusetts General Hospital

type: Expression profiling by high throughput sequencing

summary: We tested the effects of dynamic cell culture environments in stirred bioreactors (STR) using human kidney organoids. Enhanced glomerular vascularization was observed.

overall_design: Human kidney organoids were differentiated from pluripotent stem cells by our previously published protocol in static culture (Morizane, Nature Biotechnology, 2015). Day 14 organoids were transferred to stirred bioreactors and cultured for three weeks.

first: Murat

last: Tekguc


title: The effect of Stat3 loss on inflammation and endometriosis

status: Public on Aug 01 2024

city: Columbia

first: Jae-Wook

last: Jeong

state: MO

address: 1030 Hitt Street

department: Obstetrics, Gynecology and Women's Health

institute: University of Missouri

summary:
Aberrant STAT3 activation is a major contributor to endometriosis.
We used microarrays to detail the mechanism underlying Stat3 loss.

overall_design: Uteri of Stat3 f/f and Stat3 d/d mice were selected at 3.5 dpc for RNA extraction and hybridization on Affymetrix microarrays.

first: Tae

middle: H

last: Kim


title: Inorganic Ions Activate Lineage-Specific Gene Regulatory Networks to Drive Stem Cells Differentiation

status: Public on Jun 10 2024

city: Bryan

first: Irtisha

last: Singh

state: TX

address: 8447 Riverside Pkwy, Medical Research and Education Building II Suite 4344

department: Department of Molecular and Cellular Medicine

institute: Texas A&M University

summary: Bioactive aqueous inorganic ions facilitate human mesenchymal stem cell lineage commitment by directing biomolecular signaling

overall_design: Investigation of ions' impact on stem cell fate and molecular signaling cascades

first: Anna

middle: L

last: Kersey

first: Akhilesh

middle: K

last: Gaharwar


title: Microglia contribute to polyG-dependent neurodegeneration in neuronal intranuclear inclusion disease

status: Public on Jul 04 2024

city: Shanghai

first: Yangye

last: Lian

address: Xuhui No.180, Fenglin Road

country: China

institute: Zhongshan Hospital affiliated to Fudan University

summary: Neuronal intranuclear inclusion disease (NIID) is a neurodegenerative disease caused by GGC repeat expansion of NOTCH2NLC. Despite the identification of uN2CpolyG, a toxic polyglycine (polyG) protein, the pathogenic mechanisms of NIID remain unclear. Herein, we investigated the role of polyG by expressing wild-type NOTCH2NLC, expanded NOTCH2NLC with 100 GGC repeats (translating or not translating into uN2CpolyG), or mutated NOTCH2NLC that encodes a pure polyG without GGC-repeat RNA and C-terminal stretch (uN2CpolyG-dCT) in mice. Both uN2CpolyG and uN2CpolyG-dCT induced the formation of inclusions composed by filamentous materials and caused neurodegenerative phenotypes in mice, including impaired motor and cognitive performance, shortened lifespan, and pathological lesions such as white matter lesions, microgliosis, and astrogliosis. By contrast, the sole expression of GGC-repeat RNA was non-pathogenic. Combining with bulk and single-nuclei RNA-seq, we determined the common molecular signatures associated with expressing uN2CpolyG and uN2CpolyG-dCT in the brain, especially the up-regulation of inflammation and microglia markers whereas down-regulation of immediate early genes and splicing factors. Importantly, microglia-mediated inflammation was visualized in NIID patients by positron emission tomography, which correlated with white matter atrophy levels. Moreover, microglia ablation ameliorated neurodegeneration in uN2CpolyG-expressing mice but did not alter polyG inclusions. Together, these results demonstrate that polyG is critical for the pathogenesis of NIID, and microglia contribute to polyG-induced neurodegeneration.

overall_design: To analyze molecular characteristics of different cell populations, we performed single nuclei RNA-seq (snRNA-seq) on cerebral cortices of  ATG-GGC11 (n=2, 3A6)  and ATG-GGC100 (n=3,3A7) mice groups.

first: Shaoping

last: Zhong


title: Transcriptomic analysis reveals sex specific genes similar to human NAFLD associated with transgenerational NAFLD

status: Public on Jun 12 2024

first: Ramji

last: Bhandari

address: 1200 University Ave

department: Division of Biological Sciences

summary: Comparison of bulk RNA-Seq data from the livers of male and female medaka fish, in the BPA lineage vs control lineage of f2 generation, with publicly available human NAFLD-NASH patient data revealed that 20% of DEGs were mutually upregulated with those found in human NAFLD patients. Additionally, 27.69% of DEGs were mutually downregulated, and 52.31% of ancestral BPA-specific DEGs were expressed in the livers of female fish in the BPA lineage. The activation of myc, atf4, xbp1, and stat4, as well as the inactivation of igf1, were associated with pathogenesis.

overall_design: RNA seq of the liver of biological replicates from BPA lineage and control lineage of male and female Medaka fish. Three biological replicates (3 control + 3 BPA treated) for female fish and two biological replicates (2 control + 2 BPA treated) for male fish were used.

first: Sourav

last: Chakraborty

first: Santosh

last: Anand


title: Comparative transcriptomic profiling between two radiologic phenotypes in lungs from patients with pulmonary disease caused by Mycobacterium avium complex

status: Public on Jul 01 2024

city: Seoul

first: Su-Young

address: 81, Irwon-ro, Gangnam-gu

country: South Korea

institute: Samsung Medical Center

summary: The incidence of Mycobacterium avium complex (MAC)-induced pulmonary disease (PD) has been increasing in South Korea, posing considerable challenges in treatment. Treatment guidelines recommend initiating therapy after serial computed tomography (CT) monitoring. While drug treatment often improves outcomes in patients with the nodular bronchiectatic (NB) form, those with the fibrocavitary (FC) form may continue to experience persistence despite treatment. Our objective was to classify type-specific and common genes based on radiological phenotypes (NB and FC) using RNA sequencing to analyze the cellular landscape of 11 NB sections, 10 FC sections, and 21 unaffected sections from 21 MAC-PD cases. Predictive in silico analysis indicated a significant role for genes related to B cell proliferation and regulation in both forms. Additionally, NB patients exhibited predominantly regulated antimicrobial immune responses, whereas FC patients showed enrichment in genes related to extracellular matrix structure. These target genes identified through genetic analysis could potentially serve as predictive biomarkers for type-specific biological processes and cellular pathways, necessitating further validation studies.

overall_design: To examine biological variances between two distinct clinical phenotypes (NB or FC forms) based on unaffected sections, lung tissue was procured in pairs from each patient. Then, one sample originated from an affected lesion, and the other was extracted from an unaffected region. Resection criteria included consideration of the cavity lesion in FC form or the bronchiectasis lesion in NB form, along with the unaffected area.

first: Byung

middle: W

last: Jhun

first: Seong

middle: M

last: Moon

first: Sung

middle: J

last: Shin

first: Ju

last: Lee


title: Microglia contribute to polyG-dependent neurodegeneration in neuronal intranuclear inclusion disease II

overall_design: To explore the transcriptomic alterations induced by polyG proteins, we performed bulk RNA-seq on cerebral cortices of AAV-injected mice. ATG-GGC11 (3A6) , ATG-GGC100(3A7), TAG-GGC100(3A8), ATG-GGN100dCT (3A9).


title: The GPI-anchor biosynthesis pathway is critical for syncytiotrophoblast differentiation and placental development

status: Public on May 19 2024

city: Madrid

first: Vicente

middle: Pérez

last: García

state: Madrid

address: C/ Nicolás Cabrera 1

department: Tissue and organ homeostasis

country: Spain

institute: Centro de Biologia Molecular Severo Ochoa

summary: The glycosylphosphatidylinositol (GPI) biosynthetic pathway in the endoplasmic reticulum (ER) is crucial for generating GPI-anchored proteins (GPI-APs), which are translocated to the cell surface and play a vital role in cell signaling and adhesion. This study focuses on two integral components of the GPI pathway, the PIGL and PIGF proteins, and their significance in trophoblast biology. We show that GPI pathway mutations impact on placental development impairing the development of the syncytiotrophoblast (SynT), and especially the SynT-II layer, which is essential for the establishment of the definitive nutrient exchange area within the placental labyrinth. CRISPR/Cas9 knockout of Pigl and Pigf in mouse trophoblast stem cells (mTSC) confirms the role of these GPI enzymes in syncytiotrophoblast differentiation. Mechanistically, impaired GPI-AP generation induces an excessive unfolded protein response (UPR) in the ER in mTSCs growing in stem cell conditions, akin to what is observed in human preeclampsia. Remarkably, the transcriptomic profile of Pigl- and Pigf-deficient cells separates human patient placental samples into preeclampsia and control groups suggesting an involvement of Pigl and Pigf in establishing a preeclamptic gene signature. Upon differentiation, the impairment of the GPI pathway hinders the induction of WNT signaling for early SynT-II development. Our study unveils the pivotal role of GPI biosynthesis in early placentation and uncovers a new preeclampsia gene expression profile associated with mutations in the GPI biosynthesis pathway, providing novel molecular insights into placental development with implications for enhanced patient stratification and timely interventions.

overall_design: The RNA-seq experiment assesses the transcriptional changes in wild type (Vector) versus Pigl knockout (KO) or Pigf KO mouse trophoblast stem cells at three time points: under stem cell conditions at 0 days (0d), and after 3 days (3d) and 6 days (6d) of differentiation.

first: Andrea

last: Álvarez-Sánchez

first: Johanna

last: Grinat

first: Paula

last: Doria-Borrell

first: Maravillas

last: Mellado-López

first: Érica

last: Pedrera-Alcocer

first: Marta

last: Malenchini

first: Salvador

last: Messeguer

first: Myriam

last: Hemberger

last: Pérez-García


title: PanethcellTNF-signalinginducesbacterialsepsis: transcriptome of intestinal epithelial cells 8h after TNF

status: Public on Jun 05 2024

first: Steven

last: Timmermans

summary: Paneth Cells (PCs) are secretory cells located in the crypts of Lieberkühn of the small intestine. They produce antimicrobial peptides (AMPs) and are thought to keep the microbiome under control. The cytokine TNF, which functions mainly via its TNF receptor 1 (TNFR1 or P55), is a driving force in many inflammatory bowel disease (IBD) patients, but its impact on PCs is poorly known. We have generated PC-specific P55 knockout mice (P55Paneth KO) and found that these mice are protected against lethal TNF-induced systemic inflammatory response syndrome (SIRS). The impact of TNF on PCs is characterized by morphological changes and by intestinal bacterial translocation into organs, such as liver, in which these bacteria induce expression of numerous genes, associated with typical sepsis signatures. The presence of these bacteria cause lethality of the mice. Based on bulk RNA-SEQ on pure PCs, P55 signaling stimulated mainly a type-I IFN response in the PCs, which drives the bacterial translocation and lethality via IFNAR1. Interestingly, based on ER stress reporter mice, PCs constitutively have high amounts of spliced XBP1 (XBP1s) protein, presumably to support their secretion of AMPs, and TNF eradicates this XBP1s production, leading to failure of the unfolded protein response (UPR). This causes reduced antimicrobial activity in PCs, in a P55 and IFNAR1-dependent way. Our data suggest a novel axis induced by TNF in PCs, leading to interferon-induced UPR failure causing lethal bacterial translocation to the liver (and other organs)

overall_design: To investigate the effect of TNF on the intestinal epithelial cells (IEC) transcriptome, cells  were isolated 8h after TNF or PBS (control) injection in mice that were given normal dinking water of water with 25mM ZnSO4. 2 PBS and 4 TNF samples were taken for RNA isolation and sequencing from each of the water and zinc groups.

first: Charlotte

last: Wallaeys

first: Natalia

last: Garcia-Gonzalez

first: Jolien

last: Vandewalle

first: Tineke

last: Vanderhaeghen

first: Hester

last: Dufoor

first: Melanie

last: Eggermont

first: Riet

last: De Rycke

first: Serge

last: Verbanck

first: George

last: Kollias

last: Souffriau

first: Richard

middle: S

last: Blumberg

first: Takao

last: Iwawaki

first: Claude

last: Libert


title: Role of reducing Hmbox1 in cardiac ischemia/reperfusion injury

status: Public on May 17 2024

city: shanghai

first: Yujiao

last: Zhu

address: Nanchen Road 381

institute: Shanghai University

summary: This RNA-sequencing was performed to determine the differentially expressed mRNA after inhibition of Hmbox1 in cardiac ischemia/reperfusion injury

overall_design: To investigate the function of Hmbox1 in cardiac ischemia/reperfusion (I/R) injury, we established  cardiac I/R injury mouse model in which Hmbox1 was knocked down via adeno-associated virus 9 (AAV9). Total RNAs were extracted from heart tissues from mice at 3 weeks post I/R injury. Four groups were set in this animal experiment (n=3 per group): (1) control AAV9+Sham (Con_Sham); (2) sh-Hmbox1 AAV9+Sham (shHmbox1_Sham); (3) control AAV9+IRI (Con_IRI); (4) sh-Hmbox1 AAV9+IRI (shHmbox1_IRI). The mRNA expression profiling analyses were performed using data obtained from RNA-seq of 4 different groups.

first: Junjie

last: Xiao

first: Yihua

last: Bei


title: Impaired fatty acid import and metabolism in macrophages restricts intracellular growth of Mycobacterium tuberculosis

status: Public on Jun 24 2024

city: Ithaca

first: Nelson

middle: Victor

last: Simwela

state: NY

address: 930 Campus Rd

department: Microbiology and Immunology

institute: Cornell University

summary: Mycobacterium tuberculosis (Mtb) infection of macrophages reprograms cellular lipid metabolism to promote the formation of cytosolic lipid droplets. While it is clearly known that intracellular Mtb utilize host derived fatty acids and cholesterol to fuel a majority of its biosynthetic demands, the role of macrophage lipid metabolism on the bacteria’s ability to access the intracellular lipid pool remains controversial. We have utilized a CRISPR genetic knockdown approach to, systematically and comprehensively, characterize the role of macrophage fatty acid metabolism on the intracellular growth of Mtb. Our analyzes demonstrate that knockdown of lipid import, sequestration and metabolism genes collectively impair the intracellular growth of Mtb in macrophages. We further demonstrate that modulating fatty acids homeostasis in macrophages impair Mtb replication by enhancing production of pro-inflammatory cytokines, restricting the bacteria access to nutrients and increasing oxidative stress in a manner which is surprisingly divergent. We also demonstrate that impaired macrophage lipid droplet biogenesis is restrictive to intracellular Mtb growth. However, increased induction of lipid droplet formation by downstream blockade of fatty acid oxidation does not rescue the impaired Mtb growth phenotypes. Our work provides a characterization of macrophage fatty acid homeostasis and how its modulation impacts Mtb intracellular growth.

sample_organism:
Mycobacterium tuberculosis
Mus musculus

platform_organism:
Mycobacterium tuberculosis
Mus musculus

overall_design: Analysis of the transcriptional profile of both the host and Mtb in  Mtb infected macrophages with impaired fatty acid metabolism genes (Scramble (SC), PLIN2, FATP1 and CPT2)

first: N

middle: V

first: E

last: Jaecklein

first: C

last: Sassetti

first: D

middle: G

last: Russell


title: PanethcellTNF-signalinginducesbacterialsepsis: PC transcriptome 15h ater TNF

status: Public on Jun 03 2024

first: Steven

last: Timmermans

overall_design: To investigate the effect of TNF on the paneth cell (PC) transcriptome, PCs were isolated 15h after TNF or PBS (control) injection in mice using cell sorting. 3 PBS and 4 TNF samples were taken (biological replicates; each from a different mice) for RNA isolation and sequencing

first: Charlotte

last: Wallaeys

first: Natalia

last: Garcia-Gonzalez

first: Jolien

last: Vandewalle

first: Tineke

last: Vanderhaeghen


title: Manipulating the transcriptome and chromatin accessibility in T cells through CRISPR-induced alternative splicing of IKZF1 (RNAseq)

status: Public on Jun 01 2024

city: Gainesville

first: Jeremy

middle: Robert Barker

last: Newman

state: FL

address: 2033 Mowry Road

department: Genetics Institute

institute: University of Florida

summary: Genome-wide association studies have identified statistically significant associations between genetic variants in or near the IKZF1 gene and several autoimmune disorders. IKZF1, encoding the transcription factor Ikaros, produces at least 10 distinct transcripts via alternative splicing.  To explore the role of alternative splicing of IKZF1 in mature T cells, we generated a panel of Jurkat T-cell clones with various truncating mutations in IKZF1 exon 4, exon 6 or both. Differences among clones in gene expression, chromatin accessibility, and protein abundance were assessed by RNA-seq, ATAC-seq and immunoblotting. Clones with single targeting events in either exon 4 or 6 clustered separately from double-targeted clones on multiple parameters, but overall, clone responses were highly heterogeneous. Perturbation of IKZF1 splicing resulted in significant differences in expression and chromatin accessibility of other autoimmunity-associated genes and elicited compensatory expression changes in other IKZF family members. Our results suggest that even modest perturbations of IKZF1 splicing can have significant effects on gene expression and function in mature T cells.

overall_design: Exons 4 and 6 of the human IKZF1 gene in Jurkat cell lines were edited using CRISPR/Cas9. Gene expression profiling was performed using RNA-seq of four WT clones, seven clones with edits in IKZF1 exon 4, five clones with edits in IKZF1 exon 6, and eight clones with edits in IKZF1 exons 4 and 6. Analysis of gene expression was performed comparing WT clones to CRISPR-edited clones based on exons edited and protein detection.

first: Lucia

last: Pastor

middle: R

first: Colin

last: Callahan

first: M

middle: A

last: Atkinson

first: S

last: Onengut-Gumuscu

first: P

last: Concannon


title: Identification and Validation of Small Molecule Inhibitors Targeting FGFR through Molecular Docking-Based Screening

status: Public on Jun 30 2024

city: Wenzhou

first: LIjun

last: Chen

state: Other

address: Wenzhou Medical University, Wenzhou City, Zhejiang Rrovince,China

institute: Wen Zhou Medical University

summary: Fibroblast growth factors and their receptors (FGFRs) play important roles in multiple cellular processes. FGFR genetic alterations such as mutations, amplifications, and chromosomal translocations are prevalent in various cancer types, contributing to the initiation and progression of tumors by enhancing FGFR signaling. Identifying effective small molecule inhibitors that selectively target FGFR can advance cancers therapy driven by FGFR abnormalities. Here, we conducted molecular docking and dynamics simulations to discover new small compounds targeting FGFRs. By docking with 2.8 million small molecules, we identified three promising FGFR inhibitors ranked in the top average absolute difference in free energy. By evaluating the binding stability of the docking pose of these three compounds, we found that ZINC000101867325 formed the most stable binding interactions with FGFRs. After treatment with ZINC000101867325, colorectal cancer cell lines showed a decrease in FGFR phosphorylation and early apoptosis. In addition, ZINC000101867325 is also predicted to interact with FGFR2 mutations in colorectal cancer (CRC) patients. This study provides the novel FGFR inhibitors, and enriches treatment strategies for cancers.

overall_design: We docked approximately 2.8 million small molecule drugs to the crystal structure of FGFR1-4 and predicted three small molecules targeting fgfr. Among them, ZINC000101867325 showed superior bond type and stability. The effects of ZINC000101867325 on FGFR phosphorylation and apoptosis induction in two colorectal cancer cell lines were verified by RNA-seq combined with WB and flow cytometry analysis.In the RNA-seq analysis on cells, ~100 cells were used for per reaction and two or three replicates were performed for each group.

first: Ming

last: Zong

first: Jingzhang

last: Ji

first: Lijun

first: Lixin

last: Zhang


title: Aged skin exacerbates experimental osteoarthritis via enhanced IL-36R signaling

status: Public on May 31 2024

city: Guangzhou

first: Dalin

address: 295 Changxing Road

institute: Guangdong Institute of Orthopedics

summary: To investigate  skin aging is an important driver of experimental osteoarthritis(OA) progression in mice via enhanced IL-36 receptor (IL-36R) signaling. we generated epidermis keratinocyte conditional knockout mice (IL-36Ra-cKO) with topical administration of capsid-mutant Adeno-associated virus 2 (AAV2) vector23 encoding Cre recombinase (AAV2-Cre) to IL-36Rafl/fl mice.  Subsequently, epidermal skin tissues were collected from IL-36Rafl/fl and IL-36Ra-cKO mice for RNA sequencing analysis.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for IL-36Rafl/fl and IL-36Ra-cKO mice epidermal skin tissues

first: Chong

last: Wang

first: Changsheng

last: Yang

first: Liangyu

first: Panpan

first: Zilin

last: Zou

first: Honghao

last: Li

first: Yun

last: Xiao

first: Junfeng

last: Wu

first: Qinwei

last: Cheng

first: Chengliang

first: Ee

last: Ke

first: Bin

last: Huang

first: Xiaochun

last: Bai

first: Kai


title: Manipulating the transcriptome and chromatin accessibility in T cells through CRISPR-induced alternative splicing of IKZF1 (ATACseq)

first: Jeremy

last: Newman

type: Genome binding/occupancy profiling by high throughput sequencing

overall_design: Exons 4 and 6 of the human IKZF1 gene in Jurkat cell lines were edited using CRISPR/Cas9.  Chromatin accessibility profiling was performed using ATAC-seq of four WT clones, seven clones with edits in IKZF1 exon 4, five clones with edits in IKZF1 exon 6, and eight clones with edits in IKZF1 exons 4 and 6. Analysis of chromatin accessibility was performed comparing WT clones to CRISPR-edited clones based on exons edited and protein detection.

first: Lucia

last: Pastor

first: Colin

last: Callahan

first: M

last: Atkinson

first: S

first: P


title: PanethcellTNF-signalinginducesbacterialsepsis: PC transcriptome 15h ater TNF in IFNARKO mice

first: Steven

last: Timmermans

overall_design: To investigate the effect of TNF on the paneth cell (PC) transcriptome, PCs were isolated 15h after TNF or PBS (control) injection in mice using cell sorting. 3 PBS and 3 TNF samples were taken (biological replicates; each from a different mice) for RNA isolation and sequencing. All mice were KO for Ifnar1

first: Charlotte

last: Wallaeys

first: Natalia

last: Garcia-Gonzalez

first: Jolien

last: Vandewalle

first: Tineke

last: Vanderhaeghen

first: Hester

last: Dufoor

first: Melanie

last: Eggermont

first: Riet

last: De Rycke

first: Serge

last: Verbanck

first: George

last: Kollias

first: Richard

last: Blumberg

first: Takao

last: Iwawaki

first: Claude

last: Libert


title: The oocyte microenvironment is altered in adolescents compared to oocyte donors.

status: Public on May 15 2024

city: Chicago

first: Elnur

last: Babayev

state: IL

address: 303 E Superior St 10-220

department: Obstetrics and Gynecology

institute: Northwestern University

summary:
Study question: Are the molecular signatures of cumulus cells (CCs) and follicular fluid (FF) of adolescents undergoing fertility preservation differ from that of reproductively adult oocyte donors?
Summary answer: The microenvironment immediately surrounding the oocyte, including the CCs and FF, is altered in adolescents undergoing fertility preservation compared to oocyte donors.
What is known already: Adolescents experience a period of subfecundity following menarche. Recent evidence suggests that this may be at least partially due to increased oocyte aneuploidy. Reproductive juvenescence in mammals is associated with suboptimal oocyte quality.
Study design, size, duration: This was a prospective cohort study. Adolescents (10-19 years old, N=23) and oocyte donors (22-30 years old, N=31) undergoing ovarian stimulation and oocyte retrieval at the Northwestern Fertility and Reproductive Medicine Center between November 1, 2020 and May 1, 2023 were enrolled in this study.
Participants/materials, setting, method:  Patient demographics, ovarian stimulation, and oocyte retrieval outcomes were collected for all participants. The transcriptome of CCs associated with mature oocytes was compared between adolescents (10-19 years old, n=19), and oocyte donors (22-30 years old, n=19) using bulk RNA-sequencing. FF cytokine profiles (10-19 years old, n=18 vs. 25-30 years old, n=16) were compared using cytokine arrays.
Main results and the role of chance: RNA-seq analysis revealed 581 differentially expressed genes (DEGs) in cumulus cells of adolescents relative to oocyte donors, with 361 genes downregulated and 220 upregulated. Genes enriched in pathways involved in cell cycle and cell division (e.g., GO:1903047, p= 3.5 x 10-43; GO:0051983, p= 4.1 x 10- 30; GO:0000281, p= 7.7 x 10-15; GO:0044839, p= 5.3 x 10-13) were significantly downregulated, while genes enriched in several pathways involved in cellular and vesicle organization (e.g., GO:0010256, p= 1.2 x 10-8; GO:0051129, p= 6.8 x 10-7; GO:0016050, p= 7.4 x 10-7; GO:0051640, p= 8.1 x 10-7) were upregulated in CCs of adolescents compared to oocyte donors. The levels of 9 cytokines were significantly increased in FF of adolescents compared to oocyte donors: IL-1 alpha (2-fold), IL-1 beta (1.7-fold), I-309 (2-fold), IL-15 (1.6-fold), TARC (1.9-fold), TPO (2.1-fold), IGFBP-4 (2-fold), IL-12-p40 (1.7-fold) and ENA-78 (1.4-fold). Interestingly, 7 of these cytokines have known pro-inflammatory roles. Importantly, neither the CC transcriptomes or FF cytokine profiles were different in adolescents with or without cancer.
Large scale data: Original high-throughput sequencing data will be deposited in Gene Expression Omnibus (GEO) before publication, and the GEO accession number will be provided here.
Limitations, reasons for caution: This study aims to gain insights into the associated gamete quality by studying the immediate oocyte microenvironment. The direct study of oocytes is more challenging due to sample scarcity, as they are cryopreserved for future use, but will provide a more accurate assessment of oocyte reproductive potential.
Wider implications of the findings: Understanding the underpinnings of altered immediate oocyte microenvironment of adolescent patients may provide insights into the reproductive potential of the associated gametes in the younger end of the age spectrum. This has implications for the fertility preservation cycles for very young patients.

overall_design: mRNA-seq on human primary cumulus cells microdissected from MII-oocyte associated cumulus-oocyte complexes (COCs) retrieved from adolescents compared to oocyte donors.

first: Dilan

last: Gokyer


title: METI: Deep profiling of tumor ecosystems by integrating cell morphology and spatial transcriptomics

status: Public on May 23 2024

city: Houston

first: Linghua

last: Wang

address: 1881 East Road,

institute: MD Anderson Cancer Center

summary: The recent advance of spatial transcriptomics (ST) technique provides valuable insights into the organization and interactions of cells within the tumor microenvironment (TME). While various analytical tools have been developed for tasks such as spatial clustering, spatially variable gene identification, and cell type deconvolution, most of them are general methods lacking consideration of histological features in spatial data analysis. This limitation results in reduced performance and interpretability of their results when studying the TME. Here, we present a computational framework named, Morphology-Enhanced Spatial Transcriptome Analysis Integrator (METI) to address this gap. METI is an end-to-end framework capable of spatial mapping of both cancer cells and various TME cell components, robust stratification of cell type and transcriptional states, and cell co-localization analysis. By integrating both spatial transcriptomics, cell morphology and curated gene signatures, METI enhances our understanding of the molecular landscape and cellular interactions within the tissue, facilitating detailed investigations of the TME and its functional implications. The performance of METI has been evaluated on ST data generated from various tumor tissues, including gastric, lung, and bladder cancers, as well as premalignant tissues. Across all these tissues and conditions, METI has demonstrated robust performance with consistency.

overall_design: Two muscle-invasive bladder cancer samples (T2-T4) and four gastric adenocarcinoma samples were used for the Visium Spatial transcriptomics.

first: Jiahui

last: Jiang

first: Jian

last: Hu


title: A spatially resolved multiomic single-cell atlas of soybean development

status: Public on Jul 05 2024

city: ATHENS

first: Robert

middle: J.

last: Schmitz

state: GA

address: B416 Davison Life Sciences Department of Genetics University of Georgia 120 East Green Street

department: Genetics

institute: University of Georgia

summary: Cis-regulatory elements (CREs) precisely control the spatiotemporal gene expression in cells. Using a spatially resolved single-cell atlas of gene expression and chromatin accessibility across ten soybean tissues, we identified 103 distinct cell types and 303,199 accessible chromatin regions (ACRs). Nearly 40% of ACRs showed cell-type-specific patterns and were enriched for transcription factor (TF) binding motifs controlling cell-type specification and maintenance. We identified non-cell autonomous activity of NIN-LIKE PROTEIN 7 (NLP7), the Nodule Inception (NIN) gene regulatory network and de novo enriched TF motifs in the nodule infected cells. With comprehensive developmental trajectories for endosperm and embryo, we found that 13 sucrose transporters, sharing the DOF11 binding motif, were co-up-regulated in late peripheral endosperm and identified the key embryo cell-type specification regulators during embryogenesis, including a homeobox TF that promotes cotyledon parenchyma identity. This resource provides a valuable foundation for analyzing gene regulatory programs in soybean cell types across tissues and life stages.

overall_design: We generated a comprehensive accessible chromatin and transcriptome atlas across various cell types in soybean with scATAC-seq, snRNA-seq and spatial-RNA-seq(spRNA-seq), where we collected samples from ten major tissues at different stages of the soybean life cycle. These tissues included leaf, hypocotyl, root, nodule, young pod, and five stages of developing seeds: globular stage (GS), heart stage (HS), cotyledon stage (CS), early maturation stage (EMS), and middle maturation stage (MMS). For each tissue, we conducted scATAC-seq and snRNA-seq with at least two replicates.

first: Xuan

last: Zhang

first: Ziliang

last: Luo

first: Alexandre

middle: P

last: Marand


title: EIF6 conditions drug-tolerant persister-like transdifferentiation in small cell lung carcinoma (eIF6_KD)

status: Public on Apr 30 2024

city: Chengdu

first: Haoning

last: Peng

state: Sichuan

address: No.37 Guoxue Alley, Wuhou District

department: Department of Thoracic Surgery

institute: West China Hospital, Sichuan University

summary: Drug-tolerant persister cells withstand treatments by adapting their identity through lineage-dependent plasticity during systemic anti-cancer therapies. This phenomenon is evident in small-cell lung carcinoma (SCLC), a lethal neuroendocrine cancer initially responsive (60-80%) to platinum-based chemotherapy but succumbing to resistance within 6 months in advanced stages. This resistance associates with the transdifferentiation of residual tumour cells into a non-neuroendocrine state, a process intricately tied to SCLC's chemotolerance, yet molecular mechanisms governing this lineage conversion remain completed understood. Here we use paired cytoplasmic RNA-seq and polysomal RNA-seq to compare gene expression between NE and non-NE SCLC cell lines on both transcriptional and translational level. We report that first-line chemotherapy induces translation initiation factor eIF6 in drug-tolerant persister-like cells in SCLC, correlating with the non-neuroendocrine state in SCLC. Intervening eIF6 inhibits non-neuroendocrine transdifferentiation, thus enhancing SCLC responsiveness to chemotherapy. This study sheds light on eIF6's potential therapeutic interventions to mitigate treatment resistance in SCLC.

overall_design:
To investigate function of eIF6 in H69 and H69M cells, we established stable eIF6-KD H69 and H69M cell lines by lenitivirus infection containing shRNA targeting eIF6.
Comparative gene expression profiling analysis of RNA-seq data for H69_sheIF6 cells vs. H69_shNTC cells and H69M_sheIF6 cells vs. H69M_shNTC cells

first: Zhile

first: Mengyao

first: Lunxu

last: Liu

first: Shensi

last: Shen


title: MSC-small extracellular vesicles alleviated Th2-airway inflammation by regulating the metabolism of DCs in mice

first: Long-Xin

last: Huang

address: No. 58, Zhongshan 2nd Road

institute: The First Affiliated Hospital of Sun Yat-sen University

summary:
Background: Allergic asthma is one of the chronic inflammatory diseases and is generally induced by CD4+ T helper 2 cells (Th2) in the context of persistent inhaled stimuli. Dendritic cells (DCs) are essential to mounting the Th2-mediated airway inflammation by presenting inhaled antigens to prime CD4+ T cells. Small extracellular vesicles (sEV) derived from mesenchymal stem cells (MSCs) exhibited great interest in intractable diseases. However, whether MSC-sEV play a role on DCs in airway inflammation is still unclear.
Methods: We isolate MSC-sEV using anion-exchange chromatography. Mouse bone marrow-derived DCs (BMDCs) and human monocyte-derived DCs (moDCs) were used to study the effects of MSC-sEV on dendritic cell surface molecules and their cytokine release. Mice were sensitized with house dust mites (HDM) to induce airway inflammation, and treated with MSC-sEV, The effects of sEV on murine DCs were identified. Extracellular flux analysis techniques were used to study the effects of MSC-sEV on the metabolic state of dendritic cells. RNA sequencing to study altered gene expression in BMDCs after MSC-sEV treatment.
Results: MSC-sEV mitigated the accumulation of Th2-associated moDCs in mouse lung in response to HDM. MSC-sEV also decreased the activation of moDCs induced in vitro including the expression of co-stimulatory molecules and cytokines secretion. Furthermore, we identified that DCs were able to take MSC-sEV in vitro and in vivo. Mechanistically, using bulk RNA-sequencing, we found that MSC-sEV played roles in the metabolic pathway of murine DCs. Using extracellular flux analysis, we found that MSC-sEV increased the requirement of oxidative phosphorylation on moDCs. Importantly, MSC-sEV displayed similar effects on human moDCs including decreased co-stimulatory molecular and cytokine production.
Conclusions: MSC-sEV are able to alter the metabolic state of DCs, favoring DCs to maintain OXPHOS (oxidative phosphorylation) rather than glycolysis, thereby reducing DCs-initiated inflammatory responses and attenuating Th2 lung inflammation, suggesting MSC-sEV can be a potential clinical therapy for airway inflammation.

overall_design: To identify how sEV regulated DC activation, the analysis of global genes was applied to understand the mechanism. We compared geomatic profiles of PBS or sEV-treated BMDCs.

first: Lifen

last: Wen


title: Gene Expression by a Model Fungus in the Ascomycota Provides Insight Into the Decay of Fungal Necromass

status: Public on May 01 2024

city: Saint Paul

first: Jonathan

last: Schilling

state: Minnesota

address: 1445 Gortner Avenue

department: Plant and Microbial Biology

institute: University of Minnesota

summary: Fungal necromass in soil represents the stable carbon pools. While fungi are known to decompose fungal necromass, how fungi decomopose melanin, remains poorly understood.  Recently, Trichoderma species was found to be one of the most commonly associated fungi in soil, we have used a relevant fungal species, Trichoderma reesei, to characterized  Genes involved in the decomposition of melanized and non-melanized necromass from Hyaloscypha bicolor.

overall_design: We grew Trichoderma reesei RUT-C30 in liquid cultures with low- and high-melanin necromass and  Determined gene expression patterns relative to each other. We also compared the expression pattern to that on glucose carbon source.

first: Irshad

middle: U

last: Haq

first: Peter

last: Kennedy


title: Longitudinal trajectories of mild/moderate SARS-CoV2 infection reveals key precursors and predictors of convalescent virus-specific immunity

status: Public on May 30 2024

city: Seattle

first: Allen Institute

last: For Immunology

state: Washington

address: 615 Westlake Ave N, Seattle, WA

institute: Allen Institute

summary: SARS-CoV2 infection results in clinically heterogeneous manifestations that are partially driven by complex longitudinal interactions with the immune system. The public health burden of mild COVID-19 is massive given the >100 million infected worldwide, but most research has focused on severe and fatal COVID-19. We recruited a mild COVID-19 cohort for multimodal immunophenotyping of single immune cells (scRNAseq, scATACseq, flow cytometry), serum proteins, virus-specific cellular and humoral immune responses, and clinical annotation from early acute infection to convalescence. Samples were acquired longitudinally from early acute infection before 15 days to >100 days post-symptom onset. Comparison to uninfected controls revealed marked immune activation consistent with acute response to viral infection at 15 days post-symptom onset with stronger inflammatory responses in older (>=40) compared to younger (<40) participants. Type I, II, and III interferon responses were most consistently upregulated in nearly all PBMC subsets. Plasmablasts were expanded in early infection with signatures of active IFNL and IFNG signaling, linking early inflammation and IFN responses to control of viral replication via antibodies.  Longitudinal models of scRNAseq confirmed most inflammatory pathways decreased over time, including type I and II IFN signaling, as well as signaling by innate danger sensors (TLR, RIG-I) that detect viral replication. By contrast, genes with increasing expression over time were enriched for EMT and wound healing pathways. Most participants showed consistent decay of inflammatory signatures by convalescence at >d 30 post-symptom onset except those presenting with PASC. Despite similar virus-specific adaptive immune responses between post-acute sequelae PASC and recovered convalescent participants, PASC participants showed persistent serum cytokine and chemokine differences including elevated IL5, TNF, IL-12p70, and soluble CD28 at >30 d post-symptom onset. scRNAseq, scATACseq, and network analyses provided more signs of persistent inflammatory activation in multiple cell types, with multimodal evidence pointing to increased activity of innate cells (CD14 monocytes, DCs) and effector T cells (CD4 TEM, CD8 TEMRA). Cytokines at day 7 PSO were highly correlated with and predictive of both virus-specific IgG and neutralizing antibody titers. Integrative network analyses defined the most enriched ligand-receptor pathways across PBMC subsets including LTA/TNF𝛃, TNF, and IFNg in early infection, and these also showed signs of persistent activation in PASC. Common downstream targets of these pathways in PASC point to novel therapeutic targets and mechanistic hypotheses, including IL-1Ꞵ, AP-1, ZFP36/TTP, and CEBP/ꞵ. These findings revealed dynamics of immune activation and critical early signals correlated with adaptive immune responses. Serum protein signatures predicting antibody responses and correlating with PASC may provide opportunities for enhanced immunomonitoring and personalized treatment.

overall_design:
We recruited a mild COVID-19 cohort for multimodal immunophenotyping of single immune cells (scRNAseq, scATACseq, flow cytometry), serum proteins, virus-specific cellular and humoral immune responses, and clinical annotation from early acute infection to convalescence. Samples were acquired longitudinally from early acute infection before 15 days to >100 days post-symptom onset.
NOTE from submitter: The FASTQs are from human subjects and they will be submitted to dbGAP.

first: Gregory

last: Szeto

first: Aarthi

last: Talla

first: Mark-Phillip

last: Pebworth

first: Suhas

last: Vasaikar

first: Kathy

last: Henderson

first: Alexander

middle: T

last: Heubeck

first: Mark

last: Gillespie

first: Palak

middle: C

last: Genge

first: Elliot

last: Swanson

first: Charles

last: Roll

first: Adam

last: Savage

first: Ernest

last: Coffey

first: Lynne

last: Becker

first: Julian

last: Reading

last: Skene

first: Xiaojun

last: Li

first: Troy

last: Torgeson

first: Thomas

middle: F

last: Bumol

first: M

last: McElrath

first: Kristen

last: Cohen

first: Evan

last: Newell

first: Maria

last: Lemos

first: Stephen

last: DeRosa

first: Julie

last: Czartoski

first: Lamar

last: Ballweber-Fleming


first: Dalin

last: Chen

summary: To investigate  skin aging is an important driver of experimental osteoarthritis(OA) progression in mice via enhanced IL-36 receptor (IL-36R) signaling. The supernatants from the co-culture of UV-aged mouse primary keratinocytes with synovial fibroblasts were collected to induce mouse primary articular chondrocytes (CCs) to become aging SNL CCs. On this basis, Spesolimab (100ng/ml) was added to culture aging SNL CCs to study the drug's effect on OA. We then performed gene expression profiling analysis using data obtained from RNA-seq of 3 groups: control CCs, aging SNL CCs, aging SNL CCs+Spesolimab.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for control CCs, aging SNL CCs and aging SNL CCs+Spesolimab.

first: Chong

last: Wang

first: Changsheng

last: Yang

first: Liangyu

first: Panpan

first: Zilin

last: Zou

first: Honghao

first: Yun

last: Xiao

first: Junfeng

last: Wu

first: Qinwei

last: Cheng

first: Chengliang

first: Ee

last: Ke

first: Bin

last: Huang

first: Xiaochun

last: Bai

first: Kai


title: Gene-expression profiling of individuals resilient to Alzheimer's disease reveals higher expression of genes related to metallothionein and mitochondrial processes and no changes in the unfolded protein response

status: Public on Apr 25 2024

city: Amsterdam

first: Luuk

middle: Elard

last: de Vries

address: Meibergdreef 47

department: Neuroregeneration

country: Netherlands

institute: Netherlands Institute for Neuroscience

summary: Some individuals show a discrepancy between cognition and the amount of neuropathological changes characteristic for Alzheimer’s disease (AD). This phenomenon has been referred to as ‘resilience’. The molecular and cellular underpinnings of resilience remain poorly understood. To obtain an unbiased understanding of the molecular changes underlying resilience, we investigated global changes in gene expression in the superior frontal gyrus of a cohort of cognitively and pathologically well-defined AD patients, resilient individuals and age-matched controls (n=11-12 per group). 897 genes were significantly altered between AD and control, 1121 between resilient and control and 6 between resilient and AD. Gene set enrichment analysis (GSEA) revealed that the expression of metallothionein (MT) and of genes related to mitochondrial processes was higher in the resilient donors. Weighted gene co-expression network analysis (WGCNA) identified gene modules related to the unfolded protein response, mitochondrial processes and synaptic signaling to be differentially associated with resilience or dementia. As changes in MT, mitochondria, heat shock proteins and the unfolded protein response (UPR) were the most pronounced changes in the GSEA and/or WGCNA, immunohistochemistry was used to further validate these processes. MT was significantly increased in astrocytes in resilient individuals. A higher proportion of the mitochondrial gene MT-CO1 was detected outside the cell body versus inside the cell body in the resilient compared to the control group and there were higher levels of heat shock protein 70 (HSP70) and X-box-binding protein 1 spliced (XBP1s), two proteins related to heat shock proteins and the UPR, in the AD donors. Finally, we show evidence for putative sex-specific alterations in resilience, including gene expression differences related to autophagy in females compared to males. Taken together, these results show possible mechanisms involving MTs, mitochondrial processes and the UPR by which individuals might maintain cognition despite the presence of AD pathology.

overall_design: RNA was isolated from the grey matter in the superior frontal gyrus.

middle: E

first: Aldo

last: Jongejan

first: Jennifer

last: Monteiro Fortes

first: Rawien

last: Balesar

first: Annemieke

last: Rozemuller

first: Perry

middle: D

last: Moerland

first: Inge

last: Huitinga

first: Dick

last: Swaab

first: Joost

last: Verhaagen


title: RNA-seq analysis of the gene expression profile of Huh7 cells upon incubation with different TGF-β-treated hepatic stellate cells (HSCs)-derived ectosomes

status: Public on Apr 28 2024

city: Xiamen

first: Qi-tao

last: Chen

state: Fujian Province

address: Xiang annan road

department: School of Life Sciences

institute: Xiamen University

summary:
Purpose: The goals of this study are to compare the gene expression profile of Huh7 cells treated with different ectosomes or not and to evaluate the function of ectosomal Hexokianse 1 (HK1) derived from HSCs.
Methods: Firstly, we treated different LX-2 HSCs with TGF-β (2 ng/mL) for 36 hours to activate HSCs, then ectosomes from LX-2 or HK1-KD LX-2 culture medium were isolated respectively by differential centrifugation processes. Secendly, Huh7 cells were preincubated with different ectosomes or not for 12 hours and then  fresh medium for another 12 hours. Lastly, different groups of Huh7 cells were prepared for RNA-seq.
Results: By Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis of the differentially expressed genes, a total of 124 pathways were found to be enriched in the ectosome-incubated Huh7 cells compared with the control Huh7 cells, while a total of 36 pathways were enriched in the ectosome-incubated Huh7 cells compared with the HK1-KD ectosome-incubated Huh7 cells.Gene set enrichment analysis (GSEA) also revealed enrichment of the N-glycosylation pathway in the ectosome-incubated Huh7 cells compared with the control Huh7 cells. Conversely, this pathway was negatively regulated in Huh7 cells incubated with HK1-KD HSC-derived ectosomes
Conclusion: the transferal of HSC-derived ectosomal HK1 into HCC cells regulates the N-glycosylation level of Huh7.

overall_design: Three groups of Huh7 were sequenced and every group had three independent samples.

first: Qiao-ling

first: Zhi-yuan

last: Zhang


title: Cranial Photomodulation Reshapes Brain Border Immunity and Promotes Recovery After Stroke

status: Public on Apr 18 2024

city: Zhanjiang

first: Wei

last: Feng

address: 236 Yuanzhu Road

institute: Central People's Hospital of Zhanjiang

summary: The skull bone marrow represents a targetable neuroimmune interface adjacent to the brain borders with the potential to improve stroke outcomes. Here, we identified the skull as a highly responsive immune niche and developed a cranial photo-immunologic regulation (CPR) strategy using low-dose, narrow-band medical UVB. Targeted irradiation of the interparietal region reshapes brain border immune crosstalk, effectively promoting repair and functional recovery after ischemic injury. Mechanistically, UVB-based CPR reverses stroke-induced suppression of meningeal B cell activation, contributing to restoring brain border immune homeostasis. Depletion of meningeal B cells further demonstrates that these cells are required for the therapeutic effects of CPR. These findings establish the skull as an actionable target for neuroimmune modulation along the skull–meninges–brain axis and provide a potentially transformative strategy for treating neurological disease.

overall_design: Single-cell transcriptomic profiling was performed on Sham, tMCAO (transient middle cerebral artery occlusion), and tMCAO+UVB(ultraviolet B) treated mouse skull, meninges, and brain tissue separately.

first: Chao

first: Chun-Jie

last: Liu

first: Meilian

last: Li

first: Jianhao

last: Zeng

first: Lisi

last: Wang

first: Xin

last: Dai

first: Tao

last: Tao


title: Cranial photoregulation reshapes brain border immunity and promotes recovery after stroke

overall_design: Bulk RNA sequencing of mouse skull, meninges, and brain tissue at 0.5/24 hours post cranial photobiomodulation, as well as bulk RNA sequencing of mouse skull, meninges, and brain tissue following non-cranial  photobiomodulation.


title: EIF6 conditions drug-tolerant persister-like transdifferentiation in small cell lung carcinoma

first: Haoning

last: Peng

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

overall_design: Refer to individual Series


title: EIF6 conditions drug-tolerant persister-like transdifferentiation in small cell lung carcinoma (SCLC cell lines)

overall_design:
We performed RNA-sequencing for paired cytoplasmic mRNA and polysomal mRNA in H196 cells vs. H69 cells and H69M cells vs.H69 cells for comparing their gene expression on both transcriptional and translational level.
We performed RNA-sequencing for paired cytoplasmic mRNA and polysomal mRNA in H196 cells vs. H69 cells, H69M cells vs.H69 cells, H82R cells vs. H82 cells and H209A cells vs.H209 cells for comparing their gene expression on both transcriptional and translational level.

first: Zhile

first: Mengyao

first: Lunxu

first: Shensi

last: Shen


title: The allosteric landscape of the Src kinase

status: Public on Apr 27 2024

city: Barcelona

first: Antoni

last: Beltran

address: Dr Aiguader, 88

institute: CRG Barcelona

summary: Enzymes catalyze the reactions of life and are the targets of nearly all small molecule drugs. Most drugs inhibit enzymes by binding to conserved active sites, causing problems of specificity and toxicity. Targeting regulatory allosteric sites can increase specificity, overcome drug resistance and tune or activate activity.However, the vast majority of enzymes have no known allosteric sites and methods do not exist to globally identify or predict them.Here we present a general and fast method to globally chart allosteric communication in enzymes and apply it to the Src protein kinase to produce the first comprehensive map of negative and positive allosteric control of enzymatic activity. Allosteric control of Src is pervasive, anisotropic and distance dependent, but fairly predictable from simple sequence and structural features. The comprehensive allosteric map enables unbiased identification of all the allosterically active surface pockets of the Src kinase for the development of activatory and inhibitory drugs.This general approach can be used to chart global allosteric maps of many kinases, enzymes, and other biochemical activities important for medicine and biotechnology.

sample_organism:
Saccharomyces cerevisiae
Homo sapiens

platform_organism:
Saccharomyces cerevisiae
Homo sapiens

overall_design: We designed a variant library covering the kinase domain of the human tyrosine kinase Src. The library is split in 5 blocks of ~60 aa residues, and each block consists of all single substitutions in 10 genetic backgrounds (54,455 genotypes in total across the full kinase domain). Each block was cloned into a Src kinase domain alone construct (KD), and a full-length Src construct (FL), and each of these was selected in triplicate in activity and protein abundance assays in yeast. The data files here correspond to the amplicon-seq libraries of the population of variants in each block before and after selection (inputs and outputs).

first: Ben

last: Lehner


title: Construction of Danio rerio Asymmetrical Maps (DreAM)

status: Public on Apr 08 2024

city: Hang Zhou

last: cheng

state: Zhe Jiang

address: 886 Yuhangtang Road

department: Institute of genetics

institute: Zhejiang University

summary: Purpose: Construction of 3D zebrafish spatial transcriptomics data for studying the establishment of AP axis. Methods:  We performed serial bulk RNA-seq data of zebrafish embryo at three development points. Using the published spatial transcriptomics data as references, we implemented Palette to infer spatial gene expression from bulk RNA-seq data and constructed 3D embryonic spatial transcriptomics. The constructed 3D transcriptomics data was then projected on zebrafish embryo images with 3D coordinates, establishing a spatial gene expression atlas named Danio rerio Asymmetrical Maps (DreAM). Results: DreAM provides a powerful platform for visualizing gene expression patterns on zebrafish morphology and investigating spatial cell-cell interactions. Conclusions: Our work used DreAM to explore the establishment of anteroposterior (AP) axis, and identified multiple morphogen gradients that played essential roles in determining cell AP positions. Finally, we difined a hox score, and comprehensively demonstrated the spatial collinearity of Hox genes at single-cell resolution during development.

overall_design: Bulk RNA-seq was performed on the serial cryosections taken along the left-right axis of zebrafish embryos at 10 hpf, 12 hpf and 16 hpf.

first: Yang

last: Dong

last: Cheng

first: Xiang


title: RpoS acts as a global repressor of virulence gene expression in Escherichia coli O104:H4 and enteroaggregative E. coli

status: Public on Apr 04 2024

city: Münster

first: Petya

last: Berger

address: Robert-Koch Straße 41

department: Institute of Hygiene

country: Germany

institute: University of Münster

summary: In 2011, in Germany, Escherichia coli O104:H4 caused the enterohemorrhagic E. coli (EHEC) outbreak with the highest incidence rate of hemolytic uremic syndrome. This pathogen carries an exceptionally potent combination of EHEC- and enteroaggregative E. coli (EAEC)-specific virulence factors. Here, we identified an E. coli O104:H4 isolate that carried a single nucleotide polymorphism (SNP) in the start codon (ATG>ATA) of rpoS, encoding the alternative sigma factor S. The rpoS ATG>ATA SNP was associated with enhanced EAEC-specific virulence gene expression. Deletion of rpoS in E. coli O104:H4 Dstx2 and typical EAEC resulted in a similar effect. Both rpoS ATG>ATA and DrpoS strains exhibited stronger virulence-related phenotypes in comparison to wild type. Using promoter-reporter gene fusions, we demonstrated that wild-type RpoS repressed aggR, encoding the main regulator of EAEC virulence. In summary, our work demonstrates that RpoS acts as a global repressor of E. coli O104:H4 virulence, primarily through an AggR-dependent mechanism.

overall_design: We performed strand specific RNA-seq with total RNA isolated from E. coli O104:H4 wild type and rpoS ATG>ATA grown under standard laboratory conditions (LB medium; 37oC; 180 rpm) to logarithmic growth phase (log; OD600 = 0.4- 0.5) and to transition to stationary phase (transition; OD600 = 3.0-3.4). Three biological replicates per strain were included. DESeq2 analysis based on the reads mapped to the E. coli O104:H4 genome was performed to identify examples of differential gene expression.


title: Transcriptome-wide mRNA condensation precedes stress granule formation and excludes stress-induced transcripts

status: Public on Apr 26 2024

first: D Allan

last: Drummond

address: 929 E 57th Street

institute: University of Chicago

summary: Stress-induced condensation of mRNA and proteins into stress granules is conserved across eukaryotes, yet the function, formation mechanisms, and relation to well-studied conserved transcriptional responses remain largely unresolved. Stress-induced exposure of ribosome-free mRNA following translational shutoff is thought to cause condensation by allowing new multivalent RNA-dependent interactions, with RNA length and associated interaction capacity driving increased condensation. Here we show that, in striking contrast, virtually all mRNA species condense in response to multiple unrelated stresses in budding yeast, length plays a minor role, and instead, stress-induced transcripts are preferentially excluded from condensates, enabling their selective translation. Using both endogenous genes and reporter constructs, we show that translation initiation blockade, rather than resulting ribosome-free RNA, causes condensation. These translation initiation-inhibited condensates (TIICs) are biochemically detectable even when stress granules, defined as microscopically visible foci, are absent or blocked. TIICs occur in unstressed yeast cells, and, during stress, grow before the appearance of visible stress granules. Stress-induced transcripts are excluded from TIICs primarily due to the timing of their expression, rather than their sequence features. Together, our results reveal a simple system by which cells redirect translational activity to newly synthesized transcripts during stress, with broad implications for cellular regulation in changing conditions.

overall_design: S. cerevisiae cells were stressed, and their RNA was fractionated either using Sed-Seq (to identify RNA which sediments in a non-ribosome dependent fashion) or Polysome-Seq (to identify the ribosome association of RNA).

first: Jared

last: Bard

first: D

first: Edward

middle: W

last: Wallace

first: Hendrik

last: Glauninger


title: Interaction between HIV-1 Tat and EBV Zta favours immune escape of B cells by downregulating HLA-ABC expression

status: Public on Apr 01 2024

city: Villejuif

first: Anna

last: Schwager

state: Please Select

address: 114 Rue Edouard Vaillant

country: France

institute: Institut Gustave Roussy

summary: Both Human immunodeficiency virus (HIV) and Epstein-Barr virus (EBV) are associated with an increased risk of malignancies. People living with HIV frequently have EBV reactivation and develop EBV-associated B-cell malignancies. In this study, we aimed to uncover the involvement of HIV-1 and EBV co-existence in the development of B-cell malignancies. We studied two viral transcriptional activators (HIV-1 Tat and EBV Zta) and their possible interaction since they both have cell-penetration domains and can be found simultaneously in the blood or cells of people with HIV. We found that Tat and Zta directly bound each other in human B cells, T cells, and blood serum. Using RNA-sequencing, we found that combined Tat and Zta action in B cells differed from a simple combination of two proteins. A subset of genes, activated by Tat or Zta alone, that trigger an immune response and antigen presentation in B cells, remained unchanged when two proteins were combined. B cells, treated or transfected with Tat and Zta, exhibited a substantial decrease in HLA-ABC (MHC class I) expression, a critical component of the antigen processing and presentation pathway. HLA-ABC downregulation induced by Tat and Zta interaction conferred protection against cytotoxic T cell recognition of EBV-infected B cells. Tat and Zta interaction was also observed in serum from an HIV-positive individual. To conclude, we demonstrated for the first time the direct interaction between HIV-1 Tat and EBV Zta; this interaction can bring about immune evasion of EBV-infected or transformed B cells.

overall_design: Purified B cells or BLAS cells were treated with Tat (250 ng/ml ~ 25 μM), Zta (1 µg/ml ~ 37 μM) or Tat and Zta (at 250 ng/ml and 1 µg/ml, respectively) or left untreated. 6 h and 48h later cells were collected and RNA was extracted using a NucleoSpin RNA isolation kit (Macherey-Nagel, Germany) following manufacturer’s instructions. Total RNA from BLAS cell lines was also isolated using the NucleoSpin RNA kit following the manufacturer's instructions. Each condition was performed and analyzed in triplicates. The cDNA libraries were constructed and sequenced in Novogene, China.


title: Whole transcriptome sequencing of 6 pairs of esophageal squamous cell carcinoma and paracancerous tissues

status: Public on Apr 15 2024

city: Nanchong

first: kang

last: liu

state: State...

address: 55 Dongshun Road, Gaoping District, Nanchong city, Sichuan Province

institute: north sichuan medical college

summary: Screening and identification of novel biomarkers affecting the progression of esophageal squamous cell carcinoma by whole-transcriptome sequencing of 6 pairs of esophageal squamous cell carcinoma tissues and adjacent tissues

overall_design: We sequenced and analyzed 6 pairs of esophageal squamous cell carcinoma tissues and paracancerous tissues

first: liu

last: kang

first: song

last: guiqin


title: Stiffness Assisted Cell-Matrix Remodeling Trigger Mechanotransduction Regulatory Programs to Induce Stem Cell Differentiation

city: Bryan

first: Irtisha

last: Singh

address: 8447 Riverside Pkwy, Medical Research and Education Building II Suite 4344

institute: Texas A&M University

summary: The interplay of transcription and epigenetics in response to biophysical factors is important in the design of engineered matrix platforms that seek to direct cellular fate. We present a nanoparticle crosslinker to reinforce hydrogels and study the impact on hMSC transcription through RNA-seq. The study provides insight into how environmental stiffness drives global intracellular signaling through PI3AKT transction and chromatin remodeling to enable YAP/TAZ activity.

overall_design: Examination of matrix stiffness impact on transcriptome and epigenome to modulate hMSC fate

middle: L

last: Kersey

first: Daniel

first: Kaivalya

last: Deo

first: Christina

last: Dubell

first: Ting-Ching

last: Wang

first: Manish

last: Jaiswal

first: Min Hee

last: Kim

first: Tanmay

last: Lele

first: Akhilesh

middle: K

last: Gaharwar

first: Aparna

last: Murali

first: Sarah

last: Hargett


title: Selective refueling of CAR T cells using ADA1 and CD26 boosts antitumor immunity

status: Public on Mar 27 2024

city: HOUSTON

first: Xiaotong

last: Song

state: Texas

address: 2121 W. Holcombe Blvd

department: CIID

summary: CAR T cell therapy has revolutionized the treatment of hematologic malignancies, but its efficacy in solid tumors remains limited due to the immunosuppressive nature of the tumor microenvironment and the inability of T cells to persist and traffic to the tumor site. While current strategies focus on enhancing CAR T cell activity through costimulatory molecules and cytokines, a critical yet often overlooked factor is the competition for nutrients between tumor cells and T cells in the nutrient-deprived tumor microenvironment. To address this challenge, we employed a selective metabolic refueling (MR) strategy by providing T cells with inosine as an alternative fuel source for growth and functionality. In this study, we engineered CAR T cells to co-express a membrane-bound CD26 and a cytoplasmic adenosine deaminase 1 (ADA1) fused to an anti-CD3 scFv. ADA1 irreversibly converts both intracellular and extracellular adenosine to inosine, overcoming adenosine-mediated immunosuppression and providing T cells with inosine for growth. The inclusion of an anti-CD3 scFv fusion partner and overexpressing CD26 boosts ADA1 capture in a membrane proximal manner, providing inosine for T cells and minimizing feeding the tumor cells. We demonstrate that ADA1 is conditionally secreted in an autocrine manner in response to CD3/CD26 stimulation and that it activates CAR T cells through trans-signaling in a tumor-specific manner. In addition, we show that, compared to unmodified CAR T cells, CD26-overexpressing CAR T cells have better migration capacity and are less susceptible to TGF-β1 suppression. Finally, we demonstrate that, in mice models of human hepatocellular carcinoma (GPC3-MR-CAR) and human non-small cell lung cancer (HER2-MR-CAR), metabolically refueled CAR T cells (MR-CAR) are more efficient in reducing tumor growth than unmodified CAR T cells. Thus, selective refueling CAR T cells using ADA1 and CD26 holds promise for improving the efficacy of CAR T cell therapy of solid tumors.

overall_design: Comprehensive analyses of RNA-seq profiles compared between nontransduced and different CAR-T cells

first: Abhijit

last: Sarkar


title: TDG orchestrates ATF4-dependent gene transcription during retinoic acid-induced cell lineage committment

status: Public on Mar 31 2024

city: Nantes

first: Christine

last: Chauveau

address: 30 Boulevard Jean Monnet

institute: CR2TI UMR 1064

summary: Cell differentiation during development is associated to large-scale modifications in the methylome that require the engagement of an active DNA demethylation machinery including Ten-Eleven-Translocation enzymes for oxidation of 5-methylcytosine and the T:G mismatch DNA glycosylase (TDG) for removal of the oxidized bases. Despite this well-defined molecular function, the biological output of TDG activity remains elusive. Here we combined transcriptomic and epigenomic approaches in TDG knock-out embryonal carcinoma cells, an epiblast stem-like cell model, to decipher TDG function in pluripotent cells and their retinoic acid-induced differentiated progeny. We determined that TDG activity is balancing differentiation in favor of a neural fate at the expense of a cardiac mesoderm fate. This process is associated with a sustained activity of a large set of ATF4-dependent genes in relation with TDG engagement at the Atf4 gene promoter and in conjunction with a TDG-dependent regulation of the mammalian target of rapamycin complex 1. These observations highlight the central role of TDG in cell differentiation and support a model linking metabolic reprogramming to cell fate acquisition.

overall_design: To investigate the role of TDG during cell differentiation, we invalidated the Tdg gene by CRISPR-Cas9 in P19 embryonal carcinoma cells. wt and Tdg-null cells were then grown as monolayers or aggregates in the presence or not of retinoic acid. RNA-seq was then applied to the different samples in order to establish the transcriptome associated with these different cell culture conditions.

first: Jeremie

last: Poschmann

first: Gilles

last: Salbert

first: Martin

last: Braud

first: Cynthia

last: Fourgeux

first: Thierry

last: Madigou

first: Marion

last: Turpin


title: Analysis of retroviral MLT1 enhancer-driven genes identifies over-expression of EPS8L1 as an early-stage initiator of both oxidative stress and poor trophoblast invasiveness in the pregnancy disorder preeclampsia

city: Berlin

first: Amit

last: Pande

address: Robert-Rössle-Straße 10

institute: Max Delbruck Center for Medicine

summary: In the human-specific disorder of pregnancy preeclampsia (PE) we find dysregulation of multiple genes associated with the cis-regulatory elements (CREs) derived from mammalian-specific endogenous retrovirus (ERV3-MLT1)

overall_design: To identify the regulatory effect of the EPS8L1 enhancer derived from mammalian-specific endogenous retrovirus (ERV3-MLT1).

first: Laurence

last: Hurst

first: Zsuzsanna

last: Izsvak

first: Rabia

last: Anwar


title: DNA methylation status determines the sensitivity of repeats to restriction by the HUSH-MORC2 complex [Bulk RNA-seq]

status: Public on Mar 26 2024

city: Lund

first: Ninoslav

last: Pandiloski

address: Sölvegatan 17

country: Sweden

institute: Lund University

summary: The human silencing hub (HUSH) complex binds to transcripts of LINE-1 retrotransposons (L1s) and other genomic repeats, recruiting MORC2 and other effectors to remodel chromatin. However, how HUSH and MORC2 operate alongside DNA methylation, a central epigenetic regulator of repeat transcription, remains poorly understood. Here we interrogate this relationship in human neural progenitor cells (hNPCs), a somatic model of brain development that tolerates removal of DNA methyltransferase DNMT1. Upon loss of MORC2 or HUSH subunit TASOR in hNPCs, L1s remain silenced by robust promoter methylation. However, genome demethylation and activation of evolutionarily-young L1s attracts MORC2 binding. Simultaneous depletion of DNMT1 and MORC2 causes massive accumulation of L1 transcripts. We identify the same mechanistic hierarchy at pericentromeric α-satellites and clustered protocadherin genes, repetitive elements important for chromosome structure and neurodevelopment respectively. Our data delineate the independent epigenetic control of repeats in somatic cells, with implications for understanding the vital functions of HUSH-MORC2 in hypomethylated contexts throughout human development.

overall_design: Expression levels of genes and transposable elements in human neuroepithelial-like stem cells (hNSC) was examined by Bulk-RNA sequencing in five different conditions: LacZ - CRISPRi (a control targetting a bacterial gene which is not present in the human genome), TASOR - CRISPRi, MORC2 - CRISPRi, DNMT1 - CRISPRi and DNMT1&MORC2 - CRISPRi.


status: Public on Mar 19 2024

city: Rennes

address: 263 Av. Gal. Leclerc

department: IGDR UMR6290

institute: University Rennes 1

overall_design: To investigate the role of TDG during cell differentiation, we invalidated the Tdg gene by CRISPR-Cas9 in P19 embryonal carcinoma cells. wt and Tdg-null cells were then grown as monolayers in the presence or not of 1 micromolar retinoic acid. MNase-seq was then applied to the different samples in order to establish genome-wide maps of nucleosome occupancy in these different conditions..

first: Stéphane

last: Avner


title: DNA methylation status determines the sensitivity of repeats to restriction by the HUSH-MORC2 complex [ONT cDNA sequencing]

overall_design: Long transcripts of transposable elements, long, and nested genes were examined with ONT long-read cDNA sequencing in four different conditions: LacZ - CRISPRi (a control targetting a bacterial gene which is not present in the human genome), MORC2 - CRISPRi, TASOR-CRISPRi, and DNMT1-CRISPRcut.


title: DNA methylation status determines the sensitivity of repeats to restriction by the HUSH-MORC2 complex

overall_design: Refer to individual Series (listed below).


title: The long non-coding RNA Meg3 mediates imprinted gene expression  during stem cell differentiation

status: Public on Mar 28 2024

city: Gif-sur-Yvette

first: Benoit

last: Moindrot

address: 1, avenue de la terrasse

institute: Institute for Integrative Biology of the Cell

summary: The imprinted Dlk1-Dio3 domain comprises the developmental genes Dlk1 and Rtl1, which are silenced on the maternal chromosome in different cell types. On this parental chromosome, the domain’s imprinting control region activates a polycistron that produces the lncRNA Meg3 and many miRNAs (Mirg) and C/D-box snoRNAs (Rian). Although Meg3 lncRNA is nuclear and associates with the maternal chromosome, it is unknown whether it controls gene repression in cis. We created mouse embryonic stem cells (mESCs) that carry an ectopic poly(A) signal, reducing RNA levels along the polycistron, and generated Rian-/- mESCs as well. Upon ESC differentiation, we found that Meg3 lncRNA (but not Rian) is required for Dlk1 repression on the maternal chromosome. Biallelic Meg3 expression acquired through CRISPR-mediated demethylation of the paternal Meg3 promoter led to biallelic Dlk1 repression, and to loss of Rtl1 expression.  lncRNA expression also correlated with DNA hypomethylation and CTCF binding at the 5’-side of Meg3.  Using Capture Hi-C, we found that this creates a Topologically Associating Domain (TAD) organization that brings Meg3 close to Dlk1 on the maternal chromosome.  The requirement of Meg3 for gene repression and TAD structure may explain how aberrant MEG3 expression at the human DLK1-DIO3 locus associates with imprinting disorders.

overall_design: Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for CTCF in F1-hybrid mouse embryonic stem cells (BJ1 cells = C57BL/6J x JF1; JB1 cells = JF1 x C57BL/6J). Cells are described in Kota et al. (2014). Input is also provided

first: Daan

last: Noordermeer


title: Embryonic lumenogenesis is controlled by selective mRNA decay triggered by LIN28A relocation [miCLIP]

status: Public on Mar 01 2024

city: London

first: Jernej

last: Ule

address: 10-12 Russell sq

department: Neuromuscular Diseases

country: United Kingdom

institute: UCL Institute of Neurology

summary: The naïve pluripotent epiblast cells become polarized into a rosette-like structure, followed by irreversible transition into primed pluripotency during one of the fastest morphological switches termed lumenogenesis. This requires rapid decay of pluripotency-associated mRNAs, but the underlying mechanism remains unknown. Guided by machine learning and metabolic RNA sequencing, we identified RNA binding proteins (RBPs), especially LIN28A, as primary mRNA decay factors. To understand if RBP dynamics steer embryogenesis, we used mRNA-RBP interactome capture during naïve-rosette-epiblast-gastrulation progression. We identified a dramatic increase in LIN28A mRNA binding, driven by its nucleolus-to-cytoplasm translocation during the naïve-primed pluripotency transition. Cytoplasmic LIN28A binds to 3’UTRs of pluripotency-associated mRNAs to directly stimulate their decay, and thereby progression to lumenogenesis. Accordingly, forced nuclear retention of LIN28A impeded lumenogenesis, causing an unforeseen embryonic multiplication and impaired gastrulation. This reveals selective mRNA decay, driven by nucleo-cytoplasmic RBP translocation, as an intrinsic mechanism for cell identity switch that controls embryonic timing of lumenogenesis.

overall_design: Examination of iCLIP, RNA-seq, QuantSeq in the differentiation and priming of mouse embryonic stem cells and in the presence or absence of LIN28A

first: Miha

last: Modic

first: Igor

last: Ruiz de los Mozos


title: Transcriptomic analyses of a Cyberlindnera jadinii strain (NBRC0988) grown in glycerol

status: Public on Mar 14 2024

city: Tianjin

first: Feiyu

last: Fan

state: Tianjin

address: 32 West 7th Avenue, Tianjin Airport Economic Area

institute: Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences

summary: A strand-specific RNA-seq library was constructed for a single sample, specifically the wild-type strain NBRC0988 grown in Yeast Extract Peptone (YP) medium supplemented with 2% w/v glycerol. To prepare the RNA sample, NBRC0988 cells grown overnight were inoculated into 100 mL of liquid Yeast Extract Peptone Dextrose (YPD) medium, with an initial inoculation OD600 of 0.1. These cells were then cultured for 15 hours at 30°C and 250 rpm. Subsequently, the cells were collected by centrifugation at 6,000g for 5 minutes. After being washed twice with phosphate buffer saline (PBS), they were inoculated into fresh 100 mL of YP medium containing 2% w/v glycerol. Following flask culturing at 30°C and 250 rpm for an additional 5 hours, the yeast cells were collected by centrifugation for total RNA isolation and Illumina RNA-seq library construction. Total RNA from the sample was isolated using the TRIzol reagent (Invitrogen, Grand Island, USA) and then utilized for high-throughput RNA sequencing. Commercially, the 150-nt paired-end strand-specific RNA-seq libraries (SS_lib_type RF) were generated by Novogene Biotechnology Co. Ltd (Tianjin, China) using the Illumina's Novaseq 6000 platform (Illumina, San Diego, USA).

overall_design: RNA profiles of C. jadinii wild-type strain NBRC0988 grown in YP medium containing 2% w/v glycerol was generated by deep sequencing, using Illumina’s novaseq 6000 platform

first: Liping

last: Xu

first: Rongxin

last: Zhang

first: Qilong

last: Wen

first: Xuqian

first: Lishan

last: Gu

first: Kaiyu

last: Ma

first: Gen

last: Li

first: Honglei

last: Wang

first: Xueli


title: Temporal gene expression changes and affected pathways in neurodevelopment of a mouse model of Smith-Lemli-Opitz syndrome

status: Public on Feb 15 2024

city: Seattle

first: Libin

state: WA

address: 1705 NE Pacific St

department: Medicinal Chemistry

institute: University of Washington

summary: Smith-Lemli-Opitz syndrome is an autosomal recessive disorder that arises from mutations in the gene DHCR7, which encodes the terminal enzyme of cholesterol biosynthesis, leading to decreased production of cholesterol and accumulation of the cholesterol precursor, 7-dehydrocholesterol, and its oxysterol metabolites. The disorder displays a wide range of neurodevelopmental defects, intellectual disability and behavioral problems. However, an in-depth study on the temporal changes of gene expression in the developing brains of SLOS mice has not been done before. In this work, we carried out the transcriptomic analysis of whole brains from WT and Dhcr7-KO mice at four time points through postnatal day 0. First, we observed the expected downregulation of the Dhcr7 gene in the Dhcr7-KO mouse model, as well as gene expression changes of several other genes involved in cholesterol biosynthesis throughout all time points. Pathway and GO term enrichment analyses revealed affected signaling pathways and biological processes that were shared amongst time points and unique to individual time points. Specifically, the pathways important for embryonic development, including Hippo, Wnt, and TGF-β signaling pathways are the most significantly affected at the earliest time point, E12.5. Additionally, neurogenesis-related GO terms were enriched in earlier time points, consistent with the timing of development. Conversely, pathways related to synaptogenesis, which occurs later in development compared to neurogenesis, are significantly affected at the later time points, E16.5 and PND0, including the cholinergic, glutamatergic, and GABAergic synapses. The impact of these transcriptomic changes and enriched pathways is discussed in the context of known biological phenotypes of SLOS.

overall_design: Total RNA was sequenced for four replicates each of whole brains from WT and Dhcr7-KO mouse embryos/neonates at four different gestational time points, including three embryonic (E12.5, E14.5, and E16.5) and one postnatal time point (PND0).

first: Li

last: Amy

first: Tomita

last: Hideaki

first: Xu

last: Libin


title: Effect of hypoxia on canine glioma stem-like cells metabolism

city: Davis

first: Ryan

last: Toedebusch

state: CA

address: 1 Shields Avenue

institute: University of California

type: Methylation profiling by high throughput sequencing

summary:
To investigate the role of hypoxia treatment on the metabolic profiles of canine glioma cells
We then performed reduce representation bisulfite sequencing on 3 unique canine cell lines and correlated with functional assays

overall_design: Following hypoxia treatment, migration, viability, gene expression, and RRBS profiling was carried out to understand the role of metabolism-related changes in canine glioma cells

first: Christine


title: shRNA-based miRNA overexpression results in significant off-target effects caused by isomiR production [miRNA-seq PC-3]

status: Public on Apr 17 2024

city: Moscow

first: Anton

last: Zhiyanov

address: 20 Myasnitskaya ulitsa

department: Faculty of Biology and Biotechnology

country: Russia

institute: HSE University

type: Non-coding RNA profiling by high throughput sequencing

summary: miRNA overexpression strategy based on RNA Polymerase (Pol) III mediated expression of short hairpin RNA (shRNA) coding particular miRNA is widely used for miRNA functional studies. For some miRNAs, e.g., encoded in the genome as a part of a polycistronic miRNA cluster, it is most likely the only way for their individual stable overexpression. Here we have revealed that extra \texttt{U} residues (up to five) added Pol III at the 3$'$-end of the transcribed shRNA associated with a shift in the Dicer cleavage position of the shRNA. This results in the undesirable formation of 5$'$-end miRNA isoforms (5$'$-isomiRs), which have a significantly altered seed region compared to the initially encoded canonical hsa-miR-93-5p. The predominant expression of 5$'$-isomiRs without three or four first nucleotides instead of the canonical isoform could be disclosed based on miRNA-Seq analysis. Moreover, mRNA sequencing data showed that the 5$'$-isomiRs of hsa-miR-93-5p presumably regulate their own mRNA targets. Thus, omitting miRNA-Seq analysis may lead to erroneous conclusions regarding revealed mRNA targets and possible molecular mechanisms in which studied miRNA is involved.

overall_design:
PC-3 (prostate cancer) cell line was transduced by lentiviral particles coding one of the two types of shRNA to achieve stable overexpression of hsa-miR-93-5p.
All shRNA constructs contained the sequence of the canonical hsa-miR-93-5p and differed in their stem lengths. The first one contained the full sequence of hsa-miR-93-5p: 5'-CUACCUGCACGAACAGCACUUAG-3' -- upper (5p) stem, 5'-CUUCAAGAGAG-3' -- stem loop, 3'-...UUGAUGGACGUGCUUGUCGUGAAAC-5' -- lower (3p) stem, while the second one contained 4 nucloetides 3'-end shortened version: 5'-CUGCACGAACAGCACUUAG-3' -- upper (5p) stem, 5'-CUUCAAGAGAG-3' -- stem loop, 3'-...UUGACGUGCUUGUCGUGAAAC-5' -- lower (3p) stem.
Both shRNA-overexpressed cell lines were sequenced to understand the impact of shRNA length on isomiR production.

first: Diana

last: Maltseva

first: Ivan

last: Kirillov

first: Roman

last: Suvorov

first: Daria

last: Gubani

first: Anna

last: Kudriaeva

first: Alexey

last: Belogurov

first: Alexander

last: Tonevitsky


title: Embryonic lumenogenesis is controlled by selective mRNA decay triggered by LIN28A relocation

first: Jernej

last: Ule

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

overall_design: Refer to individual Series


title: DNA methylation status determines the sensitivity of repeats to restriction by the HUSH-MORC2 complex [CUT&RUN]

first: Ninoslav

last: Pandiloski

overall_design:
Comparative H3K9me3 and H3K4me3 enrichment analysis of CUT&RUN-seq data for human neuroepithelial like stem cells (hNSCs). The comparison was done between hNSCs lacking MORC2, TASOR, DNMT1 and their respective controls to investigate the changes induced by the removal of the forementioned epigenetic repressors. Additionally, CUT&RUNseq was also employed to explore the occupancy of MORC2.

Comparative H3K4me3 enrichment analysis of CUT&RUN-seq data for human neuroepithelial like stem cells (hNSCs). The comparison was done between hNSCs lacking DNMT1 and DNMT1+MORC2 and their respective controls to investigate the changes induced by the removal of the forementioned epigenetic repressors.


title: Next generation sequencing to identify GPT1 regulating signaling pathway in CRC

status: Public on Feb 07 2024

city: Guangzhou

last: Xiong

state: Guangdong

address: 58 Zhongshan Road 2

department: Department of Endocrinology

institute: The First Affiliated Hospital of Sun Yat-sen University

summary: We performed next generation sequencing between control CRC cells and CRC cells with GPT1 overexpression to identify GPT1 regulating signaling pathway

first: Zhenxing

last: Liang

first: Huashan

last: Liu


title: Effect of IFN-α and IFN-γ on gene expression of 3D organoids derived from telomerase-immortalized normal human esophageal keratinocyte cell line, EPC2-hTERT.

status: Public on Feb 26 2024

city: Philadelphia

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, allergic inflammatory disease of the esophageal mucosa. While type 2 cytokines are predominant, targeting type 2 inflammation alone is not always effective for all patients. In EoE patients, the transcriptional profiling of  esophageal biopsy reveal upregulation of type I and II interferon (IFN) response, dysregulated immune signaling and downregulation of esophageal-specific genes in EoE patients. However, the role of interferon signaling during EoE is not fully understood. This study aims to investigate the impact of IFN signaling on esophageal epithelium and elucidate its role in EoE. IFN-γ stimulation of esophageal epithelial cells triggered disruption of tissue differentiation, epithelial barrier integrity and cytotoxicity, which are relevant features of EoE immunopathology. Beyond EoE, our findings offer insights into esophageal disorders which may involve increased IFN-γ signaling, including infections, gastrophageal reflux disease, and Barrett's esophagus.

overall_design: Comparative transcriptomic profiling of 3D organoids derived from telomerase-immortalized normal human esophageal keratinocyte cell line, EPC2-hTERT

first: Melanie

last: Ruffner


title: Multiplexed Assays of Variant Effects (MAVE) of PALB2 single nucleotide variants in exon 10.

status: Public on Feb 08 2024

city: Rochester

first: Fergus

last: Couch

address: 200 1st ST SW

institute: Mayo Clinic

summary: To validate a high-throughput screening data in human cells using Multiplexed Assays for Variant Effects (MAVE), we performed a high-throughput deep mutational scanning of single nucleotide changes in exon 10 encoding p.G1000 to p.I1037 of the WD40 domain of PALB2 using a cell survival assay in haploid human HAP1 cells. We obtained MAVE scores for 276 single-nucleotide variants, leading to 9 nonsense and 68 synonymous changes, as well as 199 amino acid substitutions. Both variant groups showed an asymmetric distribution that is skewed towards low MAVE scores of nonsense and damaging variants, respectively. These MAVE data included scores for 218 unique single-nucleotide variants, leading to 9 nonsense changes and 209 amino acid substitutions. We observed a good and significant correlation between the outcomes from the MAVE and high-throughput screens (n=179, r=-0,6439, p<0.0001), indicating concordance between the outcomes of high-throughput analysis of PALB2 variants in human and mouse cells.

overall_design: Saturation genome editing was used to introduce variants and the variant count on day 5 and day 20 were measured. The log2 ratio between D5 and D20 was used to measure the variant effects

first: Huaizhi

last: Huang

first: Chunling

last: Hu

middle: J


title: DNA methylation-based classification of common renal tumors

city: Bethesda

first: Kenneth

last: Aldape

state: Maryland

address: Building 10, Room 2S235

department: NCI, LABORATORY OF PATHOLOGY (HNC7Z18)

institute: National Institute of Health

summary: In present work one of our main objectives was to collect enough DNA methylation profiles of diverse tumor types in renal cancer.  So, we used a collection of newly processed samples on DNA methylation data to have a variety of diverse kidney tumors from various sources. We processed a subset of samples (n=156) profiled as a part of our routine clinical methylation testing for kidney cancer at NIH. We collected one subset of samples (n =173) from our collaborators at UCSF and another subset of samples (n=82) from NYU.

overall_design: All samples were profiled in accordance with standard manufacturer’s protocol for Illumina EPIC array (http://www.illumina.com/).

first: Antonios

last: Papanicolau-Sengos

first: Omkar

last: Singh

first: Kyung

last: Park

first: Matija

last: Snuderl

first: David

last: Solomon


title: Glomerular voom-transformed RNAseq counts from the Neptune Cohort and healthy living donors [re-analysis of GSE197307]

city: Ann Arbor

first: Sean

last: Eddy

state: MI

address: 1150 West Medical Center Drive

institute: University of Michigan

summary: Glomerular Transcriptome in the Nephrotic Syndrome study network cohort

overall_design:
Please note that GSE197307 samples were re-analyzed as described below.
Glomerular RNAseq profiles; Raw data (fastq) files are available upon approved ancillary study request from the NEPTUNE consortium (please see https://www.neptune-study.org/ancillary-studies; email contact: NEPTUNE-STUDY@umich.edu
*data processing step:
fastqScreen 0.11.4
cutadapt 1.14 -m 50 -A CTGTCTCTTATA -a CTGTCTCTTATA
STAR 2.5.2.a --bamRemoveDuplicatesType UniqueIdentical --twopassMode Basic --outFilterIntronMotifs RemoveNoncanonical --outSAMtype BAM SortedByCoordinate --outSAMstrandField intronMotif
htseq-count --stranded=no --order=pos --mode=union
genome build/assembly: GRCh 38.86
processed data files format and content: voom transformed data were batch corrected and trimmed to remove transcripts of no detectable abundance.

first: Eichinger

last: Felix

first: Eddy

last: Sean

first: Mariani

last: Laura

first: Menon

last: Rajasree

first: Godfrey

last: Brad

first: Kretzler

last: Matthias


title: DNA methylation status determines the sensitivity of repeats to restriction by the HUSH-MORC2 complex [ONT whole genome DNA]

first: Ninoslav

last: Pandiloski

overall_design:
DNA methylation levels of genes and transposable elements in human neuroepithelial-like stem cells (hNSC) were examined by ONT long-read DNA sequencing in two different conditions: LacZ - CRISPRi (a control targetting a bacterial gene which is not present in the human genome) and MORC2 - CRISPRi.

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


title: Antisense transcription can induce expression memory via stable promoter repression

status: Public on Mar 11 2024

first: Till

last: Schwämmle

state: Berlin

address: Ihnestrasse 63

department: Otto-Warburg-Laboratory

institute: Max Planck Institute for Molecular Genetics

summary: The capacity of cells to retain a memory of previous signals enables them to adopt unique cell fates and adjust to their surrounding environment. The underlying gene expression memory can arise from mutual repression of two genes, forming a toggle switch. Such mutual repression may occur at antisense loci, where two convergently oriented genes repress each other in cis. Under which conditions antisense transcription can generate expression memory remains poorly understood. To address this question, we combine mathematical modeling, genomics and a synthetic biology approach. Through simulations we show that stable memory can emerge, if both genes in an antisense pair transcribe through the convergent promoter and induce a stable repressive chromatin state. Genome-wide analysis of nascent transcription further supports antisense-mediated promoter repression with promoter-overlapping antisense gene pairs exhibiting mutually exclusive expression. Through constructing a synthetic antisense locus in mouse embryonic stem cells (mESCs) we then show that such a locus architecture can indeed maintain a memory of a transient stimulus. Mutual repression and the capacity for memory formation are elevated, when mESCs differentiate, showing that epigenetic memory is a cell type-specific property. Our finding that stem cells adapt their ability to remember stimuli as they differentiate might help to elucidate how stemness is maintained.

overall_design: We performed Whole-genome bisulfite-sequencing (WGBS) in differentiating female mouse embryonic stem cells (TX1072dXIC XX cell line). Samples were collected at days 0, 2 and 4 of 2i/LIF-withdrawal.

first: Verena

last: Mutzel

first: Svearike

last: Oeverdieck

first: Lucia

last: Librenjak

first: Benedikt

last: Boesen

first: Melissa

last: Bothe

first: Rutger

last: Gjaltema

first: Ilona

last: Dunkel

first: Gemma

last: Noviello

first: Edda

middle: G

last: Schulz


title: Biallelic non-productive enhancer-promoter interaction precedes imprinted expression of KcnK9 during mouse neural commitment [RNA-seq]

status: Public on Feb 05 2024

city: Clermont-Ferrand

first: franck

last: court

address: 28 Place Henri Dunant

institute: GReD CNRS

summary: How constitutive allelic methylation at imprinting control regions (ICRs) interacts with other levels of regulation to drive timely parental allele-specific expression along large imprinted domains remains partially understood at most imprinted loci. To gain insight into the regulation of the Peg13-KcnK9 domain, an imprinted domain with important brain functions, during neural commitment, we performed a detailed integrative analysis of the epigenetic, transcriptomic and cis-spatial organisation in an allele-specific manner in a mouse stem cell-based model of corticogenesis that recapitulates the control of imprinted gene expression in the embryonic brain. We have evidence that despite an allelic higher order chromatin structure associated with the paternally CTCF-bound Peg13 ICR, enhancer-Kcnk9 promoter contacts can occur on both alleles, although they are only productive on the maternal allele. This observation challenges the canonical model in which CTCF binding isolates the enhancer and its target gene on either side, and suggests a more nuanced role for allelic CTCF binding at the ICR of this locus.

overall_design: RNA-seq  generated on ES and NP cells for B6 x JF1 and JF1 x B6 genetic backgrounds.

first: Jil

last: Cercy

first: Franck

last: Court

first: Philippe

last: Arnaud


title: 5-Hydroxymethylcytosine Profiling of Cell-Free DNA Identifies Bivalent Genes That Are Prognostic of Survival in High-Risk Neuroblastoma [RNA-Seq]

status: Public on Feb 01 2024

city: Chicago

first: Mark

last: Applebaum

state: IL

address: 900 E 57th St

department: Pediatrics, Section of Hematology/Oncology

institute: University of Chicago

summary:
Purpose: Neuroblastoma is the most common extracranial solid tumor in childhood. We previously showed that circulating cell-free DNA (cfDNA) and tumor biopsy derived 5-hydroxymethylcytosime (5-hmC) profiles identified patients with neuroblastoma who experienced subsequent relapse. Here, we hypothesized that 5-hmC modifications selectively enriched in cfDNA compared with tumor biopsy samples would identify epigenetic changes associated with aggressive tumor behavior and identify novel biomarkers of outcome in patients with high- risk neuroblastoma.

Methods: 5-hmC profiles from cfDNA (n = 64) and tumor biopsies (n = 48) were compared. Two neuroblastoma cell lines underwent chromatin immunopre- cipitation followed by sequencing (ChIP-Seq) for H3K27me3, H3K4me3, and H3K27ac; kethoxal-associated single-stranded DNA sequencing; hmC-Seal for 5-hmC; and RNA-sequencing (RNA-Seq). Genes enriched for both H3K27me3 and H3K4me3 in the included cell lines were defined as bivalent. Using bivalent genes defined in vitro, a bivalent signature was established in three publicly available cohorts of patients with neuroblastoma through gene set variation analysis. Differences between tumors with high or low bivalent signatures were assessed by the Kaplan-Meier method and Cox proportional hazards models.

Results: In cfDNA compared with tumor biopsy derived 5-hmC profiles, we found in- creased 5-hmC deposition on Polycomb Repressive Complex 2 target genes, a finding previously described in the context of bivalent genes. We identified 313 genes that bore bivalent chromatin marks, were enriched for mediators of neuronal differentiation, and were transcriptionally repressed across a panel of heterogeneous neuroblastoma cell lines. In three distinct clinical cohorts, low bivalent signature was significantly and independently associated with worse clinical outcome in patients with high-risk neuroblastoma.

Conclusion: Low expression of bivalent genes is a biomarker of worse outcome in patients with high-risk neuroblastoma.

overall_design: RNA-Seq performed in 8 neuroblastoma cell lines without any treatment. RNA-Seq performed in 2 neuroblastoma cell lines with and without tazemetostat treatment.

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