title: A distinct human cell type expressing MHCII and RORγt with dual characteristics of dendritic cells and type 3 innate lymphoid cells

city: Saint Louis

first: Alina

last: Ulezko Antonova

state: MO

address: 660 S. Euclid Ave

department: Path/Immuno

institute: WUSM

summary: Recent studies have characterized various mouse antigen-presenting cells (APCs) expressing the lymphoid-lineage transcription factor RORγt, which exhibit distinct phenotypic features and are implicated in the induction of peripheral regulatory T cells (Tregs) and immune tolerance to microbiota and self-antigens. These APCs encompass Janus cells and Thetis cell subsets, some of which express the autoimmune regulator AIRE. RORγt+ MHCII+ type 3 innate lymphoid cells (ILC3) have also been implicated in the instruction of microbiota specific Tregs. While RORγt+ APCs have been actively investigated in mice, the identity and function of these cell subsets in humans remain elusive. Herein, we identify a rare subset of RORγt+ cells with dendritic cell (DC) features through integrated single-cell RNA sequencing and single-cell ATAC sequencing. These cells, which we term RORγt+ DC-like cells (R-DC-like), exhibit DC morphology, express the MHC class II machinery, are distinct from all previously reported DC and ILC3 subsets, but share transcriptional and epigenetic similarities with DC2 and ILC3. We have developed procedures to isolate and expand them in vitro, enabling their functional characterization. R-DC-like cells proliferate in vitro, continue to express RORγt, and differentiate into CD1c+ DC2-like cells. They stimulate the proliferation of allogeneic T cells. The identification of human R-DC-like cells with proliferative potential and plasticity towards CD1c+ DC2-like cells will prompt further investigation into their impact on immune homeostasis, inflammation, and autoimmunity.

overall_design: Tonsils were obtained from elective tonsillectomies, mechanically disrupted, and processed for analysis. Ileum samples were obtained from surgical waste. Different enrichment methods were used for isolating specific cell populations from the samples. FACS-sorted purified CD34+ cells from bone marrow or peripheral blood were cultured for 15 days in the presence of GFP-expressing OP9-Delta-like1 cells and either 2% of FLT3L supernatant, 20ng/ml SCF and 20ng/ml GM-CSF or 0.5% of FLT3L supernatant, 20ng/ml SCF and 20ng/ml IL-7. Cells were sorted at day 15 for viability and CD45 expression and subjected to scRNAseq.  The study followed approved human protocols by the Institutional Review Board (IRB) at Washington University School of Medicine.The sorted cells were then subjected to sequencing using the 10X Genomics Platform 3' with chemistry version 2 and version 3 for lung samples only.

first: Silvia

last: Lonardi

first: Marina

last: Cella

first: Marco

last: Colonna


title: LPS-induced systemic inflammation mediates a protective effect against ischemic injury via microglia

status: Public on Dec 19 2023

city: Chuo

first: Kozo

last: Saito

address: 1110 Shimokato

department: Department of Neuropharmacology

country: Japan

institute: University of Yamanashi

type: Expression profiling by high throughput sequencing

summary: LPS-induced systemic inflammation mediates a protective action on ischemic injury via microglia. We used RNA sequencing analysis (RNA-seq) to gain molecular insights into microglia-mediated protection of cerebral infarction.

overall_design: We isolated CD11b+ cells from LPS-treatd mice or saline-treated mice 7 days after staritng administration using magnetic-activated cell sorting (MACS) procedure.

first: Le

last: Phung


title: Distinct guard cell–specific remodeling of chromatin accessibility during abscisic acid– and CO2-dependent stomatal regulation

city: La Jolla

first: Charles

middle: Anthony

last: Seller

state: CA

address: 9500 Gilman Dr

department: Cell and Developmental Biology

country: USA

institute: UC San Diego

type: Genome binding/occupancy profiling by high throughput sequencing

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

overall_design: Refer to individual Series


title: Distinct guard cell specific remodeling of chromatin accessibility during abscisic acid and CO2 dependent stomatal regulation (RNA-Seq)

summary: In plants, epidermal guard cells integrate and respond to numerous environmental signals to control stomatal pore apertures thereby regulating gas exchange. Chromatin structure controls transcription factor access to the genome, but whether large-scale chromatin remodeling occurs in guard cells during stomatal movements, and in response to the hormone abscisic acid (ABA) in general, remain unknown. Here we isolate guard cell nuclei from Arabidopsis thaliana plants to examine whether the physiological signals, ABA and CO2, regulate guard cell chromatin during stomatal movements. Our cell type specific analyses uncover patterns of chromatin accessibility specific to guard cells and define novel cis-regulatory sequences supporting guard cell specific gene expression. We find that ABA triggers extensive and dynamic chromatin remodeling in guard cells, roots, and mesophyll cells with clear patterns of cell-type specificity. DNA motif analyses uncover binding sites for distinct transcription factors enriched in ABA-induced and ABA-repressed chromatin. We identify the ABF/AREB bZIP-type transcription factors that are required for ABA-triggered chromatin opening in guard cells and implicate the inhibition of a set of bHLH-type transcription factors in controlling ABA-repressed chromatin. Moreover, we demonstrate that ABA and CO2 induce distinct programs of chromatin remodeling. We provide insight into the control of guard cell chromatin dynamics and propose that ABA-induced chromatin remodeling primes the genome for abiotic stress resistance.

overall_design: We used RNA-sequencing to examine the transcriptional response to abscisic acid (ABA) in whole leaves and guard cells of the plant Arabidopsis thaliana. 5-to-6 week-old plants were sprayed with either 50 µM ABA or a vehicle control, and after 4 hours we extracted total RNA. We conducted two biological replicates per sample.

middle: A

first: Julian

middle: I

last: Schroeder


title: Genome-wide transcriptome analysis of the brain (ventral part of telenchephalon, hypothalamus and pituitary) of medaka (Oryzias latipes) kept under constant photoperiodic conditions

city: Nagoya

first: Takashi

last: Yoshimura

address: Furo-cho, Chikusa-ku

institute: Nagoya University

summary: Expression profile of the brain (ventral part of telenchephalon, hypothalamus and pituitary) of medaka (Oryzias latipes) kept under constant photoperiodic conditions were obtained by RNA-seq analysis. Monthly transcriptomes under constant photoperiodic conditions identified 518 circannual genes.

overall_design: To investigated the expression profile of each gene under constant photoperiodic conditions, samples were collected every month and RNA-seq analysis was performed using these time-series samples.


title: Genome-wide transcriptome analysis of the brain (ventral part of telenchephalon, hypothalamus and pituitary) of medaka (Oryzias latipes) kept under outdoor natural conditions over a period of two years

summary: Expression profile of the brain (ventral part of telenchephalon, hypothalamus and pituitary) of medaka (Oryzias latipes) kept under outdoor natural conditions were obtained by RNA-seq analysis. Monthly transcriptomes for two years identified 3,341 seasonally-oscillating genes.

overall_design: To investigated the yearly expression profile of each gene in each season, samples were collected every month at around 8 pm (Japan Standard Time) for two years, from October 2015 to October 2017 from medaka kept under outdoor natural conditions. RNA-seq analysis was performed using these time-series samples.


title: Genome-wide transcriptome analysis of the brain (ventral part of telenchephalon, hypothalamus and pituitary) of medaka (Oryzias latipes) at the spring equinox, summer solstice, autumn equinox, and winter solstice [24h RNA-seq]

summary: Expression profile of the brain (ventral part of telenchephalon, hypothalamus and pituitary) of medaka (Oryzias latipes) kept under outdoor natural conditions were obtained by RNA-seq analysis. Daily transcriptomes on the four equinoxes/solstices identified 1,381 daily-oscillating genes.

overall_design: To investigated the daily expression profile of each gene in each season, samples were collected every 4 h for a 24-h period at the spring equinox, summer solstice, autumn equinox, and winter solstice in 2018 and 2019 and RNA-seq analysis was performed using these time-series samples.


title: Distinct guard cell specific remodeling of chromatin accessibility during abscisic acid and CO2 dependent stomatal regulation (ATAC-Seq)

overall_design: We used the Assay for Transposaes Accessible Chromatin with sequencing (ATAC-seq) to profile genome-wide patterns of chromatin accessibility in different Arabidopsis thaliana cell-types (seedling roots, mesophyll cells, and guard cells), and following exposure of plants to different treatments (abscisic acid, 100 ppm atmospheric carbon dioxide, or 1000 ppm atmospheric carbon dioxide). Nuclei from different cell-types were purified using FACS performed on samples derived form transgenic plants expressing cell-type specific nuclear fluorescent markers (Histone H2B tagged with EGFP). All experiments were performed with a minimum of 2 biological replicates per cell-type and treatment.


title: Dissecting crosstalk between embryonic and extra-embryonic tissues with stem cell co-culture [II]

city: Kunming

first: Mq

last: Peng

state: Yunnan

address: No. 727, Jingming South Road, Chenggong District

country: China

institute: Kunming University of Science and Technology

summary: Faithful embryogenesis requires the precise coordination between embryonic and extraembryonic tissues. Although embryonic and extraembryonic stem cells have been derived from several mammalian species including humans, they are cultured in different conditions, which makes it difficult to study their intercommunication. Here, by simultaneously activating FGF, TGF-β and WNT pathways, we derived stable pluripotent stem cells (PSCs), trophoblast stem cells (TSCs) and extraembryonic endoderm stem cells (XENs) from mouse blastocysts under the same condition (FTW). Co-culture of PSCs and XENs in the same environment uncovered, among other interactions, a previously unrecognized control of proliferation of epiblast cells by extraembryonic endoderm cells. FTW condition also supported de novo derivation of XENs from cynomolgus monkey and human blastocysts, and enabled setting up co-culture of human iPSCs and XENs. Crosspieces comparison revealed conserved and divergent processes and genes regulating XENs and ligand-receptor interactions between pluripotent and extraembryonic endoderm cells. Our study establishes a unique stem cell co-culture strategy to examine embryonic and extraembryonic lineage crosstalk during early mammalian development, and opens the door for developing more faithful in vitro models and differentiation protocols.

overall_design: Totally 7 samples were sequenced, including 1 mouse blastocyst sample, 1 mouse blastocyst derevation in FTW medium for 8 days sample, 1 FTW-mESCs sample, 1 FTW-mTSCs sample, 1 FTW-mXENs sample, and 2 replicates of FTW-mESCs/FTW-mTSCs/FTW-mXENs co-culture for 5 days samples.

first: Jun

last: Wu

first: Yulei

last: Wei


title: XBP1s-Mediated Endoplasmic Reticulum Proteostasis Network Enhancement Can Selectively Improve Folding and Secretion of an Osteogenesis Imperfecta-Causing Collagen-I Variant

city: Cambridge

first: Matthew

middle: D

last: Shoulders

state: MA

address: 77 Massachusetts Avenue

department: Chemistry

institute: Massachusetts Institute of Technology

summary: Osteogenesis imperfecta (OI) is most commonly caused by autosomal dominant mutations in genes encoding collagen type-I. Here, we test the hypothesis that modulation of the endoplasmic reticulum (ER) proteostasis network via the unfolded protein response (UPR) can improve the folding and secretion of the lethal osteogenesis imperfecta (OI)-causing G425S a1(I) variant. We show that specific induction of the UPR’s XBP1s transcriptional response enhances G425S a1(I) secretion up to ~300% of basal levels. Notably, the effect is selective – WT a1(I) secretion is unaltered by XBP1s. XBP1s pathway activation appears to post-translationally enhance the folding/assembly and secretion of G425S a1(I). Consistent with this notion, we find that the stable, triple-helical collagen-I secreted by XBP1s-activated G425S a1(I) patient fibroblasts includes a higher proportion of the mutant a1(I) polypeptide than the collagen-I secreted under basal ER proteostasis conditions.

overall_design: RNA-seq characterizing two cell lines: WT donor or G425S a1(I) primary patient fibroblast cells. These cell lines were transduced with adenovirus encoding XBP1s or GFP (in triplicate). All cells were treated with 200 uM sodium ascorbate for 24 h before harvesting.

first: Ngoc-Duc

last: Doan

first: Agata

last: Bikovtseva


title: Comprehensive analysis of mRNA 5-methylcytosine modification in Eimeria tenella oocysts

city: Jinzhong

first: Qing

last: Liu

address: Taigu, Shanxi Province, PR China

institute: Shanxi Agricultural university

type: Methylation profiling by high throughput sequencing

summary: 5-methylcytosine (m5C) is a type of RNA modifications reported to regulate diverse biological processes. However, the distribution and biological functions of m5C in E. tenella mRNAs remain largely unknown. Herein, we report transcriptome-wide profiling of mRNA m5C in E. tenella by applying m5C RNA immunoprecipitation followed by a deep-sequencing approach (m5C-RIP-seq). Our data showed that m5C peaks were distributed across the whole mRNA body. There were 2813 hypermethylated and 1850 hypomethylated m5C peaks in sporulated oocysts compared with unsporulated oocysts. Further joint analysis showed a positive correlation between m5C modification and gene expression levels. The mRNA sequencing and m5C-RIP-seq data were consistent with the results of the quantitative reverse transcription PCR (RT-qPCR) and m5C-RIP-qPCR, respectively. Gene ontology (GO) and pathway enrichment analysis predicated diverse biological functions and pathways, including microtubule motor activity, helicase activity, cGMP-PKG signaling pathway, aminoacyl-tRNA biosynthesis, glycolysis/gluconeogenesis and spliceosome. Meanwhile, stage-specific gene expression signatures of m5C-related enzymes were observed. Altogether, our findings revealed the transcriptional significance of m5C modification in E. tenella oocysts, providing clues for in-depth research.

overall_design: MeRIP-seq and RNA-seq of sporulated and unsporulated oocysts of E. tenella.

first: Lin-Mei

last: Yu

first: Zi-Rui

last: Wang

first: Qing-Xin

last: Fan

first: Tao

last: Jia

first: Tian-Hong

last: Zhang

first: Xing-Quan

last: Zhu


title: Activation domains and Coactivator direct genomic localization of fused DNA binding domains, defining the subset of activated promoters [RNA-seq]

city: Rehovot

first: Naama

last: Barkai

state: HaMerkaz

address: Herzl 234

department: Molecular Genetics

country: Israel

institute: Weizmann Institute of Science

summary: Activation domains (ADs) within transcription factors (TFs) induce gene expression by recruiting coactivators to specific regulatory regions. Within the prevailing model, TF-coactivator recruitment is independent of DNA binding, which is consistent with direct AD-coactivator interactions seen outside cells. However, this independence was not yet tested within the genomic context. Here, we targeted two Med15-interacting ADs to hundreds of budding yeast promoters through fusions with multiple DNA binding domains (DBDs), gradually controlling their abundances using libraries of synthetic promoters. Genomic profiling revealed that AD identity influences DNA binding locations and that transcription induction and Med15 recruitment are restricted to a subset of DBD-bound promoters displaying flexible expression, multiple-TFs binding, and fuzzy nucleosome architecture. Further, when fused to a DBD, Med15 redirected binding towards promoters of fuzzy nucleosomes, overcoming DBD-based preferences. Our results demonstrate that ADs and their recruited coactivators posses an inherent preference for genomic localization and, therefore, define the subset of induced promoters.

overall_design: The experiments here are RNA-seq samples, collection of samples was performed at OD600 of 0.4, for controls we used the expression profiling of non-genetically modified yeast strain (BY4741).

first: Vladimir

last: Mindel

first: Sagie

last: Brodsky

first: Hadas

last: Yung

first: Wajd

last: Manadre


title: Single-Cell Gene Expression Profiling of CD45+ Cells from Tumors and Draining Lymph Nodes in Trp1 CD4+ T Cell-Treated B16F10 Melanoma-Bearing Mice

status: Public on Dec 18 2023

city: Houston

first: Dawei

last: Zou

state: TX

address: 6670 Bertner Ave

department: Surgery

institute: Houston Methodist Research Institute

summary: CD4+ T cells play a crucial role in tumor eradication. Employing single-cell RNA sequencing (scRNA-seq), we analyzed the cell states of CD45+ immune cells from tumors and draining lymph nodes in Trp1 CD4+ T cell-treated B16F10 melanoma-bearing mice. This investigation aims to enhance our understanding of how CD4 T cells mediate tumor regression.

overall_design: B16F10 melanoma-bearing mice were treated with either naïve Trp1 cells or PBS as a control. After 8 days, CD45+ cells from tumors and draining lymph nodes in the Trp1 cell transfer group were sorted, alongside CD45+ cells isolated from the tumors of the PBS control group (no cell transfer). Subsequently, these cells were subjected to analysis using single-cell RNA sequencing (scRNA-seq).

first: Adam

middle: Q

last: He

first: Wenhao

last: Chen


title: Unravelling the heterogeneous molecular landscape of pediatric post-transplant lymphoproliferative disorders

city: Barcelona

first: Itziar

last: Salaverria

address: Rossello 153

country: Spain

institute: Hospital Clínic

type:
SNP genotyping by SNP array
Genome variation profiling by SNP array

summary: It is unknown whether pediatric monomorphic post-transplant lymphoproliferative disorders (mPTLD) display similar genetic features than the immunocompetent counterpart and if they resemble adult mPTLD. We have investigated 39 pediatric mPTLD, 33 diffuse large B-cell lymphoma (DLBCL) and six Burkitt lymphoma (BL), by an integrated approach, including fluorescence in situ hybridization, cell of origin determination (COO), targeted gene sequencing and copy-number arrays. According to COO, 24/28 DLBCL (86%) were classified as activated B-cell (ABC) and all six BL were germinal center B cell (GCB)-type. Thirty-three out of 37 investigated mPTLD were positive for EBV infection.   Overall, PTLD-BL carried mutations in MYC in addition to ID3 and DDX3X, ARID1A or CCND3 and a higher mutational burden than PTLD-DLBCL (12.3 vs 6.2, P = 0.01). CN profile of PTLD-BL was less complex than in IC-BL (1 vs 6.26; P < 0.005). PTLD-DLBCL showed a very heterogeneous genomic profile characterized by a lower number of mutations (2.4 vs 6.5, P=0.01) and less CNA (2.10 vs 4.36 ; P < 0.05) than in IC patients. Pathway enrichment analysis revealed that epigenetic modifiers and Notch pathway (4 cases each) were the most recurrently affected.   In conclusion, these findings further unravel for the first time the molecular heterogeneity of pediatric mPTLD and provide new parameters for the design of more effective therapeutic strategies.

platform_organism:
Homo sapiens
Mus musculus

overall_design: Thirty-nine monomorphic PTLD <19 years-old (mean 10y, gender 25 male/14 female) were recruited and analyzed for germinal center markers, IRF4 and EBER expression. Presence of MYC, PAX5, IRF4, BCL2, BCL6 and 11q alterations was investigated by FISH. Additional molecular studies included clonality, copy number (CN) arrays, cell of origin-COO (Nanostring) and mutational analyses (Custom 167 lymphoma related genes panel, SureSelectXT, Agilent).

first: Julia

last: Salmeron Villalobos

first: Natalia

last: Castrejon de Anta

first: Olga

last: Balagué


title: Whole-genome bisulfite sequencing analysis for intestinal budding organoids derived from induced or tissue-derived intestinal stem cells, and for mouse embryonic fibroblasts

city: Fukuoka

first: Atsushi

last: Suzuki

state: Fukuoka

address: 3-1-1 Maidashi, Higashi-ku

department: Medical Institute of Bioregulation

institute: Kyushu University

summary: We have previously succeeded in generating induced fetal intestine-derived progenitor cells (iFIPCs) by expressing a gene set (Foxa3/Cdx2/Gata6/Hnf4a) in mouse embryonic fibroblasts (MEFs), which form spherical organoids (SOs) in three-dimensional culture, and subsequently maturated them into adult-type induced intestinal stem cells (iISCs), which form budding organoids (BOs) like tissue-derived intestinal stem cells (ISCs) (Miura & Suzuki, 2017). Although CpG methylation of genomic DNA is an essential epigenetic factor affecting the transcription during cell differentiation, little is known about how DNA methylation changes and what role it plays in the direct reprogramming. Therefore, we performed a genome-wide methylome analysis for iISC- and ISC-derived BOs (iISC-BO and ISC-BO, respectively) and MEFs.

overall_design: We performed genome-wide and base-resolution methylome analysis using the post-bisulfite adaptor-tagging (PBAT) method (Miura et al., 2012) in MEFs, iISC-BOs, and ISC-BOs.

first: Kenichi

last: Horisawa

first: Shizuka

last: Miura

first: Hiromitsu

last: Araki

first: Fumihito

first: Takashi

last: Ito


title: A Whole-Cell Screening Platform for Discovering Cell Adhesion Molecules Enabling Programmable Bacterial Cell-Cell Adhesion

city: Taipei

first: See-Yeun

last: Ting

address: 128 Academia Road, Section 2, Nankang

country: Taiwan

institute: Academia Sinica

summary: To identify cell adhesion molecules (CAMs) targeting bacterial membrane proteins within a synthetic bacteria-displayed nanobody library, we present a comprehensive whole-cell screening platform. This involves targeted amplicon sequencing to discover nanobodies targeting the natural adhesin, TraN. Furthermore, we employ deep mutational engineering to enhance the binding affinity of these nanobodies toward TraN.

overall_design: Nanobody genes were sequenced after extracting plasmids containing these genes from a bacterial whole-cell display platform, both before and after 1-3 rounds of selection. The selection process aimed to identify nanobodies specifically targeting the surface-exposed domain of TraN from the pSLT plasmid of Salmonella Typhimurium LT2 strain.

first: Po-Yin


title: TMAO promoted TREM-1 expression through NLRP3 inflammasome

city: Wuhan

first: Junhao

last: Wan

state: Hubei

address: Jiefang Road

institute: Union Hospital, Tongji Medical College, Huazhong University of Science and Technology

summary: To investigate the impact of TMAO on Myeloid cells. An RNA-seq analysis of myeloid cells was performed with and without TMAO treatment.

first: Fengjing

last: Yang

first: Song

last: Tong

first: Chuangyan

last: Wu

first: Ting

last: Zhou

first: Sihua

last: Wang


title: Epithelial overexpression of IL-33 induces eosinophilic esophagitis dependent on IL-13

city: Scottsdale

first: Alfred

last: Doyle

state: Arizona

address: 13400 E Shea Blvd

institute: Mayo Clinic

summary: To examine the role of the epithelial-derived cytokine, IL-33, in eosinophilic esophagitis (EoE) pathogenesis we developed a novel mouse model (EoE33) in which a secreted and active form of IL-33 is overexpressed by esophageal epithelial cells.

overall_design: To assess transcriptional changes associated with esophageal overexpression of IL-33, bulk RNA sequencing was performed on whole esophageal tissue biopsies from eight week old male transgenic EoE33 mice (n=4) and wild-type C57BL/6J littermates (n=4).

first: Benjamin

last: Wright


title: Gene profilling of splenic CD4+ T cells from naïve H2-O Wildtype and H2-O Knockout mice

city: Baltimore

first: Scheherazade

last: Sadegh Nasseri

state: MD

address: 720 Rutland Ave, Ross Building Rm 664C

department: Pathology

institute: Johns Hopkins University

summary: Loss of the MHC Class II accessory molecule H2-O has been shown to alter the peptide repertoir presented by MHC Class II molecules. Using scRNA-Seq we interrogated what outcomes altered antigen presentation was having on the naïve CD4 T cell milieu.

overall_design: CD3+ CD4+ (B220/NK1.1 negative) T cells were FACS sorted froms the spleens of 3 H2-O Wildtpe and 3 H2-O knock-out mice and analyzed by scRNA-Seq analysis.

last: Sadegh-Nasseri


title: Association of tumor burden with the efficacy of PD-(L)1 inhibitors for treatment-naive advanced non-small-cell lung cancer

city: Osakasayama City

last: Kurosaki

address: 377-2 Ohno-higashi

institute: kindai university

summary: A high tumor mutation was associated with a poor outcome of immune checkpoint inhibitor therapy for advanced NSCLC as a result of immunosuppressive phenotypes.

overall_design: We retrospectively reviewed advanced NSCLC patients and evaluated the association between tumor burden and immune-related gene expression profiling.

first: Shinichiro

first: Koji

last: Haratani


title: Effect of PCGF6 loss of function in intestinal adult epithelial cells

city: MILANO

first: Diego

last: Pasini

state: MI

address: Via Adamello 16

country: Italy

institute: European Institute of Oncology

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

overall_design: Refer to individual Series


title: Effect of PCGF6 loss of function in intestinal adult epithelial cells [RNA-seq]

summary:
We generated RNAseq expression analyses of PCGF6 WT and KO intestinal crypts in two independent replicates using two distinct transgenic models (AhCre and Villin Cre-ERT2)
We then performed differential expression analyses of PCGF6 KO vs. WT independently in the AhCre and Villin transgenic models

overall_design: Intestinal crypts of 8-12 weeks old sex match (male) mice were collected 7 days post Tamoxifen (Villin) or BNF (AhCre) administration for total RNA isolation followed by SMART-seq NGS library preparation

first: Annachiara

last: Del Vecchio

first: Daniel

last: Fernández-Pérez


title: Effect of PCGF6 loss of function in intestinal adult epithelial cells [ChIP-seq]


title: Gene expression and metabolic activity of Streptococcus mutans during exposure to dietary carbohydrates glucose, sucrose, lactose and xylitol

city: Brno

first: Martina

last: Zapletalová

address: Kamenice 5

department: Department of Biochemistry

country: Czech Republic

institute: Masaryk University

summary: Recent RNA-seq studies have given us a deeper insight into the cariogenic impact of carbohydrate sources in the bacterium Streptococcus mutans, the principal etiological agent of human dental caries. The process of dental caries development is given by the ability of this bacterium to ferment some carbohydrates up to organic acids contributing to the pH decrease in the oral cavity and the demineralization of the enamel. Besides, in dental caries progression, an important role is played by biofilm formation, which starts and ends with free planktonic cells, and it is given by several unique properties called virulence factors. The most cariogenic carbohydrate is considered sucrose, an easily metabolizable source of energy inducing acidification and synthesis of glucans forming typical bacterial cell clumps. By using multifaceted methodological approaches, we compared the transcriptomic and metabolomic profiles of S. mutans growing in planktonic culture on preferred and non-preferred carbohydrates and fasting conditions. Lactose and xylitol showed high effectiveness in the regulation of the IPS metabolism, cell wall structure and overall virulence involved in the initial phase of biofilm formation and structure but with an opposite pattern compared to sucrose and glucose. These findings confirm the recent results that xylitol and lactose play a vital role in biofilm structure but do not reduce its formation related to their cariogenic potential.

overall_design: Identification of transcriptomic changes of S. mutans depending on different carbohydrate source - glucose, sucrose, lactose and xylitol, each in triplicate; and cultivation without sugar as control

first: Veronika

last: Zajdlikova

last: Krestakova

first: Jiri

last: Kucera

first: Katerina

last: Dadakova

last: Zapletalova

last: Paskova

first: Roman

last: Reminek

first: Zdenek

last: Glatz

first: Lydie

last: Izakovicova Holla

first: Filip

last: Ruzicka

first: Jan

last: Lochman

first: Petra

last: Borilova Linhartova


title: T cell responses in SARS-CoV-2 vaccinees with sustained or declined antibody titer

city: Suita

first: Xiuyuan

last: Lu

state: Osaka

address: 3-1 Yamadaoka

department: IFReC

institute: Osaka University

summary: SARS-CoV-2 vaccines have been used worldwide to combat COVID-19 pandemic. To elucidate the factors that determine the longevity of spike (S)-specific antibodies, we traced the characteristics of S-specific T cell clonotypes together with their epitopes and anti-S antibody titers before and after BNT162b2 vaccination over time. T cell receptor (TCR) alpha/beta sequences and mRNA expression of the S-responded T cells were investigated using single-cell TCR- and RNA-sequencing. In donors exhibiting sustained anti-S antibody titers (designated as “sustainers”), S-reactive T cell clonotypes detected immediately after 2nd vaccination polarized to follicular helper T (Tfh) cells, which was less obvious in “decliners”. Even before vaccination, S-reactive CD4+ T cell clonotypes did exist, most of which cross-reacted with environmental or symbiotic bacteria. However, these clonotypes contracted after vaccination. Conversely, S-reactive clonotypes dominated after vaccination were undetectable in pre-vaccinated T cell pool, suggesting that highly-responding S-reactive T cells were established by vaccination from rare clonotypes. These results suggest that de novo acquisition of memory Tfh cells upon vaccination contributes to the longevity of anti-S antibody titers.

overall_design: Among 8 age- and sex-matched donors who accepted 2 doses of SARS-CoV-2 vaccine BNT162b2, #4, #13, #15 and #17 had declined anti-S antibody titers from 6 weeks (wks) to 24 wks after the 1st vaccination, while #8, #25, #27 and #28 had sustained antibody titers. PBMCs from these 8 donors at 3 wks, 6wks, and 24 wks after the 1st vaccination were collected. PBMCs from donor #4, #8, #13, #15 and #17 before the vaccinations were also collected. PBMCs were stimulated with SARS-CoV-2 S peptide pool, and proliferated T cells were collected and analyzed by scRNA/TCR-seq.

first: Hiroki

last: Hayashi

first: Daisuke

last: Motooka

first: Hironori

last: Nakagami

first: Sho

last: Yamasaki


title: RNaseH1KD_RNA-seq

city: Shanghai

first: Li

last: Yang

address: 131 Dong-An Road

department: Institutes of Biological Sciences

institute: Fudan University

summary: Identify ciRNAs after RNaseH1 knockdown by RNA-seq in PA1 cell line.

overall_design: RNA-seq of RNaseH1 knockdown PA1 cells.

first: Xiang

last: Li

first: Yun

middle: N

last: Lei

first: Fang

first: Ling

middle: L

last: Chen


title: Genome-wide binding of the bromdomain protein TgBDP3 in the protozoan parasite Toxoplasma gondii

city: Indianapolis

first: Victoria

last: Jeffers

state: IN

address: Pharmacology & Toxicology

institute: Indiana University School of Medicine

summary: The bromdomain protein TgBDP3 is associated with the TFIID transcriptioanl initiation complex in Toxoplasma gondii. To determine the genes that are regulated by TgBDP3, the endogenous tgbdp3 locus was modified to introduce a C-terminal HA epitope tag. ChIP-seq was performed on intracellular, replicating tachyzoites expressing TgBDP3-HA. The protein was enriched in the upstream, promoter region of a subset of actively transcribed parasite genes indicating that TgBDP3 has a role in regulation of transcriptional initiation in Toxoplasma.

overall_design: ChIP-seq of a chromatin-associated bromodomain protein in Toxoplasma gondii

first: Rusch

last: Douglas B.

first: William

middle: J

last: Sullivan Jr


title: ANKRD52_SHAPE-MaP

summary: Probe ANKRD52 secondary structure by SHAPE-MaP in PA1 cell line.

overall_design: SHAPE-MaP of in vitro ANKRD52 with or without DNA template.


title: Transcriptional profiling of intestinal tissue from mice with and without Npm1 deficiency.

status: Public on Dec 17 2023

city: Glasgow

first: Kathryn

last: Gilroy

address: Switchback Road

country: United Kingdom

institute: Cancer Research UK Scotland Institute

summary: A profile of gene expression differences in mouse intestine with the noted transgenic alleles, in the presence and absence of functional Npm1.

overall_design: Expression data from intestine with the noted transgenes with and without Npm1 deficiency.

first: Georgios

last: Kanellos

first: Owen

last: Sansom


title: Linking circular intronic RNA degradation and function in transcription by RNase H1


title: Transcriptome analysis of striatal synaptosome and synaptosomal polyribosome of WT and  Shank3 TG mice

city: Seoul

first: Kihoon

last: Han

address: 73, Inchon-ro, Seongbuk-gu

department: Neuroscience

country: South Korea

institute: Korea University College of Medicine

summary: Shank3 protein is a core organizer of the macromolecular complex in excitatory postsynapses, and its defects cause numerous synaptopathies including autism spectrum disorders. Although Shank3 function as a postsynaptic scaffold is well established, other potential mechanisms how Shank3 broadly modulates the postsynaptic proteome are relatively unexplored. To understand potential contribution of increased proteins synthesis to the proteomic change in TG striatal synaptosome, we performed RNA-seq analyses on both whole synaptosomal and synaptic polysome-enriched fractions. Comparative analyses showed a positive correlation only between polysome-associated transcriptome and up-regulated proteome in TG striatal synaptosome.

overall_design: Transcriptome of striatal synaptosome and synaptosomal polyribosome of WT and  Shank3 TG mice

first: Chunmei

last: Jin

first: Yeunkum

last: Lee

first: Hyojin

last: Kang


title: Human cranial bone-derived mesenchymal stem cells cultured under microgravity can improve cerebral infarction in rats

city: Hiroshima

first: Masashi

last: Kuwabara

address: 1-2-3 Kasumi, Minami-ku

department: Neurosurgery

institute: Graduate School of Biomedical and Health Sciences, Hiroshima University

type: Non-coding RNA profiling by high throughput sequencing

summary: Transplantation of human cranial bone-derived mesenchymal stem cells (hcMSCs) in central nervous system disorders has the potential to improve motor function by highly expressing neurotrophic factors. However, the effects of hcMSCs cultured under microgravity (MG) conditions on cerebral infarction have not been investigated before. Thus, this study aimed to investigate the transplantation effects of hcMSCs cultured in a simulated MG environment on cerebral infarction model rats. Our results showed that neurological function was significantly improved after transplantation of hcMSCs cultured in the MG environment (MG group) compared with normal gravity environment (1G group). Protein expressions of nerve growth factor, fibroblast growth factor 2, and synaptophysin were significantly higher in the MG group than in the IG group, whereas sortilin 1 expression was significantly lower. MicroRNA analysis revealed that genes related to cell proliferation, angiogenesis, neurotrophy, anti-apoptosis, neural and synaptic organization, and cell differentiation inhibition were significantly upregulated in the MG group. In contrast, genes promoting microtubule and extracellular matrix formation and cell adhesion, signaling, and differentiation were downregulated. These results indicate that hcMSCs cultured in an MG environment may be a useful source of stem cells for recovery of neurological function after cerebral infarction.

overall_design: Cerebral infarction model rats were generated; hcMSCs cultured in normal gravity (1G) or MG environment were transplanted via the transvenous route on the first day after induction of cerebral infarction for immunohistological analysis and neurological function evaluation. The expression of endogenous neurotrophic factors, axonal, neuronal, and synaptogenic factors, angiogenic factors, and apoptosis-related factors in infarcted brain tissue at 35 days after stroke induction was examined by real-time polymerase chain reaction (PCR) and western blotting analyses. Sequencing analysis of miRNAs was performed in hcMSCs cultured under 1G or MG environments.

first: Takafumi

last: Mitsuhara

first: Masataka

last: Teranishi

first: Takahito

last: Okazaki

first: Masaaki

last: Takeda

first: Daizo

last: Ishii

first: Hiroshi

last: Kondo

first: Kiyoharu

last: Shimizu

first: Masahiro

last: Hosogai

first: Takeshi

last: hara

first: Yuyo

last: Maeda

first: Tomoyuki

last: Kurose

first: Yumi

last: Kawahara

first: Louis

last: Yuge

first: Nobutaka

last: Horie


title: Basal amygdala-nucleus accumbens glutamate neurons respond to either social reward or social aversion

city: Biberach an der Riss

first: Gregorio

last: Alanis-Lobato

address: Birkendorfer Str 65

department: Global Computational Biology and Digital Sciences

country: Germany

institute: Boehringer Ingelheim Pharma

summary: A substantial proportion of basal amygdala (BA) glutamate neurons project to nucleus accumbens (NAc). The evidence that these neurons are activated by reward and/or aversion is equivocal. Social stimuli are highly salient, and in male mice we conducted a detailed analysis of the responsiveness of BA-NAc neurons to estrous female (social reward, SR) or aggressive male (social aversion, SA). Both SR and SA activated c-Fos expression in a relatively high number of BA-NAc neurons in intermediate (int) BA. Using Fos-TRAP2 mice, the majority of social int-BA-NAc neurons were activated by either SR or SA, i.e. were monovalent, and in similar numbers. Fiber photometry provided corroborative evidence that int-BA-NAc neural pathway activity was similar in response to SR or SA. These findings contribute substantially to understanding the topography and valence-specificity of BA-NAc neurons with respect to highly salient stimuli, and to identifying molecular targets for treatment of reward- or aversion-specific psychopathologies.

overall_design: The aims of this experiment were to measure transcriptome expression of the BA-NAc neuron population at the medial a-p BA region where these neurons are responsive to social reward or social aversion and compare it with the posterior a-p region where neurons responsive to social aversion were in the majority.

first: Giulia

last: Poggi

first: Coralie

last: Viollet

first: Christopher

middle: R

last: Pryce


title: Expression data from carotid body glomus cells and neuroblasts, from young adult rats (9-11 weeks of age)

city: Seville

first: Ricardo

last: Pardal

address: Avda. Manuel Siurot, s/n

department: Medical Physiology and Biophysics

institute: University of Seville

summary:
The carotid body is a peripheral chemoreceptor organ containing a neurogenic niche with stem cells and immature neuronal cells that convert into new neurons (glomus cells) upon activation by hypoxia.
We have used microarray to compare the gene expression programs of glomus cells and neuroblasts, both in normoxic and hypoxic (3 days) conditions, to highlight the molecular mechanisms underlying fast neurogenesis

overall_design: Glomus cells and neuroblasts were sorted by flow cytometry from enzymatically dispersed carotid bodies, from both normoxic and 3-days hypoxic rats. Total RNA was extracted and hybridized on Affymetrix microarrays

first: Verónica

last: Sobrino


title: ChIP-seq analysis of HEK cells transfected with SARS-CoV-2 protein Orf8

city: Philadelphia

first: Erica

last: Korb

state: PENNSYLVANIA

address: 3400 Civic Center Boulevard

institute: University of Pennsylvania

summary: Analysis of histone mimicry by SARS-CoV-2 Orf8 encoded protein

overall_design: HEK cells were transfected with GFP or Orf8 and isolated 48 hours after transfection for ChIP-seq


title: The dilated cardiomyopathy-associated RNA Binding Motif Protein 20 regulates long pre-mRNAs in neurons [seCLIP-Seq]

status: Public on Dec 16 2023

city: basel

first: peter

last: scheiffele

address: spitalstrasse 41

country: Switzerland

institute: Biozentrum, university of Basel

summary: Precise coordination of molecular programs and neuronal growth govern the formation, maintenance, and adaptation of neuronal circuits. RNA metabolism has emerged as a key regulatory node of neural development and nervous system pathologies. To uncover novel cell-type-specific RNA regulators, we systematically investigated expression of RNA recognition motif-containing proteins in the mouse neocortex. Surprisingly, we found RBM20, an alternative splicing regulator associated with dilated cardiomyopathy, to be expressed in cortical parvalbumin interneurons and mitral cells of the olfactory bulb. Genome-wide mapping of RBM20 target mRNAs revealed that neuronal RBM20 binds distal intronic regions. Loss of neuronal RBM20 has only modest impact on alternative splice isoforms but results in a significant reduction in an array of mature mRNAs in the neuronal cytoplasm. This phenotype is particularly pronounced for genes with long introns that encode synaptic proteins. We hypothesize that RBM20 ensures fidelity of pre-mRNA splicing by suppressing non-productive splicing events in long neuronal genes. This work highlights a common requirement of two excitable cell types, cardiomyocytes and neurons, for RBM20-dependent transcriptome regulation.

overall_design: eCLIP-seq, RBM20 WT heart and olfactory bulb, size matched input used as a contorl. All samples libraries were single-end. However OBrep3 and its SMI were sequenced paired-end and only R1 fastq file was used for further analysis.

last: Di Bartolomei

first: Raul

last: Ortiz

first: Dietmar

last: Schreiner

first: Susanne

last: Falkner

first: Esther

middle: E

last: Creemers

first: Peter

last: Scheiffele


title: The dilated cardiomyopathy-associated RNA Binding Motif Protein 20 regulates long pre-mRNAs in neurons [Ribo-TRAP RNA-seq]

overall_design: Comparative gene expression profiling analysis and alternative splicing analysis of RIBO-TRAP RNA-seq data for RBM20 WT and KO mice in parvalbumin positive neurons of the neocortex or vglut positive neurons of the olfactory bulb


title: RNA-Seq analysis Ezh2 knockdown effect on vascular smooth muscle cells

city: Jinan, Shandong

first: Shuai

last: Leng

address: 105 Jiefang Road

institute: Central Hospital Affiliated to Shandong First Medical University

summary: Vascular smooth muscle cells (VSMCs) phenotype switch has been thought to be critical to the development of thoracic aneurysm/dissection. To investigate the function EZH2 in the regulation of VSMCs phenotype switch, we established mouse vascular smooth muscle cells in which each target gene has been knocked down by siRNA.

overall_design: VSMCs isolated and cultured from mouse aorta were siEzh2 or control (n=3 for each group) for 48 hours. Total RNA was isolated using Trizol reagent (Invitrogen, USA). Agilent 2100 Bioanalyzer (Agilent Technologies, USA) was used to evaluate the RNA quality.

first: Wenqian

last: Yu

first: Shishan

last: Xue


title: The dilated cardiomyopathy-associated RNA Binding Motif Protein 20 regulates long pre-mRNAs in neurons


title: gene expression of KYSE510 cells  with treatment of 100 nM PAF for 24 hours

city: Beijing

first: lingyuan

last: zhang

address: 52 Fucheng Road, Haidian District, Beijing, China

institute: BEIJING INSTITUTE FOR CANCER RESEARCH

summary:
KYSE510 cells were treated with 100 nM PAF for 24 hours,and then KYSE510 cells were collected for gene expression analysis using Agilent SurePrint G3 Human Gene Expression v3 (8×60K) Microarray.
Investigation of the gene network of KYSE510 cells regulated by PAF.

overall_design: We designed this experiment to screen for gene whose expressions are effected by PAF treatment.

first: Lingyuan

last: Zhang

first: Jie

last: Chen


title: Histone demethylase KDM5 regulates cardiomyocyte maturation by promoting fatty acid oxidation, oxidative phosphorylation, and myofibrillar organization [CUT&RUN]

status: Public on Dec 15 2023

city: Houston

first: Priyatansh

last: Gurha

state: Texas

address: 6770 Bertner Ave

department: School of Medicine

institute: UTHealth

summary: Rationale: Human pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) provide a platform to identify and characterize factors that regulate the maturation of CMs. The transition from an immature fetal to adult CM state entails coordinated regulation of the expression of genes involved in myofibril formation and OXPHOS among others. Lysine demethylase 5 (KDM5) specifically demethylate H3K4me1/2/3 and have emerged as potential regulators of expression of genes involved in cardiac development and mitochondrial function.Objectives: The purpose of this study is to determine the role of KDM5 in iPSC-CM maturation.Methods and Results:  KDM5A, B, and C proteins were mainly expressed in the early post-natal stages and their expressions were progressively downregulated in the postnatal cardiomyocytes and were absent in adult hearts and CMs. In contrast, KDM5 proteins were persistently expressed in the iPSC-CMs up to 60 days after the induction of myogenic differentiation, consistent with the immaturity of these cells. Inhibition of KDM5 by KDM5-C70 -a pan-KDM5 inhibitor, induced differential expression of 2,372 genes, including upregulation of genes involved in fatty acid oxidation (FAO), OXPHOS, and myogenesis in the iPSC-CMs. Likewise, genome-wide profiling of H3K4me3 binding sites by the CUT&RUN (Cleavage Under Targets & Release Using Nuclease) assay showed enriched of the H3K4me3 peaks at the promoter regions of genes encoding FAO, OXPHOS, and sarcomere proteins. Consistent with the chromatin and gene expression data, KDM5 inhibition increased expression of multiple sarcomere proteins and enhanced myofibrillar organization. Furthermore, inhibition of KDM5 increased H3K4me3 deposits at the promoter region of the ESRRA gene and increased its RNA and protein levels. Knockdown of ESRRA in KDM5-C70-treated iPSC-CM suppressed expression of a subset of the KDM5 targets. In conjunction with changes in the gene expression, KDM5 inhibition increased oxygen consumption rate and contractility in iPSC-CMs.

overall_design: IPSC-CM and treated group were used for  CUT and RUN Assay. The latter utilized an IgG (control antobody) and a H3K4me3 antibodies

first: Manisha

last: Deogharia

first: A

last: Marian


title: Sequence reliance of a Drosophila context-dependent transcription factor

city: Atlanta

first: Leila

last: Rieder

state: GA

address: 1510 Clifton Rd

department: Biology

institute: Emory University

summary: Despite binding similarciselements in multiple locations, transcription factors often perform context-dependent functions at different loci. How factors integratecissequence and genomic context is still poorly understood and has implications for off-target effects in genetic engineering. TheDrosophilacontext-dependent transcription factor CLAMP targets similar GA-richciselements on the X-chromosome and at the histone gene locus but recruits very different, loci-specific factors. We discover that CLAMP leverages information from bothciselement and local sequence to perform context-specific functions. Our observations imply the importance of other cues, including protein-protein interactions and the presence of additional cofactors.

overall_design: ChIP-seq for Mxc in D. melanogaster embryos.

first: Lauren

last: Hodkinson


title: Comprehensive Analysis of Differentially Expressed LncRNAs associated with Lipid Metabolism in Patients with Colorectal Cance

city: hangzhou

first: Rui

last: Zhu

address: yucai bei road

institute: xiaoshan hospital

summary: Colorectal cancer (CRC) is one of the most common cancers in the world, and its causes are related to diet and obesity. Currently, crosstalk between fat metabolism and CRC has been reported, but the specific mechanism of crosstalk remains to be further studied. In this study, we screened Differentially Expressed lncRNAs and mRNAs from primary cancer tissue, paracancer tissue and white adipose tissue of patients with CRC.Then, by screening and analyzing the genes which differed between primary and paracancer tissues, as well as between paracancer tissues and white adipose tissue, but not between primary and white adipose tissue, we found that lncRNA (MIR503HG) may be involved in the cross-talk between CRC and lipid metabolism through exosome delivery. Although our sample were limited, this study raises the possibility of cross-talk between lipid metabolism and CRC through exosome delivery of lncRNAs.

overall_design: In this study, high-throughput LncRNA sequencing assay was used to analyze human colorectal cancer tissue, para-cancer tissue and para-cancer mesenteric adipose tissue of 10 patients. we conducted the comprehensive screening and analysis of LncRNA associated with lipid metabolism in human colorectal cancer based on bioinformatics, in order to investigate the differentially expressed LncRNA related to adipose metabolism and analyze the mechanism of their involvement in colorectal cancer.

first: Xiufeng

first: Fei

last: yang

first: Zhong

last: shen


title: Absence of Anxa1 in myeloid lineage cells exacerbates acute pancreatitis (AP)

city: Fuzhou

first: Changgan

state: Fujian

address: #29 Xinquan Road, Fuzhou 350001, People’s Republic of China

institute: Fujian Medical University Union Hospital

summary: To elucidate the role of macrophage Anxa1 in the pathogenesis of AP, we intercrossed Anxa1flox/flox (Anxa1f/f or Anxa1 WT, as control) with Lyz2-Cre mice to create myeloid-specific Anxa1-knockout (Anxa1CKO) mice. We induced AP with repeated injection of cerulein in Anxa1CKO and  Anxa1f/f littermates and examined the pancreas injury and inflammation. Pancreas tissue samples were collected at 12 hours after the first cerulein injection and performed RNA-seq.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for Anxa1 CKO and  Anxa1 WT AP mice.

first: YU

last: PAN

first: Zelin

last: Hou


title: The Rv2166c Transcriptional Regulator's On/Off Switch Modulates Mycobacteria Cell Wall Biosynthesis and Cell Division, Enhancing Environmental Stress Resistance

city: Chongqing

first: Hongxiang

last: Xu

address: Tiansheng Road

institute: Southwest University

summary: Background: Mtb's cell wall comprises peptidoglycan, arabinogalactan, and mycolic acids linked to capsule proteins and polysaccharides by noncovalent bonds. Cell division requires extensive remodeling by inserting cell wall-building subunits, which require multiple enzymes to ensure precision and accuracy during the addition and conjugation of biomolecules to the cell wall. Approximately 35% of division and cell wall cluster operon genes are involved in cell wall biosynthesis, 22% in cell division, and 43% are still unstudied. Results: Rv2166c was examined in M. smegmatis, a substitute for M. tuberculosis, using three strains: CRISPR-Cas12a-knockout ∆Ms_4236, complemented ∆Ms_4236::Rv2166c, and pALACE transformed Ms_Vec were grown at 370C in 7H9 or 7H10 to evaluate phenotypic, chemical stress, and antibiotic responses in normoxia and hypoxia. We also examined the whole transcriptome to identify genes associated with MSMEG_4236 deletion of the Rv2166c homolog under normoxia and hypoxia. Deletion and overexpression of Rv2166c affect mycobacterial biofilm formation, cell elongation, colony morphology, and sliding motility in normoxia but are alleviated in hypoxia. Overexpression showed resistance to nucleic acid-target antibiotics, but sensitivity to cell wall-target drugs, yet long-term expression arrests growth. According to transcriptome investigation, the deletion of MSMEG_4236 upregulated all division and cell wall cluster operon genes, several mycolic acids and arabinogalactan biosynthesis genes and downregulated numerous promoters outside of the dcw cluster operon through binding at AAAGTG[G/T] sequence motifs. Conclusion: This study shows that the mycobacterial Rv2166c represses the division and cell wall cluster operon as a transcriptional regulator. Thus, modulating this gene's expression affects many mycobacteria phenotypes and environmental stress resistance. Several AAAGTG[G/T] motifs containing promoters have been found, demonstrating that Rv2166c regulates genes other than its operon. These data suggest that the Rv2166c can be targeted as future antituberculosis medicines.

overall_design: The M. smegmatis strains grew in 7H9 media under normoxia and hypoxia for three days Then, the bacteria were harvested and washed with 1x PBS, and pellets were preserved through liquid nitrogen and kept at -80°C. Then, the samples were quality-checked, and sequence alignment analysis was performed to evaluate the ribosomal RNA contamination rate and to confirm transcriptome quality through sequence saturation and gene coverage. Samples titled with WT,H,K,KH represented the normoxia condition for the WT strain, hypoxia conditionfor the WT strain, normoxia condition for the ∆Ms_4236 strain, hypoxia condition for the ∆Ms_4236 strain, respectively.


title: DmCFL KD in Drosophila musculature of late third instar larvae

city: New York

first: Mary

last: Baylies

state: New York

institute: Memorial Sloan Kettering Cancer Center

summary: To investigate the effect of reducing DmCFL (twinstar) exclusively in Drosophila body wall muscle

overall_design: Performed gene expression profiling analysis using data obtained from RNA-seq of 2 experimental groups (tsrRNAi vs mCherryRNAi), with 3 samples each. Each sample contains the transcriptome of 7-10 Drosophila larva muscle enriched lysates.

first: Briana

last: Christophers

middle: K


title: Individual Variability in Human Cell Type Transcriptomes and Epigenomes

city: La Jolla

first: Joseph

last: Ecker

state: CA

address: 10010 North Torrey Pines Road

department: Genomic Analysis Laboratory

institute: HHMI-Salk-Institute

summary: Diversity and individual variability are essential to human cognitive function. Identifying the conserved and variable (epi)genomic signatures of the brain’s cellular components is critical for understanding the neurobiological basis of individual variation in brain function.  We applied single nucleus methylome and transcriptome sequence (snmCT-seq) to neurons from the frontal cortex of 11 adult human donors spanning a range of ages from 23 to 74, including males and females (Broadmann Area BA46). We clustered cells into brain cell types based on methylation features. We then examined the transcriptome and epigenome features in each cell type between and within individual donors. Taking advantage of the multimodal measurements in single cells, we also identified the relation between RNA expression and methylation level.These data with multiomics measurement from donors with sex and age diversity aims to approach the dimension of inter-individual variability.

overall_design:
We apply snmCT-seq to identify the transcriptomic and epigenomic features of neurons from individual adult human frontal cortex, including 3 aged male donors (age range 70-71), 3 aged female donors (71-74 years old), 3 young male donors  (25 years old) and 2 young female donors  (23-30 years old). For each donor, 2 chunks of brain tissue were collected and processed separately, to assess within-donor variability.

This project includes the following series:
https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE193274
https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE193296
https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE193299
https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE193313
https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE193339
https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE193372
https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE193458
https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE193499
https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE201248
https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE201830
https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE201910
https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE201933
https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE202033
https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE202062
https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE202125
https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE202162
https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE249238
https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE249336
https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE249399
https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE249705

first: Jofan

last: Chien

first: Anna

last: Bartlett

first: Hanqing

last: Liu

first: Rosa

last: Castanon

last: Nery

first: Bang-An

last: Wang

first: Nick

last: Johnson

first: Julia

last: Osteen

first: M

middle: M

last: Behrens

first: Eran

last: Mukamel


title: Transcriptional analysis of TREM-1+ and TREM-1- airway eosinophils during allergic inflammation

city: IOWA CITY

last: Klesney-Tait

state: Iowa

address: 451 Newton Rd

department: Internal Medicine

institute: University of Iowa

summary: Triggering Receptor Expressed on Myeloid cells 1 (TREM-1) an innate receptor that canonically amplifies inflammatory signaling in neutrophils and monocytes, plays a central role in regulating lung inflammation. Utilizing a murine model of asthma, flow cytometry revealed TREM-1+ eosinophils in the lung tissue and airway during allergic airway inflammation. TREM-1 expression was restricted to recruited, inflammatory eosinophils. Expression was induced on bone marrow derived eosinophils by incubation with IL-33, LPS, or GM-CSF. Compared to TREM-1- airway eosinophils, TREM-1+ eosinophils were enriched for pro-inflammatory gene sets including migration, respiratory burst, and cytokine production. Unexpectedly, eosinophil-specific ablation of TREM-1 increased airway IL-5 and lung tissue eosinophil accumulation. Further investigation of transcriptional data revealed apoptosis related gene sets were enriched in TREM-1+ eosinophils. Annexin V staining demonstrated higher rates of apoptosis among TREM-1+ eosinophils compared to TREM-1- eosinophils in the inflammatory airway. In vitro, Trem1/3-/- eosinophils were protected from apoptosis.  Finally, inhibition of reactive oxygen species production with diphenyleneiodonium protected WT bone marrow derived eosinophils from apoptosis more than Trem1/3-/- eosinophils, suggesting that superoxide accounted for more apoptosis in WT cells. These data demonstrate protein level expression of TREM-1 by eosinophils for the first time, define a population of TREM-1+ inflammatory eosinophils, and reveal that eosinophil TREM-1 restricts key features of type 2 lung inflammation.

overall_design: Paired eosinophil  (SSChi CD45+ CD11b+ CD11clo/mid Ly6Glo/mid Siglec-F+) samples were collected from pooled lung lavage WT mice treated to induce allergic airway inflammation . Two samples were sorted from each lavage samples: TREM-1+ denoted by "T" and TREM-1- denoted by "N".  Each sample includes a source number ie "101" and TREM status ie "T" which combined would be "101T."

first: Jayden

last: Bowen

first: Kathy

last: Keck

first: Andrew

last: Thurman

first: Alejandro

last: Pezzulo


title: Transcriptomic response to Klotho deficiency in the renal distal convolution

city: Zurich

first: Ganesh

last: Pathare

state: Zürich

address: Winterthurerstrasse 190

department: Institute of Anatomy

institute: University of Zurich

summary: To assess the transcriptomic response to Klotho deficiency in the renal distal convolution (DC), we isolated DC cells from control and DC-specific Klotho KO mice using COPAS (Complex Object Parametric Analyzer and Sorter). Further comprehensive and unbiased RNA-seq identified altered transcripts associated with canonical MAPK pathway, as well as many novel targets of Klotho.

overall_design: We included 4 male mice in each control and KO group. The DC cells were isolated using COPAS. The transcriptomic repsonse was assesed by RNA-seq


title: Multivalent state transitions shape the intratumoral composition of small cell carcinoma II

city: Chicago

first: Mohamed

last: Abazeed

state: IL

address: 303 E Superior St Lurie

department: Radiation Oncology

institute: Northwestern University

summary: Analysis of small cell lung carcinoma human-derived xenografts. The hypothesis tested in the present study was that small cell lung carcinomas demonstrate transcriptomic diversity. Results provide important information on the variation and groupings (clustering) of distinct small cell lung carcinomas.

overall_design:
RNA obtained from small cell lung carcinoma tissue derived from a human biospy or surgical specimen then passaged through NSG mice.
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Authors state "We have submitted only the processed files and the associated metadatasheet as the raw files will have to be uploaded into dBGaP."

first: Gopal

last: Priyanka


title: Hepatic fibroblast growth factor 21 is required for two polyphenols in exerting their metabolic beneficial effect in mice

city: Toronto

first: Tianru

last: Jin

state: Ontario

address: 101 college street

department: Advanced Diagnostics

country: Canada

institute: University Health Network

summary: Recent studies have shown that FGF21 is a common target for dietary polyphenol intervention and nutritional restriction. FGF21 is also a newly recognized target of GLP-1R agonists (GLP-1RAs). Here we ask whether hepatic FGF21 is required for dietary polyphenols in exerting their metabolic beneficial effects. Liver-specific FGF21 null mice (lFgf21-/-) were utilized in current study. We found that on chow diet, no appreciable defect on glucose disposal was observed in male or female lFgf21-/- mice, while fat tolerance was impaired in male but not female lFgf21-/- mice, associated with elevated serum TG level, reduced hepatic expression of Ehhadh and Ppargc1. On high-fat-high-fructose (HFHF) diet challenge, Fgf21fl/fl mice but not lFgf21-/- mice exhibited response to curcumin intervention on reducing body weight and serum TG, and on improving fat tolerance. Resveratrol intervention also affected hepatic FGF21 expression and its downstream effectors. Metabolic beneficial effects of resveratrol intervention observed in HFHF diet challenged Fgf21fl/fl mice were either absent or attenuated in lFgf21-/- mice. Thus, hepatic FGF21 is required for curcumin and resveratrol in exerting their metabolic beneficial effect. Recognition that FGF21 is the common target of GLP-1RAs and dietary intervention brings us a novel angle in studying metabolic disease treatment and prevention.

overall_design: To investigate the beneficial effects of resveratrol in high fat high fructose (HFHF) diet-challenged mice, we fed male mice with normal chow diet (LFD), HFHF diet, and HFHF diet  with Resveratrol for 16 weeks. Liver was collected at the end of the experiment and subjected to RNA isolation.

first: Weijuan

last: Shao

first: Jia

last: Feng


title: Single-Cell RNA-Seq Reveals Collagen VI Antibody-Induced M2-Like Macrophages Expressing Lyve1 Reduce Atherosclerotic Plaque Area in Apoe-/- Mice

city: Guangzhou

first: Duan

last: Rui

address: No. 1023-1063, Shatai South Road, Baiyun District, Guangzhou

institute: Southern Medical University

summary: The macrophage subpopulations that have therapeutic benefits against atherosclerosis have not yet been fully understood. A single-cell RNA sequencing analysis were performed on the F4/80+ macrophages of apolipoprotein E-deficient (Apoe−/−) mice on a normal diet (ND), a high-fat diet (HFD), and a high-fat diet (HFD) with anti-CVI-mAb treatment. We analyze the diversity of M2 macrophages in atherosclerosis using single cell RNA sequencing (scRNA-seq).

overall_design: Four-week-old male Apoe−/− mice with C57BL/6 background were fed a high-fat diet (HFD; 21 % fat, 0.15 % cholesterol) for 20 weeks. The diet was changed to a normal diet one week before the first immunization. At week 21, the mice in the anti-CVI-mAb and HFD groups received intraperitoneal injections of anti-CVI-mAb (3 mg per 200 µL) and PBS (200 µL). C57BL/6 and Apoe-/- fed normal diet (intraperitoneal injections equal volume PBS) were control. Peritoneal cells were isolated from each group. The cell suspensions obtained individually from the peritoneal cavity were passed through a 100 µm cell strainer and were stained with anti-mouse F4/80-FITC. The total viable F4/80+ cells were selected using Beckman Coulter’s MoFlo Astrios EQ for scRNA-sequencing.

first: Rui

last: Duan

first: Yan

last: Liu

first: Dongmei

last: Tang

first: Sujun

last: Xiao

first: Run

last: Lin


title: Multifaceted SOX2-chromatin interaction underpins pluripotency progression in early embryos

first: Lijia

last: Li

address: Haidian District

department: School of Life Science

institute: Tsinghua University

summary: Pioneer transcription factors (TFs), such as OCT4 and SOX2, play crucial roles in pluripotency regulation. However, the master TF-governed pluripotency regulatory circuitry was largely inferred from cultured cells. In this work, we investigated SOX2 binding from embryonic day 3.5 (E3.5) to E7.5 in the mouse. In E3.5 inner cell mass (ICM), SOX2 regulates the ICM-trophectoderm program but is dispensable for opening global enhancers. Instead, SOX2 occupies preaccessible enhancers in part opened by early-stage expressing TFs TFAP2C and NR5A2. SOX2 then redistributes when cells adopt naive and formative pluripotency by opening enhancers or poising them for further rapid activation. Hence, multifaceted pioneer TF–enhancer interaction underpins pluripotency progression in embryos, including a state in E3.5 ICM that bridges totipotency and pluripotency.

overall_design: By employing CUT&RUN,ATAC-seq, ChIP-seq and RNA-seq, we systematically examined the genome wide SOX2 cromatin binding an function in mouse erly embryos and  mESC cell lines and differentiated cells.

first: Fangnong

last: Lai

first: Wei

last: Xie


title: Temporal-specific single-cell atlas of human type A aortic dissection

city: beijing

first: Junjie

last: Luo

address: 2 West Yuanmingyuan Road, Haidian District

institute: China Agricultural University

summary: Stanford type A aortic dissection (STAAD) is an aortic degenerative remodeling disease carrying an exceedingly high mortality worldwide. The irreversible weakening, dilatation and dissection of the ascending aorta contributes to circulatory failure and premature death. Dissections manifest distinct patterns in both clinical presentations and histophathological features during disease temporal transition. Therefore, we delineate a full repertoire of cellular landscape and dynamic interplay in distinct temporal patterns of STAAD using single-cell transcriptomics for comprehensively understanding of disease features and temporal evolution. We performed single-cell RNA sequencing with ascending aortic tissues from 14 participants, including 9 patients with STAAD (4 acute, 3 subacute, and 2 chronic) and 5 control subjects. Unsupervised clustering was performed based on transcriptional profiles of highly variable genes. Single T cell analysis by RNA sequencing and TCR tracking (STARTRACT) was performed to quantitatively delineate the developmental trajectory of aortic T lymphocytes. On the basis of molecular signatures and distinct disease patterns, we draw a spatiotemporal map of disease subtype-specific alterations and complex interplay among cell types. The transcriptional profiles of 93,397 individual cells enable us to identify 12 major cell types and 37 subtypes in human ascending aorta. Comparisons of transcriptomes of disease subtypes reveal striking stage-specific cellular diversity and transcriptional dynamics in phenotypic switch, vascular inflammation, cell death and survival, extracellular matrix remodeling, and cell-cell interactions during disease temporal transition. Modeling of developmental trajectory revealed that the defects in mitochondrial OXPHOS drives phenotypic switch of contractile smooth muscle cells to synthetic myofibroblasts through AP-1 transcriptional complex. STARTRACT analysis revealed that CD8+ HSP are preferentially enriched and potentially clonally expanded in acute AD. Intersection of disease risk genes with our dataset reveals distinct gene panel in discriminating temporal-specifc AD. This compendium of transcriptome data provides valuable insights and a rich resource for understanding the cellular diversity and heterogeneity in human aorta. The distinct alterations of cell type- and disease subtype-specific spatiotemporal features will ultimately facilitate the design of more precisely tailored anti-AD therapies.

overall_design: single-cell prolifes of vascular cells from 14 human ascending aorta

first: Yongting

first: Peng

last: An

first: Fazheng

last: Ren

first: Hongjia

last: Zhang


title: Gene expression by RNA-seq in Liver of WT and panetch cell specific  P55 KO mice 18 hours after TNF

city: Gent

first: Steven

last: Timmermans

address: Technologiepark 71

country: Belgium

institute: VIB-UGent

summary: After a challenge with TNF, gene expression changes in the liver may result from direct effect of TNF, through liver TNF receptors, and indirect effect. Baterial infiltration of the liver due to leaky gut after TNF is a primary contributor of this effect. In order to evaluate only the direct effects of TNF on lier transcriptome, we performed TNF challenge in normal WT mice and P55 panetch cell KO mice, which do not show the leaky gut syndrome.

overall_design: To investigate the direct effects of TNF on liver gene expression, an RNA-seq was performed 18 hours after TNF or PBS (control) injection in P55 KO mice and P55 WT littermates (4 mice/group except KO+PBS: 3 mice/group)

first: Charlotte

last: Wallaeys

first: Natalia

last: Garcia-Gonzalez

first: Jolien

last: Vandewalle

first: Tineke

last: Vanderhaeghen

first: Hester

last: Dufoo

first: Melanie

last: Eggermont

first: Riet

last: De Rycke

first: Serge

last: Verbanck

first: George

last: Kollias

first: Richard

middle: S

last: Blumberg

first: Takao

last: Iwawaki

first: Claude

last: Libert


title: Inhibition of METTL3 attenuates renal injury and inflammation by alleviating TAB3 m6A modifications via IGF2BP2-dependent mechanisms

city: Hefei

first: Xiao ming

last: Ment

address: No. 81, Meishan Road, Shushan District, Hefei City, Anhui Province

institute: Anhui Medical University

summary: The role of N6-methyladenosine (m6A) modifications in renal diseases is largely unknown. Here, we characterised the role of N6-adenosine-methyltransferase METTL3, whose expression is elevated in renal tubules in different acute kidney injury (AKI) models as well as in human biopsies and cultured tubular epithelial cells (TECs). METTL3 silencing alleviated renal inflammation and programmed cell death in TECs in response to stimulation by tumor necrosis factor alpha (TNF-α), cisplatin, and lipopolysaccharide (LPS), whereas METTL3 overexpression had the opposite effects. Conditional knockout (cKO) of METTL3 from mouse kidneys attenuated cisplatin- and ischemic/reperfusion (I/R)-induced renal dysfunction, injury, and inflammation. Moreover, TAB3 was identified as a target of METTL3 by m6A methylated RNA immunoprecipitation sequencing and RNA sequencing. The stability of TAB3 was increased through binding of IGF2BP2 to its m6A-modified stop codon regions. The pro-inflammatory effects of TAB3 were then explored both in vivo and in vitro. Adeno-associated virus 9 (AAV9)-mediated METTL3 silencing attenuated renal injury and inflammation in cisplatin- and LPS-induced AKI mouse models. We further identified Cpd-564 as a METTL3 inhibitor that had better protective effects against cisplatin- and I/R-induced renal injury and inflammation than S-adenosyl-L-homocysteine, a previously-identified METTL3 inhibitor. Collectively, METTL3 promoted m6A modification of TAB3 and enhanced its stability via IGF2BP2-dependent mechanisms. Both pharmacological and genetic inhibition of METTL3 attenuated renal injury and inflammation, suggesting that the METTL3/TAB3 axis is a potential target for treatment of AKI.

overall_design: To identify specific targets of METTL3 in renal inflammation, we conducted RNA-seq and MeRIP-seq analyses in TNF-α-stimulated HK2 cells with or without silencing of METTL3

first: Jia-nan

last: Wang

first: Xiao-ming

last: Meng


title: 18S rRNA methyltransferase EMG1 non-enzymatically influence m6A in mRNA

city: philadelphia

first: Kathy

last: Liu

state: PA

address: 415 curie blvd

institute: University of Pennsylvania

summary: Defects in ribosome biogenesis can trigger a battery of cellular events to alter gene expression and cell growth. 18S rRNA methyltransferase EMG1 is an important assembly factor in ribosome biogenesis. Here, we report that downregulation of EMG1 non-enzymatically leads to decreased N6-methyladensine (m6A) levels in polyadenylated RNA. We revealed the identities of the messenger RNAs with decreased m6A intensities upon EMG1 deficiency using m6A-IP-seq. This group of mRNAs significantly overlap with the CLIP targets of IGF2BP3, which is an m6A reader protein stabilizing m6A-containing transcripts. Furthermore, both RNA-seq and qRT-PCR results suggest that the expression level of this group of mRNAs significantly decreased upon EMG1 deficiency, possibly through the regulation of IGF2BP3. Gene ontology analysis suggests that this group of mRNAs function in a list of cell signaling pathways important for cell growth. These findings collectively suggest that decreased ribosome assembly may influence gene expression via decreasing m6A levels in mRNAs.

overall_design: m6AIP-seq after knocking down Emg1 in N2a cells

middle: F

first: Heidi

last: Elashal

first: Hui

last: Shen


title: Carboxypeptidase M modulates BMSCs osteogenesis-adipogenesis via the MAPK/ERK pathway: An integrated single-cell and bulk transcriptomic study

status: Public on Dec 14 2023

first: Zheting

last: Liao

address: 1838 North Guangzhou Avenue

institute: Sourthern Medical University

summary: The pathogenesis of osteoporosis (OP) is closely associated with the disrupted balance between osteogenesis and adipogenesis in bone marrow-derived mesenchymal stem cells (BMSCs). We analyzed published single-cell RNA sequencing (scRNA-seq) data to dissect the transcriptomic profiles of bone marrow-derived cells in OP, reviewing 56,377 cells across eight scRNA-seq datasets from femoral heads (osteoporosis or osteopenia n=5, osteoarthritis n=3). Seventeen genes, including carboxypeptidase M (CPM), were identified as key osteogenesis-adipogenesis regulators through comprehensive gene set enrichment, differential expression, regulon activity, and pseudotime analyses. In vitro, CPM knockdown reduced osteogenesis and promoted adipogenesis in BMSCs, while adenovirus-mediated CPM overexpression had the reverse effects. In vivo, intraosseous injection of CPM-overexpressing BMSCs mitigated bone loss in ovariectomized mice. Integrated scRNA-seq and bulk RNA sequencing analyses provided insight into the MAPK/ERK pathway's role in the CPM-mediated regulation of BMSC osteogenesis and adipogenesis; specifically, CPM overexpression enhanced MAPK/ERK signaling and osteogenesis. In contrast, the ERK1/2 inhibitor binimetinib negated the effects of CPM overexpression. Overall, our findings identify CPM as a pivotal regulator of BMSC differentiation, which provide new clues for the mechanistic study of OP.

overall_design: Total RNA was isolated from human bone marrow derived mesenchymal stem cells (BMSCs) subjected to osteogenic induction and CPM knockdown (siCPM, n=3) or control siRNA (siNC, n=3) on day 7. Samples were then processed for high-throughput sequencing on the NovaSeq 6000 PE150 platform.

first: Liang

last: Zhao


title: Antisense transcription and its roles in response to environmental changes in E. coli K12

city: Charlotte

first: Ehsan

last: Tabari

state: NORTH CAROLINA

address: 9201 University City Blvd

department: Bioinformatics

institute: UNC Charlotte

summary: Antisense RNAs (cis-asRNAs) in prokaryotes are more pervasive than originally thought. However, little is known about their expression patterns and functions. Here we determined transcriptomes and proteomes of E. coli at multiple time points in five culture conditions using directional RNA-seq and tandem mass spectrometry.

overall_design: A frozen stock of Escherichia coli K12 strain MG1655 was thawed, inoculated in LB medium in a test tube by 1:100 dilution and cultured overnight at 37°C and 250 rpm. The cells were then transferred to fresh LB medium in a flask by 1:100 dilutions, and cultured at 37°C and 250 rpm. When the cells grew to an optical density at 600 nm (OD600) of 1.0, they were spun down at 3,200g for 25 min. For the minimal medium MOPS culture (MOPS), the cell pellets were resuspended in the same volume of MOPS medium (100ml of 10X MOPS mixture, 880ml of sterile H2O, 10ml (0.132M) KH2PO4 and 10ml of 20% glucose, Teknova, Hollister, CA). For heat shock treatment (HS), the cell pellets were resuspended in the same volume of MOPS medium, and incubated at 48°C and 250 rpm. For phosphorus-starvation treatment (M-P), the cell pellets were resuspended in the MOPS medium without KH2PO4. For carbon-starvation treatment (M-C), the cell pellets were resuspended in the MOPS medium without glucose. Each culture was done in three replicates in parallel, and an equal volume of cells was collected from each replicate to minimize experimental variations during sampling. Three milliliters of such pooled cell suspension were collected in a test tube containing 1.5ml RNA Later (Invitrogen) immediately after the cell pellets were resuspended in the indicated medium (0 min) and at the indicated time points thereafter (HS: 15min, 30min, 1h, 2hrs and 4hrs; MOPS, M-C and M-P: 1hr, 2hrs, 4hrs, 6hrs; M-C: 1hr, 2hrs, 4hrs, 6hrs). For the LB culture, samples were collected when the cell suspension OD600 reached to 0.5, 1.0 and 3. Cells were spun down at 6,000g for 8 mins (-4°C), and the pellets were resuspended in 1.5 ml of RNAlater. The samples were stored at -80°C until use.

first: Shan

last: Li

first: Xia

last: Dong

first: Harry

last: Lee

first: Sunil

last: Hwang

first: Zhengchang

last: Su


title: Cellular Heterogeneity of Pluripotent Stem Cell Derived Cardiomyocyte Grafts is Mechanistically Linked to Treatable Arrhythmias

city: Brisbane

first: Onkar

middle: Rajesh

last: Mulay

state: QLD

address: Carmody Road

country: Australia

institute: University of Queensland

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

sample_organism:
Homo sapiens
Sus scrofa

platform_organism:
Homo sapiens
Sus scrofa

overall_design: Refer to individual Series


title: Cellular Heterogeneity of Pluripotent Stem Cell Derived Cardiomyocyte Grafts is Mechanistically Linked to Treatable Arrhythmias [Spatial Transcriptomics]

summary: Exciting pre-clinical data have confirmed that human pluripotent stem cell derived cardiomyocytes (PSC-CMs) can remuscularise the injured or diseased heart, with several clinical trials now in planning or recruitment stages worldwide. However, ventricular arrhythmias are a predictable complication following engraftment of intramyocardially injected PSC-CMs. Therefore, there is an urgent unmet need to gain mechanistic insights and treatment strategies to control or prevent these engraftment arrhythmias (EAs). We used a porcine model of myocardial infarction and PSC-CM transplantation to investigate efficacy of pharmacologic and catheter based anti-arrhythmic strategies in mitigating EAs. Furthermore, cell doses were robustly phenotyped using single cell ribonucleic acid sequencing and high parameter flow cytometry to identify cellular characteristics predictive of arrhythmogenesis. Combination therapy with amiodarone and ivabradine significantly reduced EA rate and burden following PSC-CM transplantation. Catheter ablation was also a feasible and effective treatment strategy which could be considered in the case of pharmacologically refractory arrhythmias. In addition, we show that EAs are mechanistically linked to cellular heterogeneity in the input PSC-CM and resultant graft. Specifically, we identify atrial and pacemaker-like cardiomyocytes as culprit arrhythmogenic subpopulations. We further describe two unique surface marker signatures, SIRPA+/CD90-/CD200+ and SIRPA+/CD90-/CD200-, which identify arrhythmogenic and non-arrhythmogenic cardiomyocytes respectively. Our data deepens mechanistic understanding of EAs and suggests that modifications to current PSC-CM production and/or selection protocols could ameliorate this problem. We further show that current clinical pharmacologic and interventional anti-arrhythmic strategies can control and potentially abolish these arrhythmias, an important safety consideration given several impending clinical trials.

overall_design: Spatial transcriptomics analysis of engrafted PSC-CMs. Using a tissue microarray punch, the tissue samples of 3mm diameter were collected from regions confirmed to contain PSC-CM grafts.

first: Quan

last: Nguyen

first: Guiyan

last: Ni


title: Cellular Heterogeneity of Pluripotent Stem Cell Derived Cardiomyocyte Grafts is Mechanistically Linked to Treatable Arrhythmias [scRNA-Seq]

overall_design: Frozen day15 PSCs derived cardiomyocytes were thawed, hashtagged, and pooled for scRNA-seq.

first: Nathan

last: Palpant

first: Zixuan

last: Wu


title: A transcriptomic taxonomy of mouse brain-wide spinal projecting neurons (10X)

city: Boston

first: Yu

last: Zhang

state: MA

address: 3 Blackfan Street, Room 13060

department: Neurobiology

institute: Boston Childrens Hospital

summary: The brain executes control of nearly all bodily functions via spinal projecting neurons (SPNs) that carry command signals from numerous supraspinal regions to the spinal cord. Despite their physiological and clinical relevance, a comprehensive molecular characterization of SPNs is still lacking. Here, we use retrograde labeling, whole-brain imaging, and high-throughput transcriptional profiling to generate a unified brain-wide anatomic and transcriptomic atlas of adult mouse SPNs at single-cell resolution. We transcriptionally profiled a total of 65,002 SPNs, identified 76 region-specific SPN types, and mapped these types into a companion atlas of the whole mouse brain generated by the Allen Institute for Brain Science within the BRAIN Initiative Cell Census Network (BICCN). This SPN taxonomy reveals a three-component organization of SPNs: (1) molecularly homogeneous excitatory SPNs from the cortex, red nucleus, and cerebellum with somatotopic spinal terminations suitable for point-to-point communication; (2) highly heterogeneous excitatory and inhibitory populations in the reticular formation with broad spinal termination patterns, thus suitable for relaying commands related to the activities of the entire spinal cord; and (3) modulatory neurons expressing slow-acting neurotransmitters and/or neuropeptides in the hypothalamus, midbrain, and reticular formation for gain control of brain-spinal signals. Within each of these components, this atlas revealed additional insights. From components (2) and (3), we discovered a LIM homeobox transcription factor code that parcellates the most transcriptionally complex population, reticulospinal neurons, into five molecularly distinct and spatially segregated populations. For neurons in component (1), we found transcriptional signatures of a subset of SPNs with large soma size and correlated these with fast-firing electrophysiological properties; thus, at least two different cable lines (namely, Pvalb/Kcng4/Spp1 positive and negative) with different electrophysiological properties might underlie the transmission of brain signals to the spinal cord. Together, by integrating the anatomy, molecular identity, and physiological properties, this study establishes a comprehensive taxonomy of brain-wide SPNs and provides insight into the functional organization of SPNs in mediating brain control of bodily functions.

overall_design: Spinal projecting neuron (SPN) nuclei of C57Bl6/J mice were retrogradely labeled via injection of recombinant retrograde adeno associated virus (rAAV2/retro-Syn-H2B-fluorescent protein) into multiple segments centering the cervical (GFP) and lumbar (mScarlet) spinal cord of postnatal day 42 C57BL/6J mice. At postnatal day 56, the mice were sacrificed and nuclei were isolated via tissue dissection and dissociation of brain regions containing SPNs (i.e., rostral forelimb area cortex [RFA], primary and secondary motor cortex / primary somatosensory cortex [M1M2S1], secondary somatosensory cortex [S2], hypothalamus [HY], midbrain [MB], cerebellum [CB], pons [PONS], and medulla [MED]). Subsequently, SPNs were enriched via fluorescence activated nucleus sorting. Nuclei were then analyzed with single nucleus RNA sequencing using the 10x genomics and SmartSeq platforms.

first: Carla

middle: C

last: Winter

first: Anne

last: Jacobi

first: Junfeng

first: Leeyup

last: Chung

first: Cindy

last: van Velthoven

first: Zizhen

last: Yao

first: Changkyu

first: Zicong

first: Shuguang

last: Yu

first: Kun

last: Gao

first: Geraldine

last: Salazar

first: Evgenii

last: Kegeles

first: Makenzie

last: Tomihiro

first: Yiming

first: Zhiyun

last: Yang

first: Junjie

last: Zhu

first: Jing

last: Tang

first: Xuan

last: Song

first: Ryan

last: Donahue

first: Qing

last: Wang

first: Delissa

last: McMillen

first: Michael

last: Kunst

first: Ning

first: Kimberly

last: Smith

first: Gabriel

last: Romero

first: Michelle

last: Frank

first: Alexandra

last: Krol

first: Riki

last: Kawaguchi

first: Daniel

middle: H

last: Geschwind

first: Guoping

last: Feng

first: Lisa

middle: V

last: Goodrich

first: Yuanyuan

first: Bosiljka

last: Tasic

first: Hongkui

last: Zeng

first: Zhigang

last: He


title: Individual Variability in Human Cell Type Transcriptomes and Epigenomes [OM3B]

first: Joseph

last: Ecker

summary: Diversity and individual variability are essential to human cognitive function. Identifying the conserved and variable (epi)genomic signatures of the brain’s cellular components is critical for understanding the neurobiological basis of individual variation in brain function.  We applied single nucleus methylome and transcriptome sequence (snmCT-seq) to neurons from the frontal cortex of 11 adult human donors spanning a range of ages from 23 to 74, including males and females (Broadmann Area BA46). We clustered cells into brain cell types based on methylation features. We then examined the transcriptome and epigenome features in each cell type between and within individual donors. Taking advantage of the multimodal measurements in single cells, we also identified the relation between RNA expression and methylation level.These data with multiomics measurement from donors with sex and age diversity aims to approach the dimension of inter-individual variability.

overall_design: We apply snmCT-seq to identify the transcriptomic and epigenomic features of neurons from individual adult human frontal cortex, including 3 aged male donors (age range 70-71), 3 aged female donors (71-74 years old), 3 young male donors  (25 years old) and 2 young female donors  (23-30 years old). For each donor, 2 chunks of brain tissue were collected and processed separately, to assess within-donor variability. This sample is from chunk A of a 73 years old female.

first: Jofan

last: Chien

first: Anna

last: Bartlett

first: Hanqing

last: Liu

first: Rosa

last: Castanon

last: Nery

first: Bang-An

first: Nick

last: Johnson

first: Julia

last: Osteen

first: M

last: Behrens

first: Eran

middle: A

last: Mukamel


title: Versatile toolkit for high-efficiency and scarless overexpression of circular RNAs

city: Adelaide

first: Simon

last: Conn

state: South Australia

address: Sturt Road, Bedford Park

department: College of Medicine and Public Health

institute: Flinders University

summary: Circular RNAs (circRNAs) are a class of ubiquitously expressed, single-stranded, covalently covalently-closed (i.e. circularised) RNA that contain a unique nucleotide sequence created by the ligation of their 5' and 3' ends, called the back-splice junction. Understanding the cellular roles of circRNAs involves, in part, investigating the effects on cell phenotype of increased expression of individual circRNAs. This is frequently done by transfecting cells with plasmid DNA containing cloned exons from which the circRNA is transcribed, flanked by sequences that promote circularisation. We observed that all commonly used plasmids we tested unexpectedly incorporated molecular scars comprising vector sequence vector sequence as a molecular scar into the circRNA back-splice junction upon circularisation. Stepwise redesign of the cloning vector corrected this problem, ensuring bona fide circRNAs are produced with their natural back-splice junction at high efficiency. The fidelity of circRNAs produced from this new construct was validated by RNA sequencing. To increase the utility of this modified resource for expressing circRNA, we developed an expanded set of vectors incorporating this design that enable selection with a variety of antibiotics and fluorescent proteins, a range of promoters varying in promoter strength and plus a complementary set of lentiviral plasmids for difficult-to-transfect cells. These resources provide a versatile toolkit for high-efficiency and scarless overexpression of circular RNAs that fulfil a critical need for the investigation of circRNA function, including the role of the unique back-splice junction.

overall_design: Human HEK293 cells expressing circRNAs cloned into minicirc (BCREx9_14) and/or mc2 vectors (BCREx9_14, EGFREx2_20, EGFREx19_20, MLLEx9, MLLT2Ex3_4) followed by amplicon seq, single biological replicate.

first: Brett

middle: W

last: Stringer

first: Vanessa

first: Shashikanth

last: Marri


title: Harnessing T-cell exhaustion and trogocytosis to isolate patient-derived tumor-specific TCR

city: Segrate

first: Ivan

last: Merelli

state: Milano

address: via F.lli Cervi 93

department: Istituto di Tecnologie Biomediche

institute: Consiglio Nazionale delle Ricerche

summary: Exhausted Tumor-specific T cells circulate in patients and their specificity can be typed by trogocytosis and Peptidome on chip screening.

overall_design: T cells specific for  tumor antigens were expanded from HSCT-transplanted patients by means of Leukemic Dendridic Cells sequential stimulation or via Dextramer sorting and anti-CD3/anti-CD28 stimulation.

first: Francesco

last: Manfredi

first: Eliana

last: Ruggiero

first: Chiara

last: Bonini


title: TIGIT contributes to the regulation of 4-1BB and does not define NK cell dysfunction in glioblastoma

city: West Lafayette

first: Sagar

last: Utturkar

state: IN

address: 201 S University St

department: Institute for Cancer Research

institute: Purdue University

summary: This study evaluates the genotype of NK cells sorted for expression of TIGIT into TIGIThigh, TIGITmedium and TIGITnegative NK cells. RNAseq was performed on these three NK cell populations.

overall_design: Human NK cells isolated from PBMCs were sorted as TIGIThigh, TIGITmedium and TIGITnegative, and total RNA was isolated using the mirVana miRNA Isolation Kit (Thermo Fisher Scientific, Waltham, MA) according to manufacturer instructions, with >700 ng of RNA per sample. All samples were checked for quality prior to analysis by Agilent Bioanalyzer, with DV200 > 80 and RIN > 8.4 for all samples. Samples were sent to GeneWiz for library preparation and sequencing. Each sample contained at least 30 million paired-end reads with length of 250 bp. Data quality control (adapter trimming, minimum Phred score 30, minimum read length 50 bp) was performed using fastp 27 (version 0.19.5). Quality trimmed reads were mapped to human reference genome (GRCh38) using the STAR aligner 28 (version 2.7.9a). Reads aligned to each gene feature were counted using the featureCounts 29 program from Subread package (version 1.6.1). Differential expression (DE) analysis was performed using the edgeR method 30 (version 3.32.1) and genes with False Discovery Rate (FDR) ≤ 0.01 were denoted as significant.

first: Kyle

middle: B

last: Lupo

first: Sandra

last: Torregrosa-Allen

first: Bennett

middle: D

last: Elzey

first: Nadia

last: Atallah-Lanman

first: Aaron

last: Cohen-Gadol

first: Veronika

last: Slivova

first: MacKenzie

last: McIntosh

first: Karen

last: Pollok

first: Sandro

last: Matosevic


title: The role of FLASH in EMT plasticity in cancer cells

city: Shreveport

first: Ana-Maria

last: Dragoi

state: Louisiana

address: 1501 Kings Hwy, Molecular and Cellular Physiology Room 4-208

institute: LSU-Health Shreveport

summary: Metastasis consists of sequential steps initiated by cancer cells invading from the primary tumor site into neighboring tissues, followed by entry into the circulatory system and completed by extravasation and growth in distal organs. Circulating tumor cells, thus, encounter and adapt to multiple environmental changes during this transition. Epithelial-to-mesenchymal transition (EMT) is a developmental program that consists of loss of epithelial features concomitant with acquisition of mesenchymal features. Activation of EMT in cancer facilitates acquisition of aggressive traits and cancer invasion. EMT plasticity (EMP), the dynamic transition between multiple hybrid states in which cancer cells display both epithelial and mesenchymal phenotypes, confers survival advantages for cancer cells in the constantly changing environment. Therefore, understanding the molecular mechanisms regulating intermediate phenotypic states along the E–M spectrum is critical. Core EMT transcription factors (EMT-TFs), ZEB, SNAI and TWIST families, play an important role in EMT and its plasticity. In the present study we characterize FLASH as a regulator of EMP and multiple EMT-TFs. We demonstrate that loss of FLASH gives rise to a hybrid E/M phenotype with high epithelial scores even in the presence of TGFβ, as determined by computational methods using expression of predetermined sets of epithelial and mesenchymal genes. We demonstrate that FLASH is regulating expression of multiple cell junction proteins with an established role in cancer progression and that its role in EMT is independent of its histone biogenesis role. Further, we show that FLASH expression in cancer lines is inversely correlated with the epithelial score, consistent with its function as a repressor of the epithelial phenotype. Nonetheless, activation of a distinct set of mesenchymal markers concomitant with epithelial markers reveals the complex role of FLASH in EMT and indicates that intermediate E/M states could arise from opposing control by FLASH on different families of EMT-TFs.

overall_design: To investigate the roles of FLASH, NPAT, and SLBP in EMT, we knocked-down individually the 3 genes in untreated and TGFbeta-treated PANC-1 cells. RNAseq analysis was performed 4 days post-siRNA transfection.


title: Plasma and CSF Proteomic Signatures of Acutely Sleep-Deprived Humans: An Exploratory Study

city: St. Louis

first: Jinsheng

last: Yu

state: MO

address: 660 S. Euclid Ave.

department: Genetics

institute: Washington University School of Medicine

summary: Study objectives: Acute sleep deprivation affects both central and peripheral biological processes. Prior research has mainly focused on specific proteins or biological pathways that are dysregulated in the setting of sustained wakefulness. This pilotexploratory study’s objective wasaimed to provide a comprehensive view of the biological processes and proteins impacted by acute sleep deprivation in both plasma and cerebrospinal fluid (CSF). Methods: We collected plasma and CSF from human participants during one night of sleep deprivation and control normal sleep conditions.  1300 proteins were measured at hour 0 and hour 24 using a high-scale aptamer-based proteomics platform (SOMAomascan) and a systematics biological database tool (Metascape) was used to reveal dysregulated biological pathways.    Results: Acute sleep deprivation lead to opposite effects in plasma and CSF, decreasingdecreased the number of upregulated and downregulated differential protein expression and biological pathways and proteins in plasma but increased upregulated and downregulated protein and biological pathwayssing them in CSF. Predominantly affected pProteins and n pathways were associated with that were predominantly affected by sleep deprivation included immune response, inflammation, phosphorylation, membrane signaling, cell-cell adhesion, and extracellular matrix organization. Conclusions: The identified modification across biofluids adds to evidence that acute sleep deprivation has important impacts on biological pathways and proteins that can negatively affect human health. As a hypothesis-driving study, these findings may help with the exploration of novel mechanisms that mediate sleep loss and associated conditions, drive the discovery of new sleep loss biomarkers, and ultimately aid in the identification of new targets for intervention to human diseases.

overall_design:
Plasma and CSF samples were collected at hour 0 (07:00) and hour 24 (07:00 on the next day) from 5 healthy humans. The same humans were subjected to sleep deprivation and plasma and CSF collection at the same time points with 4-6 months apart from nomral sleep conditions.
The assays are subjected to SOMAscan proprietary oligonucleotide-based aptamer-protein binding and aptamer hybridization onto microarray slides for protein abundance measurement.

first: Ana

last: Vaquer-Alicea

first: Haiyan

last: Liu

first: Brendan

middle: P

last: Lucey