title: Sequencing of the effect of mechanical stimulation on mesenchymal stem cells

status: Public on Nov 13 2023

city: Beijing

first: Yikun

last: Zhou

state: China

address: 22# South Zhongguancun Ave

institute: Peking University

summary: Mechanical force is a crucial external stimulus that plays a significant role in regulating bone structure and remodeling. Excessive loading of the bone and joint can lead to increased catabolism, chondrocyte necrosis, apoptosis and damage to the collagen network of bone(3–5). Osteoarthritis (OA), a degenerative osteoarticular disease, is associated with abnormal mechanical force stimulation, which can occur in various joints such as knee, temporomandibular joint [TMJ], shoulder and hip(6). Conversely, the absence of mechanical loading, such as prolonged bed rest or exposure to a microgravity environment in space, can result in a rapid decrease in bone mass and strength. Understanding how mechanical stimuli regulate bone homeostasis is crucial for exploring therapeutic strategies for bone metabolic diseases.Mesechymal stem cells (MSCs) act as the external force sensoring and compression-bearing elements.What we want to explore is how mechanical stimulation affects the genome changes of mesenchymal stem cells.

overall_design: In order to explore the influence of mechanical stimulation on mesenchymal stem cells, we designed two groups: the control group and the mechanical stimulation group. The mechanical stimulation group was stimulated by large mechanical force for 24 hours, and the expression differences were compared.


title: Bulk ATAC-seq of mouse neural stem and progenitor cells (NSPC) derived from embryonic stem cells, with biallelic Sox2 enhancer cluster deletions

city: Toronto

first: Jennifer

last: Mitchell

state: Ontario

address: 25 Harbord Street

department: Cell and Systems Biology

country: Canada

institute: University of Toronto

type: Genome binding/occupancy profiling by high throughput sequencing

summary: We have generated ATAC-sequencing datasets of the regulome of mouse neural stem and progenitor cells derived from embryonic stem cells, with allele-specific deletions of Sox2 enhancer cluster regions. ATAC-seq experiments were conducted to evaluate the alterations in chromatin accessibility at candidate regulatory elements genome-wide in neural stem and progenitor cells with Sox2 enhancer loss-of-function.

overall_design: Bulk ATAC-seq of mouse neural stem and progenitor cells (NSPC) derived from embryonic stem cells, with Sox2 enhancer deletions

first: Ian

last: Tobias

middle: A


title: Bivalent histone modification of BCG1 enables fungal pathogen to evade host immunity (ChIP-Seq)

status: Public on Dec 04 2023

city: Hefei

first: zhao

middle: xiao

last: zhen

address: Anhui Agricultural University, 130 Changjiangxilu, Shushan District, Hefei, Anhui Province, China

country: China

institute: Anhui Agricultural University

summary: Bivalent chromatin modification containing opposing H3K4me3 and H3K27me3 marks controls various biological processes by fine-tuning gene expression in animals and plants, however how this bivalent modification regulates pathogenicity of fungal pathogen remains exclusive. Here, we provided a genome-wide landscape of H3K4me3 and H3K27me3 of wheat head blight fungus Fusarium graminearum (Fg), leading to the identification of infection-induced bivalent chromatin-marked genes (BCGs). Among those, BCG1, which encodes a novel xylanase with a G/Q rich motif, is required for the full virulence of Fg pathogenicity through degradation of host cell wall. However, the G/Q rich motif is recognized by pattern-recognition receptors and triggers plant innate immunity. Further data illustrates that Fg employs H3K4me3 modification to induce BCG1 expression rapidly during the early infection, and then switches to bivalent H3K4me3-H3K27me3 chromatin state that renders rapid epigenetic silencing of BCG1 for escaping from host immune monitor, therefore leading to the successful invasion. Collectively, our study highlights the molecular mechanism of how fungal pathogen employs bivalent epigenetic modification to facilitate the successful infection by escaping of host immunity, which provides conceptual insights into plant-microbe interaction.

overall_design: It was proved by ChIP-seq data and sequential ChIPd ata that modulation of BCG1 expression dynamics by bivalent histone methylations enables fungal pathogen to evade host immunity

first: Qin

last: Gu

first: Xiaozhen

last: Zhao

first: Bingqin

last: Yuan


title: Bulk RNA-seq of mouse neural stem and progenitor cells (NSPC) derived from embryonic stem cells, with allele-specific deletions of a Sox2 enhancer cluster

type: Expression profiling by high throughput sequencing

summary: We have generated RNA-sequencing datasets of the regulome of mouse neural stem and progenitor cells derived from embryonic stem cells, with allele-specific deletions of Sox2 enhancer cluster regions. RNA-seq experiments were conducted to evaluate the regulatory function of Sox2 candidate enhancers in neural stem and progenitor cells.

overall_design: Bulk RNA-seq of mouse neural stem and progenitor cells (NSPC) derived from embryonic stem cells, with allele-specific deletions of clustered Sox2 regulatory regions (SRRs). Samples of clones harboring a biallelic deletion of Sox2 Regulatory Regions 2-18 (SRR2-18) and parent line controls were cultured in a selective medium to induce differentiation of NSPCs for 10 days.

first: Sakthi

last: Moorthy


title: KAT5 activity regulates quiescent-like states in human gliomas [scRNA-seq]

status: Public on Dec 01 2023

city: Seattle

first: Sonali

last: Arora

state: WA

address: 1100 Fairview Ave N,

country: USA

institute: FHCRC

summary: In solid tumors, quiescent/G0 cell populations likely play important roles in maintaining cellular heterogeneity and promoting recurrence after stand of care. However, little is known about the mechanisms of tumor cell G0 ingress and egress. To discover regulators of G0-like states for glioblastoma (GBM), we performed a genome-wide CRISPR-Cas9 screen in patient-derived GBM stem-like cells (GSCs) for genes that when inhibited trap cells in G0-like states. We identify the protein acetyltransferase KAT5 as a key regulator of G0 and cell cycle dynamics in GSCs and GSC-derived tumors. In primary gliomas, KAT5low cells display quiescent properties, while overall KAT5 activity increases as tumors become more aggressive.  Further, we find that KAT5 activity suppresses the emergence of non-dividing subpopulations with oligodendrocyte progenitor and radial glial cell characteristics both in vitro and in a GSC tumor model. These results reveal that KAT5 activity regulates transitions between non-dividing, neurodevelopmental, and proliferative states in GBM tumors.

overall_design: samples of GSC827 were analyzed through single-cell RNA-sequencing using the GemCode platform (10x Genomics). The samples were either the entire poulation of cells or sorted for G0/G1 throught the presence of a G1 reporter, mCherry-CDT1 (aa30/120).

first: Anca

last: Mihalas

first: Samantha

last: Oconnor

first: John

last: Bassett

first: Christopher

last: Plaisier

first: Anoop

last: Patel

first: Patrick

middle: J

last: Paddison


title: Microbiome remodeling through bacterial competition and host behavior enables rapid adaptation to environmental toxins

status: Public on Nov 09 2023

city: Albany

first: Dan

middle: Jinwon

last: Kim

state: CA

address: 675 kula gulf way

department: Integrative Biology

institute: University of California, Berkeley

summary: Human activity is altering the environment at a rapid pace, challenging the adaptive capacities of genetic variation within animal populations. Animals also harbor extensive gut microbiomes, which play diverse roles in host health and fitness and may help expanding host capabilities. The unprecedented scale of human usage of xenobiotics and contamination with environmental toxins describes one challenge against which bacteria with their immense biochemical diversity are particularly suited to offer solutions. To explore the paths leading to bacteria-assisted rapid adaptation, we used Caenorhabditis elegans harboring a defined microbiome, and the antibiotic neomycin as a model toxin, harmful for the worm host and neutralized to different extents by microbiome members. Worms exposed to neomycin showed delayed development and decreased survival but were protected when colonized by neomycin-resistant members of the microbiome. Through a combination of 16S gene sequencing, counting of live bacteria and behavioral assays we identified two distinct mechanisms that facilitated adaptation: gut enrichment for a neomycin-modifying strain driven by altered bacterial competition; and host avoidance behavior, which depended on the stress-activated KGB-1/JNK and enabled preference of neomycin-protective bacteria. The straightforwardness of these mechanisms  suggests that bacteria-assisted host adaptation may be more common than currently appreciated, protecting animals from novel stressors. However, gut remodeling may also cause dysbiosis, and additional experiments identified fitness trade-offs including increased susceptibility to infection as well as metabolic remodeling. Extending these results to other toxins suggests yet unaccounted-for microbiome-dependent long-term consequences of toxin exposure.

overall_design: To investigate how neomycin affects C. elegans, worms raised on E. coli or a complex community (CeMbio) were subjected to 0.25mg/ml neomycin for 8 hours. Comparative gene expression profiling was performed using the RNA-seq data of worms with or without exposure to neomycin with respect to each bacterial community.

first: Michael

last: Shapira


title: Wnt signaling regulates ion channel expression to promote smooth muscle and cartilage formation in developing mouse trachea

status: Public on Nov 06 2023

city: Cincinnati

first: Debora

last: Sinner

state: OH

address: 3333 Burnet Ave

department: Neonatology and Pulmonary Biology

institute: CCHMC

summary: Ion channels play critical roles in the physiology and function of the nervous system and contractile tissue; however, their role in non-contractile tissue and embryonic development has yet to be understood. Tracheobronchomalacia (TBM) and complete tracheal rings (CTR) are disorders affecting the muscle and cartilage of the trachea and bronchi, whose etiology remains poorly understood. We demonstrated that trachealis muscle organization and polarity are disrupted after epithelial ablation of Wls, a cargo receptor critical for the Wnt signaling pathway, in developing trachea. The phenotype resembles the anomalous trachealis muscle observed after deletion of ion channel encoding genes in developing mouse trachea. We sought to investigate whether and how the deletion ofWlsaffects ion channels during tracheal development. We hypothesize that Wnt signaling influences the expression of ion channels to promote trachealis muscle cell assembly and patterning. Deleting Wls in developing trachea causes differential regulation of genes mediating actin binding, cytoskeleton organization, and potassium ion channel activity. Wnt signaling regulated expression of Kcnj13, Kcnd3, Kcnj8, and Abcc9 as demonstrated by in vitro studies and in vivo analysis in Wnt5a and b-catenin deficient tracheas. Pharmacological inhibition of potassium ion channels and Wnt signaling impaired contractility of developing trachealis smooth muscle and formation of cartilaginous mesenchymal condensation. Thus, in mice, epithelial-induced Wnt/b-catenin signaling mediates trachealis muscle and cartilage development via modulation of ion channel expression, promoting trachealis muscle architecture, contractility, and cartilaginous extracellular matrix. In turn, ion channel activity may influence tracheal morphogenesis underlying TBM and CTR.

overall_design: E13.5 tracheal Epithelial cells and Myoblasts were isolated by FACS selection from, Bmp4 f/f; gammaSMAeGFP mouse trachea using APC-Epcam stain to select and isolate epithelial cells and GFP to isolate muscle cells.

first: Nicholas

last: Russell

first: Natalia

last: Bottasso Arias

last: Snowball


title: Activation of automethylated PRC2 by dimerization on chromatin

city: Cologne

first: Simon

last: Poepsel

address: Robert-Koch-Str. 21

department: Center for Molecular Medicine Cologne

country: Germany

institute: University of Cologne

summary: Polycomb Repressive Complex 2 (PRC2) trimethylates lysine 27 of histone 3 (H3K27me3). This modification transcriptionally represses chromatin and its dynamics are essential for embryonic development. PRC2 also methylates its cofactor JARID2 and both methylated H3 tail and JARID2 allosterically activate the complex. PRC2 undergoes auto-methylation of its catalytic subunit, which stimulates its activity by an unknown mechanism. We show that automethylated PRC2 forms a dimer on substrate chromatin that results in autoactivation in trans. An inactive PRC2 protomer distal to the substrate nucleosome serves as an allosteric activator of the second PRC2 protomer, which is poised to methylate H3. Functional assays support our model of allosteric activation via dimerization, suggesting a novel mechanism of PRC2 trans-autoactivation that may be important for installation of H3K27me3 into naïve chromatin.

overall_design: To investigate PRC2-mediated effects during retinoic acid-driven cell differentiation, we established Ezh2 rescue cell lines in Ezh1-/-;Ezh2-/- murine embryonic stem cells (N/A Strain of origin 129P2/Ola; Lavarone etal., 2019) and analysed transcriptomic changes over the course of 4 days. To investigate PRC2-mediated effects during retinoic acid-(RA-)driven cell differentiation, we established SUZ12 rescue cell lines in SUZ12-/- murine embryonic stem cells and analysed transcriptomic changes over the course of 4 days.

first: Paul

last: Sauer

first: Egor

last: Pavlenko

first: Trinity

last: Cookis

first: Linda

last: Zirden

first: Juliane

last: Renn

first: Ankush

last: Singhal

first: Pascal

last: Hunold

first: Michaela

middle: N

last: Hoehne

first: Olivia

last: van Ray

first: Robert

last: Hänsel-Hertsch

first: Karissa

middle: Y

last: Sanbonmatsu

first: Eva

last: Nogales


title: SuperSeries=Mouse neural stem and progenitor cells (NSPC) derived from embryonic stem cells, with biallelic Sox2 enhancer cluster deletions

status: Public on Nov 10 2023

first: Jennifer

last: Mitchell

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

overall_design: Refer to individual Series


title: Mitochondria as key therapeutic targets for the treatment of corneal fibrosis

status: Public on Nov 02 2023

city: Rush

first: keith

last: nehrke

state: NY

address: 1514 Rush Scottsville Rd

department: Medicine

institute: University of Rochester Medical Center

summary: Over the past decade, our laboratory developed a cat model of corneal wound healing after photorefractive keratectomy (PRK). Recently, we used this model in a combination of in vivo and in vitro approaches to show that myofibroblasts directly inhibit nerve regeneration in the wounded stroma, sub-basal plexus and epithelium. In vitro, two distinct phases to this inhibition have been observed: (1) initial slowing of neurite elongation via a releasable factor, and (2) neurite contact inhibition by myofibroblasts. For phase 1, transforming growth factor beta 1 (TGF-1) was shown to be the soluble, anti-neuritogenic factor released by myofibroblasts, and the specific signaling cascade it triggered to slow neurite elongation was identified. Ultimately, preventing myofibroblast differentiation suppressed both phase 1 and phase 2 inhibition, and resulted in faster nerve regeneration. However, in most cases of accidental injury, patients present after myofibroblast differentiation/fibrosis have occurred. The issue then is no longer preventing fibrosis, but rather overcoming its effects. We recently discovered that the synthetic PPAR ligand and Thiazolidinedione (TZD) drug, Troglitazone, stimulates myofibroblast de-differentiation in vitro and in vivo through a PPAR independent pathway. We now show that this TDZ’s ability to promote myofibroblast de-differentiation can be mimicked by UK-5099, a compound that spares PPAR, instead inhibiting a recently identified alternative target of TZDs: the mitochondrial pyruvate carrier (MPC). The recent literature and our preliminary results thus form a strong premise for refocusing our efforts on mitochondria, and critically testing the hypothesis that inhibiting the MPC facilitates myofibroblast de-differentiation, which together with metabolic remodeling in regenerating nerves can treat both fibrosis and overcome the blocked reinnervation inherent in its context. Work is ongoing to understand the metabolic changes that underlie the ability of TDZs to facilitate wound healing though mito-centric regulation of both myofibroblasts and corneal nerves. Here, we present the results of an RNA sequencing experiment designed to examine changes to the global transcriptome that occur as a result of TGF-β1-induced activation of corneal fibroblasts, and of TDZ- or UK5099-stimulated myofibroblast de-differentiation.

overall_design: Cat corneal fibroblasts in culture (d0FB) were treated for two days with TGFb1 to induce differentiation to myofibroblasts (d2TGFb). Myofibroblasts were then treated for three days with either Troglitazone (d5Trog), Troglitazone and GW9662 (d5T-G), or UK-5099 (d5UK). Five independent replicates of cells from each of these five conditions were harvested for RNA sequencing.

first: Huxlin

last: Krystel R.

first: Nehrke

last: Keith W

first: Jeon

last: Kyeim

first: Kumar

last: Ankita


title: Single-cell analysis of chromatin accessibility in the adult mouse brain

city: La Jolla

first: Songpeng

last: Zu

address: 9500 Gilman Dr

department: Department of Cellular and Molecular Medicine

institute: University of California San Diego

summary: Here we report a comprehensive atlas of candidate cis-regulatory DNA elements (cCREs) in the adult mouse brain, generated through examination of the chromatin accessibility in 2.3 million individual brain cells from 117 anatomical dissections. The atlas includes approximately 1 million cCREs and their chromatin accessibility across 1482 distinct brain cell populations, adding over 446,000 new cCREs to the most recent such annotation in the mouse genome. Raw fastq files are demultiplexed. Out of 234 samples, fastq files for 6 samples are available in NCBI under the accession number GSE126724 (CEMBA180308_3B, CEMBA180312_3B, CEMBA180226_1A, CEMBA180227_1A, CEMBA180305_2B, CEMBA180306_2B). Processed data are SnapATAC2 (<= 2.4.0) h5ad files per sample, with raw fragments and 500-bp bmat. NOTE: due to the break changes in SnapATAC2 >= 2.5.0, the data can not be handled rightly in SnapATAC2 >= 2.5.0.

overall_design: We generated single-nucleus assys for transposase-accessible chromatin using sequencing (snATAC-seq) libraries from 117 anatomic dissections of mouse brain and two replicates for each dissection.

first: Yang

middle: E

last: Li

first: Seoyeon

last: Lee

first: Sebastian

last: Preissl

first: Bing

last: Ren


title: Cellular and Extracellular microRNA Dysregulation in LRRK2-Linked Parkinson’s Disease

status: Public on Oct 31 2023

city: Tübingen

first: Jun-Hoe

address: Otfried-Müller-Str. 37

department: Quantitative Biology Centre (QBiC)

institute: University of Tuebingen

type: Non-coding RNA profiling by high throughput sequencing

summary: The discovery of cell-free micro-RNAs in body fluids has made them a promising biomarker target in the field of neurodegenerative diseases. Although they have been reported to be differentially expressed in biofluids and tissue from sporadic Parkinson’s disease patients, it remains unclear whether similar observations can be made in patients with genetic forms of the disease. Since induced pluripotent stem cell derived neurons represent a widely used research model for both sporadic and familial Parkinson’s disease, we sought to assess the usability of this model for the identification of differentially expressed cell-free micro-RNAs in the context of the Parkinson’s disease related LRRK2 G2019S mutation in a proof-of-concept study. We isolated extracellular vesicles carrying cell-free RNA from patient-derived induced pluripotent stem cell lines carrying the LRRK2 G2019S mutation and their gene corrected isogenic controls. After generation of small-RNA libraries and differential expression analysis, we validated fourteen micro-RNAs in an independent batch of cell-free and cellular RNA via RT-qPCR. Finally, we selected eleven differentially expressed micro-RNAs from our cell culture experiments and quantified their expression levels in cerebrospinal fluid derived from two LRRK2 G2019S patients and two healthy controls.

overall_design: Isolated extracellular vesicles carrying cell-free RNA from patient-derived induced pluripotent stem cell lines carrying the LRRK2 G2019S mutation and their gene corrected isogenic controls.

first: Felix

last: Knab

first: Kevin

last: Menden

first: Fatma

last: Busra Isik

first: Anto

middle: P

last: Rajkumar

first: Stefan

last: Czemmel

first: Julia

last: Fitzgerald

first: Patrizia

last: Rizzu

first: Thomas

last: Gasser


title: Transcriptional reprogramming during human osteoclast differentiation identifies regulators of osteoclast activity

status: Public on Nov 05 2023

city: Odense

first: Alexander

last: Rauch

address: Campusvej 55

department: Department of Clinical Research

country: Denmark

institute: University of Southern Denmark and Odense University Hospital

summary:
Enhanced osteoclastogenesis and osteoclast activity contribute to the development of osteoporosis, which is characterized by increased bone resorption and inadequate bone formation. As novel anti-osteoporotic therapeutics are needed, understanding the genetic regulation of human osteoclastogenesis could help identify potential treatment targets. This study aimed to provide an overview of the transcriptional reprogramming during human osteoclast differentiation. Osteoclasts were differentiated from CD14+-monocytes from eight female donors. RNA-sequencing during differentiation demonstrated 8980 differentially expressed genes grouped into eight temporal patterns conserved across donors. These patterns showed distinct molecular functions, associated with postmenopausal osteoporosis susceptibility genes based on RNA from iliac crest biopsies, and bone mineral density SNPs. Network analyses showed mutual dependencies between the temporal expression patterns and provides insight into subtype-specific transcriptional networks. Donor specific expression patterns identified genes at monocyte stage, such as filamin B (FLNB) and oxidized low density lipoprotein receptor 1 (OLR1, encoding LOX-1), that are predictive for the resorptive activity of mature osteoclasts. Differentially expressed G-protein coupled receptors showed strong expression during osteoclast differentiation and associated with bone mineral density SNPs, implying a pivotal role in osteoclast differentiation and activity. The regulatory effects of three differentially expressed G-protein coupled receptors were exemplified by in vitro pharmacological modulation of complement 5A receptor 1 (C5AR1), somatostatin receptor 2 (SSTR2), and free fatty acid receptor 4 (FFAR4/GPR120). Activating C5AR1 enhanced osteoclast formation, while activating SSTR2 decreased resorptive activity of mature osteoclasts, and activating FFAR4 decreased both number and resorptive activity of mature osteoclasts.
In conclusion, we report the transcriptional reprogramming during human osteoclast differentiation and identified SSTR2 and FFAR4 as anti-resorptive G-protein coupled receptors as well as FLNB and LOX-1 as potential molecular markers of osteoclast activity. These data can help future investigations to identify molecular regulators of osteoclast differentiation and activity and provide the basis for novel anti-osteoporotic targets.

overall_design: RNA-expression profiles from 4 time points during human osteoclasts differentiated in vitro from CD14 positive peripheral blood cells of 8 anonymous donors.

first: Morten

middle: S

last: Hansen

first: Kaja

last: Madsen

first: Maria

last: Price

first: Kent

last: Søe

first: Yasunori

last: Omata

first: Mario

middle: M

last: Zaiss

first: Caroline

last: Gorvin

last: Frost


title: NicOPURE: Nickless RNA Circularization and One-Step Purification with Engineered Group II Introns and Cyclizing UTRs

status: Public on Oct 29 2023

city: Sg

first: XIANG

last: GAO

country: Singapore

institute: GIS

summary: Circular RNAs (circRNAs) have gained significant attention in recent years due to their diverse biological functions and potential as novel therapeutic modality. Two general mechanisms for generating circRNAs involves the utilization of group 1 and group 2 introns, which are self-splicing ribozymes found in many organisms. Although group 1 intron has been demonstrated to be highly effective in circularization, the reaction requires high temperature plus cofactors such as GTP. Consequently, undesired byproducts such as nicked RNA were generated, which requires complex purification steps before downstream applications. In this study, we have strategically designed structural elements and incorporated sequence features to enhance the efficacy of RNA circularization by group 2 introns. This innovative approach occurred at a reduced temperature and resulted in notably fewer nicks compared to group 1 introns. Furthermore, to streamline the purification process of circular RNA, we incorporated two halves of streptavidin aptamer tags into the enzymatic sites of the group 2 intron. This strategic architecture guarantees the creation of a fully operational streptavidin aptamer tag solely at the circular RNA junction site during in vitro circularization. This unique attribute facilitates a one-step purification process. The resulting "NicOPURE" system emerges as both straightforward and efficient in generating therapeutic circular mRNAs.

overall_design: To understand the structural and functional distinctions existing within Group 1 and Group 2 intron circular mRNAs. 100 μg of circular mRNA (circGLUC) for the same target gene were generated using two distinct approaches: one utilizing a Group 1 intron (Anabaena) and the other employing the NicOPURE system with Group 2 intron (Clostridium tetani). We probed the group 1 and group 2 circular mRNA secondary structure using RNA structure probing method: Selective 2’-hydroxyl acylation analyzed by primer extension and mutational profiling (SHAPE-MaP). SHAPE reactivities were computed for the GLUC open-reading frame (ORF) in both group 1 and group 2 circular mRNA using Shapemapper2.

first: Jovi

middle: L

last: An

first: GAO

last: Xiang


title: MicroRNA-1 protects the endothelium in acute lung injury

status: Public on Oct 25 2023

city: New Haven

first: Asawari

last: Korde

state: CT

address: 300 Cedar street

institute: Yale University School of Medicine

summary: Acute lung injury (ALI) and its most severe form, acute respiratory distress syndrome (ARDS), cause severe endothelial dysfunction in the lung and vascular endothelial growth factor (VEGF) is elevated in ARDS. We have found that the levels of a VEGF-regulated microRNA, microRNA-1 (miR-1) is reduced in the lung endothelium after acute injury. Pulmonary endothelial cell (EC)-specific overexpression of miR-1 protects the lung against cell death and barrier dysfunction in both murine and human models and increases the survival of mice after pneumonia-induced ALI. MiR-1 has an intrinsic protective effect in pulmonary and other types of ECs; it inhibits apoptosis and necroptosis pathways and decreases capillary leak by protecting adherens and tight junctions. Comparative gene expression analysis and RISC recruitment assays identified miR-1 targets in the context of injury, including  phosphodiesterase 5A (PDE5A), angiopoetin-2 (ANGPT2), connector enhancer of kinase suppressor of ras 3 (CNKSR3) and TNF alpha induced protein 2 (TNFAIP2). We validated miR-1-mediated regulation of ANGPT2 in both mouse and human ECs and found that in a 119-patient pneumonia cohort, miR-1 correlated inversely with ANGPT2. These findings illustrate the novel role of miR-1 as a cytoprotective orchestrator of endothelial response to acute injury with prognostic and therapeutic potential.

overall_design: mRNA sequencing of HUVEC transfected with microRNA-1 or control RNA after RNA immunoprecipitation with Argonaute-2 protein, 3 replicates per condition

last: Haslip

first: Prachi

last: Pednekar

first: Alamzeb

last: Khan

first: Maurizio

last: Chioccioli

first: Sameet

last: Mehta

first: Francesc

last: Lopez-Giraldez

first: Santos

last: Bermejo

first: Mauricio

last: Rojas

first: Charles

last: Dela Cruz

first: Michael

last: Matthay

first: Jordan

last: Pober

first: Richard

last: Pierce

first: Shervin

last: Takyar


title: RNA-sequencing analysis to identify genes regulated by MYCN in rhabdomyosarcoma cell line RH4

status: Public on Oct 19 2023

city: Bethesda

first: Zhihui

last: Liu

state: Maryland

address: 9000 rockville pike

institute: National Cancer Institute

summary: In this study we identified genes regulated by MYCN in RH4 cells

overall_design: RNA was isolated from RH4 cells that transfected with siCtrl or siMYCN for 72 h.

first: Carol

last: Thiele


title: RNA-sequencing analysis to identify genes regulated by MYCN in neuroblastoma cell line SHEP by MYCN overexpression

summary: In this study we identified genes regulated by MYCN in SHEP cells

overall_design: SHEP cell line stably transfected with doxycycline inducible MYCN overexpression vector was named SHEPtetMYCN. RNA was extracted from SHEPtetMYCN cells treated with or without 0.25 µg/ml doxycycline for 48 h.


title: UDCA regulated the proliferation and function of DNT cells via FXR

first: Chunpan

last: Zhang

address: No. 95 Yong-An Road

institute: Beijing Friendship Hospital

summary:
Primary biliary cholangitis (PBC) is a liver-specific autoimmune disease. Treatment of PBC with ursodeoxycholic acid (UDCA) is not sufficient to prevent disease progression, new and creative approaches to treating patients with PBC are urgently required. Double-negative T (DNT) cells, unique regulatory T cells, play crucial roles in maintaining immune system homeostasis. However, both the absolute number and proportion of liver DNT cells were reduced in PBC patients. Hence, it is worth pursuing that whether replenishment of DNT cells can improve the hepatic function and pathology of PBC. In this study, adoptive transfer of DNT cells significantly decreases plasma levels of ALT, AST, AMA-M2 and IgM in both dnTGFβ RII and 2OA-BSA-immunized PBC mouse models. In addition, DNT cells exhibit a strong suppression on liver T cells. While combination therapy with DNT cells and UDCA predominantly ameliorate liver inflammation and extensively inhibit hepatic T and B cells. In vitro study further reveals that UDCA promotes the proliferation of DNT cells, increased functional molecule perforin and reduced inhibitory molecule NKG2A and EPCR expression, resulting in augmenting the suppressive function of DNT cells via FXR/JNK signaling pathway in both mice and human DNT cells. In conclusion, UDCA augments the suppressive function of DNT cells via FXR/JNK. A single transfer of DNT cells in combination with UDCA strongly ameliorates PBC in mice. This study provides a potential application of DNT cell-based therapy for PBC.
Methods: The DNT cells incubation with anti-CD3/CD28 antibodies were stimulated with UDCA (60uM) for 48 hours, then transcriptome sequencing studies were performed on DNT cells RNA.Transcriptome sequencing libraries were generated using NEBNext® Ultra™ RNA Library Prep Kit for Illumina® (NEB, USA) following manufacturer’s recommendations and sequenced on an Illumina Hiseq platform (Illumina, San Diego, CA). Sequences were aligned to the reference genome with TopHat and processed with Cufflinks, which quantifies each transcript in each sample using reference annotations produced by the University of California Santa Cruz UCSC. Differentially expressed genes with a fold change of >=1.41 and pvalue <= 0.05 between UDCA treated and control DNT cells were submitted to GO and KEGG enrichment analysis, which uses unbiased methods to assess pathway enrichment.

overall_design: DNT cells mRNA profiles of control and UDCA stimulated were generated by deep sequencing in duplicate, using Illumina HiSeq.

first: Guangyong

last: Sun

first: Hua

last: Jin

first: Yunxiong

last: Wei

first: Shimeng

last: Zheng

first: Xiyu

last: Wang

first: Xinyan

last: Zhao

first: Dong

first: Jidong

last: Jia


title: The cooperation between MYCN and its cofactors WDR5 and G9a on gene transcription regulation [siWDR5]

summary: MYCN cooperates with WDR5 to activate canonical MYC targets and cooperates with G9a to repress neuronal differentiation genes.

overall_design: IMR32 cells were transfected with control siRNA (siCtrl) or WDR5 siRNA (siWDR5) and cultured in complete medium for 72 hr, then ChIP experiments were performed using  ChIP-IT High Sensitivity Kit (Active Motif ).


title: The cooperation between MYCN and its cofactors WDR5 and G9a on gene transcription regulation [siMYCN]

overall_design: IMR32 cells were transfected with control siRNA (siCtrl) or MYCN siRNA (siMYCN) and cultured in complete medium for 72 hr, then ChIP experiments were performed using  ChIP-IT High Sensitivity Kit (Active Motif ).


title: RNA-sequencing analysis to identify genes regulated by MYCN in neuroblastoma cell line KCNR.

summary: In this study we identified genes regulated by MYCN in KCNR cells

overall_design: RNA was isolated from KCNR cells that transfected with siCtrl or siMYCN for 72 h.


title: Active regulatory elements recruit cohesin to establish cell-specific chromatin domains [RNA-seq]

status: Public on Oct 13 2023

city: Oxford

first: Emily

last: Georgiades

address: John Radcliffe Hospital/Headley Way

country: United Kingdom

institute: Weatherall Institute of Molecular Medicine

summary: The genome adopts varied and specific 3D conformations in different cell types.  The loop extruding factor, cohesin, is thought to play a key role in the formation of this 3D structure, however it is unknown how cell-type specificity is achieved. One proposed mechanism is that cis-regulatory elements, which are active in unique combinations in different cell types, recruit cohesin and therefore direct the formation of specific 3D conformation.  We have identified genome-wide natural variation in enhancer sites in healthy human donors, by performing cohesin ChIP-seq in these cells we show that cohesin occupancy is directly correlated with enhancer activity.  Identification of the genetic variants associated with the gain or loss of enhancer-like elements and associating this with the recruitment of cohesin provides evidence of a direct link between widespread genetic variation in the non-coding genome and cell-specific changes in 3D genome organization. Therefore, enhancer elements have evolved not only the ability to upregulate transcription at promoters, but also the ability to stimulate the mechanisms that insure robust interactions between them.   Next, using enhancer KO models in the well characterised alpha globin locus, we show how cohesin recruitment and 3D genome organization correlates with changes in the local regulatory landscape. We then selected the strongest of the alpha globin enhancers and inserted it into a neutral region of the genome, to test if a single active enhancer was capable of recruiting cohesin and initiating the formation of a 3D chromatin domain when isolated from its native context.  Our results show that an enhancer is able to direct the recruitment of cohesin when in an active state and that this is correlated with changes in the local chromatin environment, giving rise to open chromatin sites and de novo chromatin interactions in an otherwise “neutral” region of the genome.  These results support the hypothesis that tissue-specific recruitment of cohesin is facilitated by enhancers and that this plays a role in establishing cell-specific changes in 3D genome conformation.

overall_design: CD71+ erythroid cells were harvested from day 7 embryoid bodies. PolyA+ and polyA- RNA seq was performed on three WT replicates (A-C) as controls, and three independently targeted R2-insertion clones (B6, E6 and D10).

first: Caz

last: Harrold

first: Nigel

last: Robert

first: Marieke

last: Oudelaar

first: Joe

last: Blayney


title: The cooperation between MYCN and its cofactors WDR5 and G9a


title: Active regulatory elements recruit cohesin to establish cell-specific chromatin domains [ATAC-seq]

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: CD71+ erythroid cells were harvested from day 7 WT embryoid bodies and three replicates of ATAC-seq was performed.  CD34+ cells were harvest on day 13 for donors 001, 002 and 030 and two replicates of ATAC-seq performed for each.


title: Active regulatory elements recruit cohesin to establish cell-specific chromatin domains [Capture-C]

overall_design: Capture-C was performed on CD71+ erythroid cells harvested from day 7 WT and R2-insertion embryoid bodies (EB) as per the published protocol.

first: Caroline

first: Mira

last: Kassouf

first: Helena

last: Francis

first: Joseph

first: Simone

middle: G

last: Riva

first: Edward

last: Sanders

first: Thomas

last: Milne

first: Douglas

middle: R

last: Higgs

first: Jim

last: Hughes


title: RNA-sequencing analysis to identify genes regulated by MYCN, WDR5 and G9a in neuroblastoma cell line IMR32

first: Zhihui

last: Liu

summary: In this study we identified genes regulated by MYCN, WDR5 and G9a in IMR32 cells

overall_design: RNA was isolated from IMR32 cells that transfected with siCtrl, siMYCN, siWDR5 or siG9a for 48 h.

first: Carol

last: Thiele


title: Active regulatory elements recruit cohesin to establish cell-specific chromatin domains [ChIP-seq]

overall_design: ChIP-seq was performed on aliquots of 3-10x10^6CD71+ cells. For calibrated ChIP-seq, a 1% or 4% spike-in of WT HEK293 cells were added prior to sonication. Cells were double cross-linked using disuccinimidyl glutarate (DSG, Sigma) and 1% formaldehyde (Sigma) for a total fixation time of 1 hour. Fixed chromatin samples were fragmented using the Covaris sonicator (ME220) for 10 minutes (75 power, 1000 cycles per burst, 25% duty factor) at 4°C. 50μL per sample was removed as an input control. Sonicated samples were pre-cleared to remove background signal through incubation with a 1:1 mix of Protein A/G Dynabeads (InVitrogen). Antibody was added at a concentration of 1 μg/μL per sample and immunoprecipitated at 4°C overnight.  Immunoprecipitated samples were incubated with a 1:1 mix of Protein A/G Dynabeads for 5 hours at 4°C. Beads were then washed 4x in RIPA buffer on a magnetic stand, followed by 1x wash with TE (Sigma Aldridge) + 50mM NaCl (ThermoFisher). Chromatin was eluted from the beads using elution buffer and incubated for 30 minutes at 65°C with shaking. Input samples were diluted 1:1 with elution buffer. Samples and input controls were incubated at 65°C overnight for de-crosslinking, before RNase (Roche) and proteinase K (BioLabs) treatment. DNA fragments were purified using the Zymo ChIP DNA Clean & Concentrator kit (Zymo Research) and eluted in water. DNA concentration was quantified using the Qubit dsDNA HS assay (Invitrogen) as per manufacturers protocol. To assess sonication efficiency the D1000 Tapestation (Agilent) assay was performed on input sample.  An approximately equal mass of input and IP DNA was used for indexing (0.5 ng – 1 μg). NEBNext Ultra II DNA Library Prep Kit (New England Biolabs) was used to prepare indexed sequencing libraries following the manufacturer supplied protocol.  PCR amplification was performed for 7-11 cycles (depending on input DNA concentration) using NEBNext Mulitplex Oligos (New England Biolabs). Indexed sample concentration was quantified using the KAPA Library Quantification Complete Kit (Universal)(Roche).  Samples were pooled as a 4 nM library and sequenced with a high-Output v2 75 cycle kits on the Illumina NextSeq platform.

last: Roberts


title: Ultrafast Bisulfite Sequencing for Efficient and Accurate 5-Methylcytosine Detection in DNA and RNA

city: Chicago

first: Chang

last: Ye

state: Illinois

institute: University of Chicago

type: Methylation profiling by high throughput sequencing

summary: We present an ultrafast bisulfite sequencing (UBS-seq) method, which accelerates the bisulfite reaction by about 13-fold, resulting in minimal DNA damage and much lower background in all DNA sequences. This ultrafast BS protocol is also ideal for mapping of RNA 5-methylcytosine (m5C) because of much shorter reaction time and dramatically reduced RNA background. And allowed detection of m5C stoichiometry in highly structured RNA species.

overall_design: Compare bisulfite conversion efficiency of UBS and conventional-BS reagent


title: Active regulatory elements recruit cohesin to establish cell-specific chromatin domains.

first: Emily

overall_design: Refer to individual Series


title: Descending thoracic aorta perivascular adipocytes adipogenesis is defined by their anatomical location

status: Public on Oct 11 2023

city: East Lansing

first: G. Andres

last: Contreras

state: MI

address: 736 Wilson Road Room D202, Michigan State University

department: Large Animal Clinical Sciences

institute: Michigan State University

summary: Hypertension, atherosclerosis, and aneurysms alter thoracic aorta structure. Aortic lesions found in these diseases show a unique anatomical distribution. For instance, calcifications and atherosclerotic lesions tend to occur more frequently in the posterior wall of the aorta compared to other regions. The role that the outer layer of the aorta, its perivascular adipose tissue (PVAT), plays in the pathogenesis of these lesions is unknown. The descending thoracic aorta's PVAT is distributed in three strips of tissue: one strip is located anterior to the aorta (AP), while the other two are positioned laterally to the vessel and are adjacent to the thoracic vertebrae (LP). Genetic tracing indicates LP's adipocytes descend from sm22a+ and Myf5+ progenitors while the anterior are from sm22a+ only. The implications of this ontology and aortic PVAT distribution on the development of adipocytes are unknown. We hypothesize that the anatomical location of adipocyte progenitors influences their adipogenic potential. PVAT from LP and AP was collected from male SD rats at 10 wks of age (n=7) to harvest progenitors by outgrowth expansion. Progenitors were differentiated for 4 d in adipogenic media. Adipogenesis was evaluated by lipid droplet and triglyceride quantification using the IncuCyte Live-Cell® system. RNA from progenitors and adipocytes was sequenced in Illumina NextSeqData, and Differential Expressed Genes (DEG) identified. Enrichment and network analyses were performed in Ingenuity Pathways (IPA). Our findings provide evidence supporting differences in adipogenic activity and extracellular matrix secretion between the LP and AP PVAT of the aorta. These differences may explain the anatomical location of aortic lesions associated with hypertension, atherosclerosis, and aneurysms.

overall_design: To investigate the adipogenic potential of adipocyte progenitors in the descending thoracic aorta perivascular adipose tissue

first: C. Javier

last: Rendon

first: Adam

last: Lauver

first: Stephanie

last: Watts


title: Mucosal immune alterations at the early onset of tissue destruction in Chronic Obstructive Pulmonary Disease

status: Public on Oct 09 2023

first: John

last: McDonough

address: 300 Cedar St

institute: Yale School of Medicine

summary: In 11 control (non-used donors) and 11 COPD (end-stage) explant frozen lungs, 4 cylinders/cores were processed per lung for microCT and tissue transcriptomics. MicroCT was used to quantify tissue percentage and alveolar surface density to classify the COPD cores in mild, moderate and severe zones, as well as to quantify terminal bronchioles. Transcriptomics of each core assessed fold changes in innate and adaptive cells and pathway enrichment score between control and COPD cores. Immunostainings of immune cells was performed for validation.

overall_design:
We sampled multiple regions in human lungs collected following lung transplantation for severe COPD and determined the extent of emphysema using microCT quantitative imaging and tissue histology to classify them as early, progressive, or end-stage disease. We then performed RNA-seq profiling to determine gene expression at these selected stages and applied a linear mixed-effects model to identify differentially expressed genes.
24 control Samples from GSE184316 (GSM5583910-GSM5583933, PRJNA763978) are reanalyzed in this study.

first: Charlotte

last: de Fays

first: Bart

last: Vanaudenaerde


title: UBE2A/B is the trans-acting factor mediating mechanotransduction and contact inhibition

city: Tianjin

first: Fumihiko

last: Nakamura

address: 92 Weijin Road, Nankai District

institute: Tianjin University

summary: Mechanotransduction converts mechanical stimuli into biochemical signals to regulate cell growth, differentiation, and survival. However, how mechanotransduction is linked to gene expression remains poorly understood. Here, we report the identification of UBE2A as an mechano-sensitive trans-acting factor, which translocates from the cytosol to the nucleus upon mechanical stimuli. UBE2A is an E2 enzyme that catalyzes ubiquitination of histone, which eventually results in transcription activation of downstream genes. Using ChIP-seq and RNAseq analysis, we identify downstream genes of UBE2A, including YAP1, a well-known mechanotransducer. Knock-down of UBE2A/B significantly downregulates YAP1 protein expression. Our findings identify the new mechanotransduction pathway that regulates gene expression through histone ubiquitination by UBE2A/B.

overall_design:
Examination of trans-acting factors involved in mechanotransduction, identify candidates' biological function using RNA-seq in KD/normal HEK cell.
mRNA profiles of wild type (WT) and UBE2A/B knock-down (KD) HEK293 cells.

first: Mingwei

last: Feng


title: Gene expression in diabetic RIP-LCMV-GP mice

status: Public on Oct 01 2023

city: Frankfurt am Main

first: Urs

last: Christen

address: Theodor-Stern Kai 7

department: Pharmacenter

institute: Goethe University Frankfurt

summary:
Gene expression profile in laser-dissected islets of Langerhans in the inducible RIP-LCMV-GP mouse model for type 1 diabetes (T1D)
RIP-LCMV-GP mice express the glycoprotein (GP) of the lymphocytic choriomeningitis virus (LCMV) in the beta-cells (rat insulin promotor, RIP); T1D develops 10-14 after LCMV-infection

overall_design: RNA has been isolated from the entire islet microenvironment (including the cellular infiltrates) before and at several times after diabetes initiation by LCMV infection

first: Christine

last: Bender


title: The cooperation between MYCN and its cofactors WDR5 and G9a on gene transcription regulation [siG9a]

overall_design: IMR32 cells were transfected with control siRNA (siCtrl) or G9a siRNA (siG9a) and cultured in complete medium for 72 hr, then ChIP experiments were performed using  ChIP-IT High Sensitivity Kit (Active Motif ).


title: Transcriptome analysis of Streptococcus mutans ΔSMU.1147 strain

status: Public on Oct 03 2023

city: Busan

first: Ryoo

last: SiUk

address: busandaehakro 63-2

department: Microbiology

country: South Korea

institute: Pusan univ.

summary: RNA-seq was performed to examine the overall gene expression in a S. mutans ΔSMU.1147 strain and to identify potential genes that can mainly contribute to the changes of gene expression in S. mutans. Using RNA-seq technique, the differentially expressed genes that are implicated in phenotypic changes of the ΔSMU.1147 strain were studied.

overall_design: Comparative analysis of the S. mutans wild-type strain with the ΔSMU.1147 strain

first: Si-Uk

last: Ryoo

first: Jeong

last: Kim


title: LRRK2 G2019S variant triggers transcriptional changes in Parkinson’s disease human myeloid cells under pro-inflammatory environment

status: Public on Sep 30 2023

city: NEW YORK

first: Hyomin

last: Seo

address: 1 Gustave L. Levy Pl

department: Neurosciences

institute: Icahn school of medicine at Mount Sinai

summary: The G2019S mutation in the LRRk2 gene is a major Parkinson’s disease risk factor. We studied its effect on microglial activity and found that it remains inactive under normal conditions but strongly affects gene regulation when exposed to inflammation such as LPS and IFNg. The mutation increases lipid metabolism and phagolysosomal genes in microglia, enhancing their ability to clear myelin debris. It also disrupts cell cycle genes, decreasing proliferation and E2F4 regular.These findings reveal G2019S’s significant impact on myeloid cell gene activity during inflammation, highlighting lipid imbalance as a key factor in functional disruptions.

overall_design:
All experiments were performed with monocytes derived from cryopreserved PBMCs to avoid any technical variability, and were performed in 2 independent experimental batches
Genotypes: LRRk2 (Wt), LRRK2 G2019S mutation (Het), LRRK2 G2019S mutation (Hom)
Treatments: Basal, IFNg, LPS

first: Elisa

last: Navarro

first: Anastasia

last: Efthymiou

first: Madison

last: Parks

first: Giulietta

last: Riboldi


title: Heat shock response and transposon control in plant shoot stem cells [RNA-seq]

city: Vienna

first: Ruben

last: Gutzat

state: Vienna

address: Dr. Bohr-Gasse 3

country: Austria

institute: Gregor Mendel Institute GmbH

summary: Post-embryonic plant development must be coordinated in response to and with environmental feedback. Development of above-ground organs is orchestrated from stem cells in the center of the shoot apical meristem (SAM). Heat can pose significant abiotic stress to plants and induce a rapid heat shock response, developmental alterations, chromatin decondensation, and activation of transposable elements (TEs). However, most plant heat-stress studies are conducted with seedlings, and we know very little about cell-type-specific responses. Here we use fluorescent-activated nuclear sorting to isolate and characterize stem cells of wild type and mutants defective in TE defense and chromatin compaction after heat shock and after a long recovery. Our results indicate that stem cells can suppress heat shock response pathways to maintain developmental programs.  Furthermore, mutants defective in DNA methylation fail to recover efficiently from heat stress and persistently activate heat shock factors and heat-inducible TEs. Heat stress also induces DNA methylation epimutations, especially in the CHG context, and we find hundreds of DNA methylation changes three weeks after stress. Our results underline the importance of disentangling cell type-specific environmental responses for understanding plant development.

overall_design: Six-day old wt, poliv and, ddm1 plants were exposed to 37 celsius degree for 1 day (D7), then plants were transferred to soil to grow for 21 more days (D28). The plants stem cell nuclei and seedlings at D7 as well as stem cell nuclei and inflorescence apices at D28 were collected for mRNA-seq.

first: Vu

last: Nguyen

first: Ortrun

last: Mittelsten Scheid


title: Pulmonary ketogenesis promotes tolerance to bacterial infection

status: Public on Oct 12 2023

city: New York

first: Sebastian

last: Riquelme

address: 650W 168th Street Black Building 4th Floor Room 427

institute: Columbia University

summary: Tolerance is a major defense strategy against infection. During the host response to pathogens, tolerance restricts inflammatory damage to tissues, maintaining the long-term integrity of organs. The mechanisms that establish tolerance are poorly understood. We analyzed pulmonary isolates of Pseudomonas aeruginosa that evolved to coexist with tolerant hosts and found that these opportunistic pathogens facilitate tolerance by stimulating ketogenesis. Ketone body production in the airway limits the accumulation of detrimental factors that injure the lung, such as inflammatory cytokines and effector phagocytes. Although ketones are typically synthesized in the liver, in situ metabolo-transcriptomic studies revealed that P. aeruginosa drives their generation in the lung by co-opting the metabolism of alveolar macrophages, which are pulmonary cells of hepatic origin. This phenomenon was restricted to clinical isolates and not observed with laboratory strains of P. aeruginosa , confirming the unique metabolo-tolerogenic properties of ESKAPE pathogens adapted to the human lung.

overall_design: WT or Irg1-/- C57bl/6 mice (8 week old) were treated with PBS or infected either with WT PAO1 or a mix of the 17 host-adapted P. aeruginosa clinical isolates (106 total CFU per mouse in 50uL). 16 hours after infection, mice were euthanized, their lungs were harvested, and single cell suspensions of the lungs were prepared as described. Briefly, the lungs were placed in an Eppendorf tube containing an enzymatic digestion solution of collagenase I (2 mg/mL), dispase (20 mg/mL), elastase (1 mg/mL), and DNAse (1 uL/mL) in PBS. The lungs were minced within the tube and then incubated with shaking at 37C for 30 minutes. 4 volumes of PBS supplemented with 10% FBS was added to quench the digestion, and the digestion solution was strained over a 70-micron filter. The cell suspension was spun down at 4C and 1400 rpm for 7 min. Red blood cell lysis was performed using the Invitrogen RBC lysis buffer. The resulting cell pellet was resuspended in PBS supplemented with 0.04% BSA before being loaded onto the 10X Genomics Chromium Single Cell Controller. Cell viability analysis was performed before loading the samples, and was above 95% per sample. A total of ~8000 cells were analyzed per sample. FASTQ file generation, alignment, filtering, barcode counting, and UMI counting were done using the 10X Cell Ranger software.

first: Kira

last: Tomlinson


title: Heat shock response and transposon control in plant shoot stem cells [Bisulfite-seq]

overall_design: Six-day old wt, poliv and, ddm1 plants were exposed to 37 celsius degree for 1 day (D7), then plants were transferred to soil to grow for 21 more days (D28). The plants stem cell nuclei at D28 were collected for bisulfite-seq.


title: Heat shock response and transposon control in plant shoot stem cells

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


title: Nanopore sequencing unveils the complexity of the murine brown adipose tissue transcriptome: Illumina RNA-Seq of eWAT from cold treated mice [05_eWAT_illumina]

status: Public on Sep 18 2023

first: Jan-Wilhelm

last: Kornfeld

department: BMB

institute: University of Southern Denmark

summary: Alternative splicing contributes to transcriptomic complexity and plays a role in the regulation of cellular identity and function, but the correct assembly of transcripts of complex loci as well as their quantification based on short-read sequencing is non-trivial. Recent long-read sequencing methods such as those from ONT and PacBio overcome these problems by potentially sequencing full transcripts. The activation of brown adipose tissue e.g., by reduced ambient temperature (cold) exposure, positively affects metabolism by increasing energy expenditure and releasing endocrine factors and has been shown to involve specific alternative splicing events. Here we assessed important features of ONT long read sequencing protocols in relation to Illumina short read sequencing: (i) Alignment characteristics to the reference genome and transcriptome, (ii) Gene and transcript detection and quantification, (iii) Detection of differential gene and transcript expression events, (iv) Transcriptome reannotation and (v) Detection of differential transcript usage events. We find that ONT long-read sequencing is advantageous in terms of transcriptome reassembly, especially when the reads are enriched for full length reads. Illumina sequencing, due to the higher number of counts available, has a higher statistical power for calling differentiall expressed/used features, whereas long-read sequencing has a lower risk of calling false positive events due to the better ability to unambiguously map reads to transcripts. Finally we describe novel transcript isoforms in cold-activated murine iBAT reassembled from ONT long reads.

overall_design: PolyA RNA from epididymal white adipose tissue of 20 weeks old male wild type C57BL/6N mice singly housed at 4°C (n = 3) for a period of 24 h sequenced by Illumina short-read sequencing.

first: Christoph

last: Engelhard

first: Sajjad

last: Khani

first: Sophia

last: Derdak

first: Martin

last: Bilban


title: mtran-Transcriptome analysis of mutants in uncharacterised mitochondrial proteins

status: Public on Sep 03 2023

city: Gif sur Yvette

first: Véronique

last: BRUNAUD

address: avenue des sciences

country: France

institute: INRAE

summary: ngs2021_04_mac-mtran-Assessment of the impact of loss-of-function of poorly characterised mitochondrial proteins on the nuclear and organellar transcriptome - Transcriptome analysis of mutants in uncharacterised mitochondrial proteins

overall_design: 6 samples with 2 biological replicats. gene knock out- double mutant AT4G15640-AT3G21465

first: Miguel

last: Botella

first: Alexandra

last: Launay-Avon

first: Olivier

last: Van aken

first: Etienne

last: Delannoy


title: Nanopore sequencing unveils the complexity of the murine brown adipose tissue transcriptome: ONT PCR RNA-Seq (TeloPrime) of iBAT from cold treated mice [03_iBAT_teloprime]

overall_design: PolyA RNA from interscapular brown adipose tissue of 20 weeks old male wild type C57BL/6N mice singly housed at 4°C (n = 3) for a period of 24 h sequenced by Oxford Nanopore Technologies  sequencing after enrichment for full length transcript using the Full-Length cDNA Amplification kit (Lexogen).


title: Nanopore sequencing unveils the complexity of the murine brown adipose tissue transcriptome: ONT direct RNA-Seq of iBAT from cold treated mice [02_iBAT_rna]

overall_design: Pooled polyA RNA from interscapular brown adipose tissue of 20 weeks old male wild type C57BL/6N mice singly housed at 4°C (n = 3) for a period of 24 h sequenced by Oxford Nanopore Technologies direct RNA sequencing.


title: Nanopore sequencing unveils the complexity of the murine brown adipose tissue transcriptome: Illumina RNA-Seq of iBAT from cold treated mice [01_iBAT_illumina]

overall_design: PolyA RNA from interscapular brown adipose tissue of 20 weeks old male wild type C57BL/6N mice singly housed at 4°C (n = 3) for a period of 24 h sequenced by Illumina short-read sequencing.


title: Nanopore sequencing unveils the complexity of the murine brown adipose tissue transcriptome.


title: Multi-Lineage Heart-Chip Models Drug Cardiotoxicity and Enhances Maturation of Human Stem Cell-Derived Cardiovascular Cells

status: Public on Sep 01 2023

city: Los Angeles,

first: Arun

last: Sharma

state: California

address: 127 S San Vicente Blvd,

department: Regenerative Medicine Institute

institute: Cedars Sinai Medical Center

summary: Cardiovascular toxicity causes adverse drug reactions and may lead to drug removal from the pharmaceutical market. Cancer therapies can induce life-threatening cardiovascular side effects such as arrhythmias, muscle cell death, or vascular dysfunction. New technologies have enabled cardiotoxic compounds to be identified earlier in drug development. Human induced pluripotent stem cell (hiPSC)-derived cardiomyocytes (CMs) and vascular endothelial cells (ECs) can screen for drug-induced alterations in cardiovascular cell function and survival. However, most existing hiPSC models for cardiovascular drug toxicity utilize two-dimensional, immature cells grown in static culture. Improved in vitro models to mechanistically interrogate cardiotoxicity would utilize more adult-like, mature hiPSC-derived cells in an integrated system whereby toxic drugs and protective agents can flow between hiPSC-ECs that represent systemic vasculature and hiPSC-CMs that represent heart muscle (myocardium). Such models would be useful for testing the multi-lineage cardiotoxicities of chemotherapeutic drugs such as VEGFR2/PDGFR-inhibiting tyrosine kinase inhibitors (VPTKIs). Here, we develop a multi-lineage, fully-integrated, cardiovascular organ-chip that can enhance hiPSC-EC and hiPSC-CM functional and genetic maturity, model endothelial barrier permeability, and demonstrate long-term functional stability.

overall_design:
We've used the 02iCTR iPSC and gCaMP iPSC,  and differentiated them into cardiomyocytes and endothelial cells, both used in a single channel of the Emulate chips. We've subjected both cell lines to flow and a cyclic mechanical stretch and the resultant cells are assessed for their gene distribution against static cultures. We've further tested the effect of a TKI drug Sorfenib on both cell types through the platform.
The cells are human-derived iPSCs. We've used 4 replicates of iPSC-ECs and iPSC-CMs for RNA sequencing of the static conditions. 3 smaples of CMs under stretch and flow and 3 samples under flow only were tested for RNA-Seq as well. Finally we have tested 3 replicates of EC under flow and stretch and 2 replicate of ECs under flow only conditions, for RNA-Seq.

first: Maedeh

last: Mozneb

first: Amelia

last: Jenkins

first: Stephany

last: Pohlman


title: Single-Cell Transcriptomic Profiling of Heart Reveals ANGPTL4 Linking Fibroblasts and Angiogenesis inHeart Failure with Preserved Ejection Fraction [bulk RNA-seq]

city: Chongqing

first: Li

last: Guoxing

state: Chongqing

address: Street of shiyou Road, Yuzhong District, Chongqing, China

institute: Chongqing Medical University

summary: Heart failure with preserved ejection fraction (HFpEF) is a complex, heterogeneous disease involving multiple cellular lineages and mechanisms. Here, we aimed to characterize the cellular heterogeneity and potential mechanisms of HFpEF at single-cell resolution.

overall_design:
Eight-week-old male C57BL/6N mice were randomly divided into two groups: an HFpEF group administered with a high-fat diet and N-nitro-L-arginine methyl ester (L-NAME, 0.5 g/L in drinking water) and a control group fed on normal diet and water. Both groups were exposed to a 12-h light/dark cycle daily. After 5 weeks, cardiac ultrasound was conducted to assess the mice's ejection fraction and diastolic function. Then, mice were euthanized by excess inhalation of CO2, followed by the isolation of the hearts and the removal of atrial tissue. Three and five ventricular samples for single-cell sequencing and bulk RNA-seq. respectively, were then obtained from each group.
IMPORTANT: Please note that there was an inconsistency when referring to the control group, i.e., control group vs. CON. To avoid confusion, I have used “control group” consistently throughout the manuscript.

first: Guoxing

last: Li

first: Huilin

last: Zhao

first: Junjin

last: Liu

first: Zhe

last: Cheng

first: Yuhong

last: Qin

first: Gang

first: yongzheng

last: Guo


title: LocusMasterTE: long-read assisted short-read TE quantification [short]

city: Hong Kong

first: Sojung

last: LEE

address: 21 Sassoon Rd, Pok Fu Lam

department: Faculty of Medicine

country: Hong Kong

institute: The Hong Kong University

summary: With an ability to compromise genome integrity, transposable elements (TEs) have significant associations with human diseases. Short-read sequencing has been used to study the expression of TEs; however, the highly repetitive nature of these elements makes multimapping a critical issue. Here we implement LocusMasterTE, an improved quantification method by integrating long-read sequencing. Introducing computed transcript per million(TPM) counts from long-read sequencing as prior distribution during Expectation-Maximization(EM) model in short-read TE quantification, multi-mapped reads are re-assigned to correct expression values. Based on simulated short reads, LocusMasterTE outperforms current quantitative approaches and is significantly favorable in capturing newly inserted TEs. We also verified that TEs quantified by LocusMasterTE clearly related to euchromatins and heterochromatins in cell line samples. With LocusMasterTE we anticipate that more accurate quantification can be performed, allowing novel functions of TEs to be uncovered.

overall_design: Short read RNA-seq of HCT116 cell line

last: Lee

first: Jayne

last: Barbour

first: Jason

middle: W

last: Wong


title: Single-Cell Transcriptomic Profiling of Heart Reveals ANGPTL4 Linking Fibroblasts and Angiogenesis inHeart Failure with Preserved Ejection Fraction

overall_design: Refer to individual Series


title: LocusMasterTE: long-read assisted short-read RNA-seq TE quantification [long]

overall_design: Oxford nanopore PCR cDNA sequencing of HCT116 cell line


title: Circulating CD3+ CD14+ T Cell-Monocyte complexes are dynamic, metabolically glucose-dependent HIV reservoirs and increased in individuals with glucose intolerance

city: Nashville

first: Samuel

last: Bailin

address: 1161 21st Avenue South, MCN A-2200

institute: Vanderbilt Universit Medical Center

summary: Persistent and elevated systemic inflammation contributes to the increased risk of cardiovascular and metabolic diseases in persons with HIV (PWH). , but aspects of the innate and adaptive immune response to HIV and other chronic co-infections that promote the development of these comorbid conditions remain unclear. We used mass cytometry and the tracking responders expanding (T-REX) workflow to define differences in circulating cells of the innate and adaptive immune arms, which differentiate non-diabetic and prediabetic/diabetic persons with HIV (PWH) on long-term antiretroviral therapy (ART). We found a significantly higher proportion of circulating CD14+ monocytes complexed with T cells, B cells, and NK cells among PWH with diabetes. The CD3+ T cells in these complexes were predominantly CD8+ memory T cells. The CD3+ CD14+ T cell-monocyte complexes are pro-inflammatory and negatively associated with plasma interleukin-10 levels and circulating CD4+ T regulatory cells. Using transmission electron microscopy, we observed heterogeneous interactions between CD3+ T cells and monocytes within the complexes, including formation of immune synapses and phagocytosis. 10x Chromium single-cell sequencing RNA transcriptomic analysis of CD3+ CD14+ doublets showed differential expression of genes involved in T cell activation and the adaptive immune response compared to monocytes that were not complexed with other immune cells. Notably, there were significantly more copies of HIV RNA  per cell in the CD3+ CD14+ T cell-monocyte complexes compared to CD14+ monocytes or CD3+ CD4+ T cells alone, and HIV virions were observed in both T cells and monocytes within the doublets?. Metabolically, CD3+ CD14+ T cell-monocyte complexes had high glucose-dependence with minimal mitochondrial dependence. Taken together, CD3+ CD14+ T cell-monocyte pairs are functional and physically interacting immune cell complexes which might be enriched with HIV. These complexesare increased among individuals with diabetes and may be a target for future HIV cure studies and diseases of aging.

overall_design: Circulating peripheral blood mononuclear cells were isolated and analyzed using scRNAseq.

first: Wanjalla

last: Celestine

first: Simmons

last: Joshua

first: Oakes

last: Jared

first: Bailin

last: Samuel

first: Nochowicz

last: Cindy

first: Warren

last: Christopher

first: Beasley

last: Heather

first: Wang

last: Jian

first: Sheng

last: Quanhu

first: Stolze

last: Joey

first: Gangula

last: Rama

first: Mashayekhi

last: Mona

first: Priest

last: Stephen

first: Zhang

last: Xiuqi

first: Chopra

last: Abha

first: Gabriel

last: Curtis

first: Temu

last: Tecla

first: Pakala

last: Suman

first: Wilfong

last: Erin

first: Gianella

last: Sara

first: Phillips

last: Elizabeth

first: Harrison

last: David

first: Hinton

last: Antentor

first: Kalams

last: Spyros

first: Mallal

last: Simon

first: Koethe

last: John


title: Systematic mapping of 3D enhancer interactions reveals lineage specific regulatory networks during early mouse developmental decisions [ChIP-Exo]

status: Public on Aug 21 2023

first: Effie

last: Apostolou

address: 1161 York Avenue, Apt 8A

institute: Weill Cornell Medicine

summary:
To model and characterize the chromatin regulatory landscape of the very first developmental cell fates, we made use of three previously derived and well-characterized Trophoblast Stem Cell (TSCs), Embryonic Stem Cell (ESC) and Extra Embryonic Stem Cell lines (XENs)
We performed ChIP-Exo for CTCF to map the insulator regions  in ESC, TSC and XEN

overall_design: Characterization of binding sites of CTCF with ChiP-Exo for all three cell types.

first: Prokopios

last: Polyzos


title: Systematic mapping of 3D enhancer interactions reveals lineage specific regulatory networks during early mouse developmental decisions [HiChIP]

summary:
To model and characterize the chromatin regulatory landscape of the very first developmental cell fates, we made use of three previously derived and well-characterized Trophoblast Stem Cell (TSCs), Embryonic Stem Cell (ESC) and Extra Embryonic Stem Cell lines (XENs)
We performed HiChIP to profile putative enhancer interactions in TSC, ESC, XEN and EPISC cells

overall_design: Identification of Enhancer interactions and enhancer rewiring between different developmental stages


title: Systematic mapping of 3D enhancer interactions reveals lineage specific regulatory networks during early mouse developmental decisions


title: Single-Cell Transcriptomic Profiling of Heart Reveals ANGPTL4 Linking Fibroblasts and Angiogenesis inHeart Failure with Preserved Ejection Fraction [scRNA-seq]

first: Li

last: Guoxing

first: Guoxing

last: Li

first: Huilin

last: Zhao

first: Junjin

last: Liu

first: Zhe

last: Cheng

first: Yuhong

last: Qin

first: Gang

first: Yongzheng

last: Guo


title: Systematic mapping of 3D enhancer interactions reveals lineage specific regulatory networks during early mouse developmental decisions [Hi-C]

summary:
To model and characterize the chromatin regulatory landscape of the very first developmental cell fates, we made use of three previously derived and well-characterized Trophoblast Stem Cell (TSCs), Embryonic Stem Cell (ESC) and Extra Embryonic Stem Cell lines (XENs)
We performed HiC to map large-scale 3D architectural rewiring  in ESC, TSC and XEN

overall_design: Characterization of Connectivity, TAD and Compartments shifts in all cell types.


title: Systematic mapping of 3D enhancer interactions reveals lineage specific regulatory networks during early mouse developmental decisions [ChIP-Seq]

summary:
To model and characterize the chromatin regulatory landscape of the very first developmental cell fates, we made use of three previously derived and well-characterized Trophoblast Stem Cell (TSCs), Embryonic Stem Cell (ESC) and Extra Embryonic Stem Cell lines (XENs)
We performed ChIP-seq for H3k27ac to map the putative active enhancers and promoters  in ESC, TSC, XEN and EPISC

overall_design: Characterization of active enhancers and promoters with H3k27ac ChiP-seq for all cell types.


title: Reconstruction of Dynamic Mammary Mini Gland in vitro for Normal Physiology and Oncogenesis

status: Public on Aug 20 2023

city: HangZhou

first: Xiaoqin

state: ZheJiang

address: Shilongshan Road No.18, Cloud Town, Xihu District

institute: Westlake University

summary: Organoid culture has been extensively exploited for normal tissue reconstruction and disease modeling, however, it is still challenging to establish physiologically relevant architecture, size and functions in homeostasis. Here, we describe the development of a long-term adult stem cell derived mammary mini-gland culture system that supports robust 3D outgrowths recapitulating the morphology, cellular context, transcriptional heterogeneity of the normal mammary gland. The self-organization ability of stem cells and the stability of the outgrowths were determined by a coordinated combination of extracellular matrix, environmental signalings and dynamic physiological cycles. These mini glands are hormone responsive and reproduce the entire postnatal mammary development including puberty, estrous cycle, lactation and involution. Most intriguingly, these mini glands maintained the mammary stem cells and represented the fate transition from embryonic bipotency to postnatal unipotency in lineage tracing assays. In addition, induced oncogene expression in the mini glands drove the clinically relevant cancer initiation that can be monitored chronologically. Together, this study provides an experimental system fostering a dynamic organ miniature that can be studied chronologically and spatially for physiologically relevant biological processes with complexity.

overall_design: Single cell RNA profiles of mammary gland basal cells and luminal cells from primary mammary gland and mammary mini gland

first: Lei

last: Yuan

first: Shaofang

last: Xie

first: Huiru

last: Bai

first: Pei

last: Cai

first: Jing

last: Lu

first: Chunhui

last: Wang

first: Zuobao

last: Lin

first: Shuying

first: Yajing

first: Shang


title: Systematic mapping of 3D enhancer interactions reveals lineage specific regulatory networks during early mouse developmental decisions [RNA-Seq]

first: Effie

summary:
To model and characterize the chromatin regulatory landscape of the very first developmental cell fates, we made use of three previously derived and well-characterized Trophoblast Stem Cell (TSCs), Embryonic Stem Cell (ESC) and Extra Embryonic Stem Cell lines (XENs)
We performed gene expression profiling in ESC, TSC, XEN and EPISC with RNA-seq technique.

overall_design: Characterization of each cell line by RNA-seq analysis and differential analysis among them to identify gene signatures

first: Prokopios


title: Nanopore sequencing unveils the complexity of the murine brown adipose tissue transcriptome: ONT direct cDNA-Seq of iBAT from cold treated mice [04_iBAT_cdna]

city: Odense

first: Jan-Wilhelm

last: Kornfeld

address: Campusvej 55

overall_design: PolyA RNA from interscapular brown adipose tissue of 20 weeks old male wild type C57BL/6N mice singly housed at 4°C (n = 3) for a period of 24 h sequenced by Oxford Nanopore Technologies direct cDNA sequencing.

first: Christoph

last: Engelhard

first: Sajjad

last: Khani

first: Sophia

last: Derdak

first: Martin

last: Bilban


title: Redox-sensitive High-mobility group box-1 isoforms contribute to liver fibrosis progression and resolution in mice

first: Xiaodong

last: Ge

state: IL

address: 909 South Wolcott Ave, COMRB 6140

department: Pathology

institute: The University of Illinois at Chicago

summary: To study role of different HMGB1 isoforms in liver fibrosis and resolution

sample_organism:
Mus musculus
Rattus norvegicus

platform_organism:
Mus musculus
Rattus norvegicus

overall_design: RNA sequencing was performed in total liver at peak fibrosis and resoluand, rHSC was treated with different HMGB1 isoforms for 6 hours

first: Natalia

last: Nieto


title: RNA sequencing of healthy human airway epithelia with IL-13 treatment time course

status: Public on Aug 16 2023

city: Iowa City

first: Alejandro

middle: A.

last: Pezzulo

state: Iowa

address: 6316 Pappajohn Biomedical Discovery Building

department: Department of Internal Medicine

institute: Universty of Iowa

summary: The goal of this study was to examine changes to gene expression induced by IL-13 treatment of air-liquid interface cultures of healthy primary human airway epithelia over time.

overall_design: Following treatment with DMSO or IL-13 for 2 days, 7 days, or 21 days, well-differentiated, primary human airway epithelia at the air liquid interface were dissociated, prepared for RNA sequencing, and sequenced on Illumina HiSeq 4000 system using 2x150bp paired end SBS chemistry.

first: Andrew

last: Thurman


title: Single cell multi-omics analysis deconstructs diabetes-associated β-cell heterogeneity in human islets(scRNASeq)

status: Public on Aug 09 2023

city: Cleveland

first: Shanshan

last: Zhang

address: 10900 Euclid Avenue

department: Department of Genome and Genetics

institute: Case Western Reserve University

summary: The functional heterogeneity of β-cells is important for metabolism and diseases, but the nature and mechanism of such variation at molecular level remain elusive. Here we explore this question by comparing both single cell RNA-seq and single cell ATAC-seq maps of healthy and type II diabetic (T2D) human islets. We dissect the T2D-associated single cell trajectory and identified signature genes and enhancers in β-cells. We also map the 3D genome of both α- and β-cells with low-input eHi-C approach to unravel the cell type-specific gene regulatory circuits. Strikingly, more than one third of the T2D signature genes show significant intra-donor heterogeneity at single cell level; these genes are functionally distinct from other signature genes that are largely invariant among cells from the same individual but differentially expressed between donors, suggesting different roles in T2D pathogenesis. Importantly, we identified consistent intra-donor variations at both transcriptomic and epigenomic levels, which strongly support the heterogenous β-cell states and transcription programs. Finally, we construct the disease-signature regulatory networks and pinpointed HNF1A as one of the top transcription factors governing T2D-associated β-cell heterogeneity. Taken together, we provide the first multi-omic characterization of β-cell heterogeneity and reveals its connection to T2D.

overall_design: Single cell sequencing (Drop-seq) for Human Pancreatic islet from 11 individuals respectively. Including 7 Healthy donor and 4 Type II diabetes patient donor.

first: Chen

last: Weng

first: Anniya

last: Gu

first: Fulai

last: Jin

first: Yan

last: Li


title: Knockout of Hsp70 genes significantly affects locomotion speed and gene expression in leg skeletal muscles of Drosophila melanogaster

status: Public on Aug 01 2023

city: Moscow

first: Pavel

last: Makhnovsky

address: 76A Khoroshevskoye shosse

country: Russia

institute: Institute of Biomedical Problems of the Russian Academy of Sciences

summary: The functions of the Hsp70 genes were studied using a line of D. melanogaster with knockout of six these genes out of thirteen. Namely, effect of knockout of Hsp70 genes on negative geotaxis climbing (locomotor) speed and the ability to adapt to climbing training (0.5-1.5 h/day, 7 days/week, 19 days) were examined. Seven- and 23-day-old Hsp70– flies demonstrated a comparable reduction (2-fold) in locomotor speed and widespread changes in leg skeletal muscle transcriptome (RNA-seq), compared to w1118 flies. To identify the functions of genes related to decreased locomotor speed the overlapped differentially expressed genes at both time points were analyzed: the up-regulated genes encoded extracellular proteins, regulators of drug metabolism and antioxidant response, while down-regulated genes encoded regulators of carbohydrate metabolism and transmembrane proteins. Additionally, in Hsp70– flies, activation of transcription factors related to disruption of the fibril structure and heat shock response (Hsf) were predicted, using the position weight matrix approach. In the control flies, adaptation to chronic exercise training was associated mainly with gene response to a single exercise bout, while the predicted transcription factors were related to stress/immune (Hsf, NF-kB, etc.) and early gene response. In contrast, Hsp70– flies demonstrated no adaptation to training, as well as significantly impaired gene response to a single exercise bout. In conclusion, the knockout of Hsp70 genes not only reduced physical performance, but also disrupted adaptation to chronic physical training, which is associated with changes in leg skeletal muscle transcriptome and impaired gene response to a single exercise bout.

overall_design:
The w1118 males (Vienna Drosophila Resource Center, ID 60000) and Hsp70– males (genotype w1118; Df(3R)Hsp70A, Df(3R)Hsp70B, Bloomington Drosophila Stock Center, ID 8841) with a knockout of 6 genes (Hsp70Aa, Hsp70Ab, Hsp70Ba, Hsp70Bb, Hsp70Bbb, Hsp70Bc) were used in this study.
1) Flies (both strains) were anesthetized with diethyl ether on the 7th, 23rd, and 47th days of life (after eclosion), legs were removed in phosphate-buffered saline and then placed in RNA extraction buffer for RNA isolation and RNA-sequencing (n = 4 pools (15 flies per pool) for each strain).
2) Flies (both strains) were trained from the 4th to the 22nd day after eclosion (19 days) using a homemade device, which rotated the tubes around the transverse axis every 15 s. Each training session began at 12:00; the training duration was 30 min on the first 2 days, 60 min on the second 2 days, and 90 min on the remaining days. Flies were anesthetized 24 h after the last training session; legs were removed for RNA isolation and RNA-sequencing (n = 4 pools (15 flies per pool) for each strain).
3) Untrained flies (7 days old, both strains) exercised for 2 h. Immediately thereafter, exercised and non-exercised flies were anesthetized and legs were removed for RNA isolation and RNA-sequencing (n = 4 pools (15 flies per pool) for each strain).

https://link.springer.com/article/10.1134/S0026893324020109

first: Daniil

last: Popov

first: Popov

last: Makhnovskii


title: Single cell profile of human iPSC-derived midbrain organoids from healthy individual, Parkinson disease patient harboring a Miro1R272Q mutation and its respective isogenic control

status: Public on Jul 31 2023

city: Belvaux

first: Claudia

last: Saraiva

state: Luxembourg

address: 6, avenue du Swing

department: Luxembourg Centre for Systems Biomedicine

country: Luxembourg

institute: University of Luxembourg

summary: Midbrain organoids are 3D in vitro models, which represent the human midbrain and can be used as Parkinson's disease (PD) models. In the current study, we used  single cell RNA sequencing (scRNA-seq) to investigate the effect of a novel mutation in the RhoT1 gene (Miro1 R272Q) in the PD pathology, and how it affect the dopaminergic neuron population present in the model.

overall_design: iPSC-derived midbrain organoids generated from a PD patient harboring the Miro1 R272Q mutation, its respective isogenic control and from a healthy donor were generated and kept in culture for 30 days. Organoids were then dissociated into single cells and used for scRNA-seq analysis.

first: Cláudia

first: Jens

middle: C

last: Schwamborn

first: Kyriaki

last: Barmpa

first: Alise

last: Zagare


title: Single cell multi-omics analysis deconstructs diabetes-associated β-cell heterogeneity in human islets (Hi-C)

overall_design: 3D genome structure mapping in sorted alpha and beta cells from human islets using easy Hi-C method


title: Understanding environmental and biocide survival of Candida auris through transcriptional profiling

status: Public on Aug 07 2023

city: Glasgow

first: Alicia

middle: Ann

last: Ware

address: Cowcaddens Rd

department: Biological & Biomedical Sciences

institute: Glasgow Caledonian University

summary: Candida auris is an enigmatic fungal pathogen, recently elevated to the Critical Priority group of fungal pathogens by the World Health Organisation. Of key concern is its ability to cause outbreaks within intensive and chronic care units, which is facilitated through biofilm formation. We investigated the susceptibility of phenotypically-diverse C. auris isolates to sodium hypochlorite (NaOCl) disinfection, and the subsequent role of biofilms in surviving disinfection using a dry-surface biofilm (DSB) model and transcriptomic profiling. Planktonic cells were tested for susceptibility to NaOCl using minimum inhibitory concentration and disinfection protocols. Biofilm formation using the DSB model consisted of consecutive 48 hr cycles with/without media across a 12-day period, assessed using viable counts, biomass assays, and microscopy. Disinfection efficacy was assessed using clinically relevant protocols of 500-1000 ppm for 1-5 min. RNA-sequencing was performed on untreated DSBs in comparison to planktonic cells. Isolates were found to be sensitive to NaOCl planktonically at concentrations ≤62.5 ppm, and grew robust biofilms using the DSB protocol. Biofilms developed tolerance to all NaOCl treatment parameters, with 2-4 log10-reductions in viable cells observed at highest concentrations. Transcriptomics identified ABC transporters and iron acquisition pathways as strongly upregulated in DSBs relative to planktonic cells. We have optimised a DSB protocol in which C. auris biofilms can mediate tolerance to adverse conditions such as NaOCl disinfection, suggesting a lifestyle through which this problematic yeast can environmentally persist and transmit. Mechanistically, the upregulation of small-molecule and iron transport pathways have been identified as potential facilitators of survival.

overall_design:
To investigate mechanisms behind C. auris survival and persistence in the healthcare environment, we grew in vitro dry surface biofilms (DSBs) using C. auris NCPF8973 (clade I, single-celled) and NCPF8978 (clade III, aggregating) and harvested RNA after 12 days of growth.
Transcriptional profiling using RNA-seq data of DSB cells and planktonic cells of the 2 isolates.
Differential gene expression was determined with DSB cells as the experimental group, and planktonic cells the control group.

first: Ryan

last: Kean

first: Christopher

last: Delaney


title: Cell-specific and shared regulatory elements control a multi-gene locus active in mammary and salivary glands

status: Public on Jul 28 2023

city: Bethesda

first: Hye Kyung

last: Lee

state: MD

address: 8 CENTER DR RM 107

department: National Institute of Diabetes and Digestive and Kidney (NIDDK)

institute: National Institutes of Health (NIH)

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

overall_design: Refer to individual Series


title: Cell-specific and shared regulatory elements control a multi-gene locus active in mammary and salivary glands [RNA-Seq]

summary: Regulation of high-density loci harboring genes with different cell-specificities remains a puzzle. Here we investigate a locus that evolved through gene duplication and contains eight genes and 20 candidate regulatory elements, including one super-enhancer. Five genes (Csn1s1, Csn2, Csn1s2a, Csn1s2b, Csn3) are expressed in mammary glands and account for 50% of mRNAs during lactation, two (Prr27, Fdcsp) are salivary-specific and one (Odam) has dual specificity. We probed the function of 12 candidate control elements, individually and in combination, in the mouse genome. The super-enhancer is essential for the expression of Csn3, Odam and Fdcsp but largely dispensable for Csn1s1, Csn2 and Csn1s2a. Csn3 activation also requires its own local enhancer. Synergism between local enhancers and cytokine-responsive promoter elements facilitated the 10,000-fold activation of Csn2 during pregnancy. Our work identifies regulatory complexity of a multigene locus with an ancestral super-enhancer active in mammary and salivary tissue and local enhancers and promoter elements.

overall_design: Total RNA-seq was done for the mammary tissues at virgin, day one and ten of lactation (L1, L10) and the salivary tissues of male mice at 2-month-old age for wild-type and mutant mice carrying a deletion of regulatory elements in the casein locus.

first: Hye

middle: K


title: Genome-scale chromatin interaction dynamic measurements for key components of the RNA Pol II general transcription machinery [RNA-seq]

status: Public on Jul 27 2023

city: Charlottesville

first: David

last: Auble

state: VA

address: 1349 Jefferson Park Avenue

department: Biochemistry and Molecular Genetics

institute: University of Virginia Health System

summary: A great deal of work has revealed in structural detail the components of the machinery responsible for mRNA gene transcription initiation. These include the general transcription factors (GTFs) which assemble at promoters along with RNA Polymerase II (Pol II) to form a preinitiation complex (PIC) aided by the activities of cofactors and site-specific transcription factors (TFs). However, less well understood are the in vivo PIC assembly pathways and their kinetics, an understanding of which is vital for determining on a mechanistic level how rates of in vivo RNA synthesis are established and how cofactors and TFs impact them. We used competition ChIP to obtain genome-scale estimates of the residence times for five GTFs: TBP, TFIIA, TFIIB, TFIIE and TFIIF in budding yeast. While many GTF-chromatin interactions were short-lived (< 1 min), there were numerous interactions with residence times in the several minutes range. Sets of genes with shared function also shared similar patterns of GTF kinetic behavior. TFIIE, a GTF that enters the PIC late in the assembly process, had residence times correlated with RNA synthesis rates. The datasets and results reported here provide kinetic information for most of the Pol II-driven genes in this organism and therefore offer a rich resource for exploring the mechanistic relationships between PIC assembly, gene regulation, and transcription. The relationships between gene function and GTF dynamics suggest that shared sets of TFs tune PIC assembly kinetics to ensure appropriate levels of expression. Scripts and processed fileas are available from: https://github.com/AubleLab/PIC_competition_ChIP_scripts

sample_organism:
Schizosaccharomyces pombe
Saccharomyces cerevisiae

overall_design: To establish RNA synthesis rates, we measured newly synthesized RNA using dynamic transcriptome analysis (DTA) method. We obtained replicated samples (n=2) 60 and 20 min post galactose induction. Thiouracil (4sU) was added to label nascent RNA. Negative control samples do not have 4sU added. Collected were following fractions: labeled RNA (L), unlabeled RNA (U) and total RNA (T).

first: Kristyna

last: Kupkova

first: Savera

last: Shetty

first: Stefan

last: Bekiranov

middle: T


title: Genome-scale chromatin interaction dynamic measurements for key components of the RNA Pol II general transcription machinery

platform_organism:
Schizosaccharomyces pombe
Saccharomyces cerevisiae


title: Systematic mapping of 3D enhancer interactions reveals lineage specific regulatory networks during early mouse developmental decisions [ATAC-Seq]

first: Effie

last: Apostolou

summary:
To model and characterize the chromatin regulatory landscape of the very first developmental cell fates, we made use of three previously derived and well-characterized Trophoblast Stem Cell (TSCs), Embryonic Stem Cell (ESC) and Extra Embryonic Stem Cell lines (XENs)
We performed ATAC to map accessible regions (AR)  in enhancer and promoter regions  in ESC, TSC, XEN and EPISC

overall_design: Characterization of AR with ATAC-seq for all cell types.

first: Prokopios

middle: A

last: Polyzos


title: Cell-specific and shared regulatory elements control a multi-gene locus active in mammary and salivary glands [ChIP-Seq]

overall_design: ChIP-seq for TFs and histone markers in the mammary tissues of wild type and mutant female mice for regulatory elements in the casein locus at virgin, day six of pregnancy (p6) and day one of lactation (L1). ChIP-seq for TFs and histone markers in the salivary tissues of wild type and mutant male mice.


title: Cell-specific and shared regulatory elements control a multi-gene locus active in mammary and salivary glands [Bisulfite-seq]

overall_design: Bisulfite-seq in WT lactating mammary tissues and liver tissues


title: Systematic mapping of 3D enhancer interactions reveals lineage specific regulatory networks during early mouse developmental decisions [4C-Seq]

summary:
To model and characterize the chromatin regulatory landscape of the very first developmental cell fates, we made use of three previously derived and well-characterized Trophoblast Stem Cell (TSCs), Embryonic Stem Cell (ESC) and Extra Embryonic Stem Cell lines (XENs)
We performed high-resolution in situ 4C-seq analysis around enhancers and promoters of cell-type specific genes (e.g. Sox17 for XEN and Nanog for ESC), which showed high concordance both with the virtual 4C of HiChIP and the called HiChIP contacts in the respective cell type

overall_design: 4C-seq in duplicates / triplicates was performed for selected genes in ESC, TSC and XEN


title: differential expression analysis to hight co2 of the brachypodium distachyon-Differential expression of project NGS20_04_HIPATH

first: Véronique

last: BRUNAUD

summary: ngs2020_04_hipath-differential expression analysis to hight co2 of the brachypodium distachyon-Analysis response of the Brachypodium distachyon to hight CO2 -treatment hight CO2

overall_design: 9 samples with 0 biological replicats. dose response,treated vs untreated comparison

first: Alexandra

last: Avon-Launay

first: Marie

last: Dufresne

first: Etienne

last: Delannoy

first: Ludivine

last: Taconnat-Soubigou


title: Differences in DNA methylation of HAMP in blood cells predicts the development of type 2 diabetes [WGBS]

status: Public on Jul 17 2023

city: Nuthetal

first: Heike

last: Vogel

state: BRB

address: Arthur-Scheunert-Allee 114-116

department: Department of Experimental Diabetology

institute: German Institute of Human Nutrition Potsdam-Rehbruecke

summary: Diabetes-resistant and diabetes-prone female New Zealand Obese mice were classified based on liver fat content and early blood glucose concentrations at 10 weeks of age before the onset of T2D. By using transcriptome and DNA methylome analysis of liver samples, we identified early epigenetic alteration in mice and humans which could serve as putative epigenetic biomarkers

overall_design: DNA methylation sequencing was performed with 6 diabetes resistant (DR) and 6 diabetes prone (DP) animals, DR animals are healthy animals and are considered as control group

first: Annette

last: Schürmann

first: Meriem

last: Ouni

first: Markus

last: Jähnert

first: Sophie

last: Saussenthaler

first: Pascal

last: Gottmann

first: Peter

last: Huypens

first: Johannes

last: Beckers


title: Differences in DNA methylation of HAMP in blood cells predicts the development of type 2 diabetes [RNA-seq]

overall_design: RNA-seq was performed with 5 diabetes resistant (DR) and 6 diabetes prone (DP) animals, DR animals are healthy animals and are considered as control group


title: Differences in DNA methylation of HAMP in blood cells predicts the development of type 2 diabetes.


title: Eurycomalactone switches hepatocellular carcinoma cells into a quiescence state through 5'tRFAla&5'tiRNAAla/DVL/β-catenin pathway inhibition

status: Public on Jul 10 2023

city: Zhanjiang

first: z

middle: z

last: p

state: Guangdong

address: wenmingdonglu

department: school of pharmacy

institute: GuangDongMedicialUniversity

summary: Although tsRNA was reported to have the ability to modulate a variety of physiological processes analogous to miRNA, the possible regulatory functions and mechanisms of tsRNAs related to the pharmacological effects of small molecules are still obscure. Herein it is shown that the natural product eurycomalactone (ELT) can reversibly switch hepatocellular carcinoma (HCC) PLC/PEF/5 and HUH7 cells into a quiescence state characterized by cell proliferation inhibition without cell death, cell cycle arrest at G0/G1 phase and cell reactivation after drug withdrawal. On the basis that β-catenin activity might mediate the proliferation or quiescence of cancer cells, the total, cytoplasmic and nuclear β-catenin, as well as its target proteins, c-myc, CD44 and cyclin D1, are all shown to be significantly weakened by ELT. Subsequently, two new tsRNAs, namely 5'tRFAla and 5'tiRNAAla, which match well with the mRNAs of Dishevelled proteins DVL2 and DVL3, two vital upstream regulators of β-catenin, are found to be decreased significantly in ELT-treated PLC/PEF/5 cells. The concentrations of the DVL2 and DVL3 proteins are similarly reduced. Accordingly, it is suggested that ELT decreases 5'tRFAla and 5'tiRNAAla levels and so suppressing the translation of their matched DVL2 and DVL3 mRNAs that thereby inhibits the β-catenin pathway and so reversibly switching HCC cells into a quiescence state. As such, this study suggests that, like miRNAs, tsRNAs might activate the translation of their matched mRNAs in non-dividing cells and so providing a possible means for suppressing tumor cell growth

overall_design: tsRNAs’ expression in hepatocellular carcinoma (HCC) PLC/PEF/5 cell was measured at 48h after exposure to doses of 8 μM ELT. The total cellular RNA of each sample was respectively extracted by Trizol and was mixed by three cell culture dishes to minimize errors.

first: Zhipeng

last: Zhang

first: Yanmei

last: Wu

first: Wenqiang

last: Liang

first: Hongbo

last: Liao

first: Xingxing

last: Xing

first: Wenxin

last: Yi

first: Zixuan

last: Liu

first: Yicheng

last: Li

first: Dongling

last: Lin

first: Ting

last: Gu

first: Mingzhi

last: Zou

first: Martin

last: Banwell

first: Huilai

last: Miao

first: Xin