title: Single cell level characterization of injury-induced immune changes in MRL/MpJ knee joints

city: Livermore

first: Stephen

middle: Patrick

last: Wilson

state: CA

address: 7000 East Ave

department: BBTD

institute: LLNL

summary: Post traumatic osteoarthritis (PTOA) is a painful joint disease characterized by the degradation of bone, cartilage, and connective tissues. PTOA is initiated by trauma to joint-stabilizing tissues, such as the anterior cruciate ligament, medial meniscus or by intra-articular fractures. In humans, ~50% of joint injuries progress to chronic PTOA, while the rest spontaneously resolve. To understand molecular programs contributing to PTOA development, we examined injury-induced fluctuations in immune cell populations and transcriptional shifts by single-cell sequencing of synovial joints in PTOA -susceptible C57BL6/J (B6) and -resistant MRL/MpJ (MRL) mice. A potent myeloid driven anti-inflammatory response was discovered in MRL injured joints that significantly contrasted the pro-inflammatory signaling seen in B6 joints. We found Trem2+ macrophages classically described as M2 (CD206+Trem2+) to be enriched in the uninjured joints of MRL mice, and these populations were consistently elevated in this strain, post injury. To evaluate whether Trem2 is essential to macrophage-mediated homeostasis in the joint, we examined Trem2 deficient mice and determined that these mice develop spontaneous OA. Overall, these data suggest that the PTOA resistant MRL mouse strain displays enhanced capacity of clearing debris and apoptotic cells induced by inflammation after injury due to an increase in activated macrophages within the synovial tissue and joint space. Furthermore, we showed that Trem2 is essential for preserving cartilage homeostasis in B6, and we postulate that the elevated levels of Trem2+ joint-resident macrophages in MRL are in part responsible for the PTOA-resistant phenotype in this strain.

overall_design: Right knee joints of 10-week-old MRL/MpJ mice were injured using a tibial compression injury model. Uninjured joints and injured joints from 1-, 3-, 7-, 15- and 30 days post-injury  were collected and immune (Cd45+) cells were analyzed using scRNA-seq.

first: Jillian

last: McCool

first: Aimy

last: Sebastian

first: Gabriela

last: Loots


title: Mutant FUS induces chromatin reorganization in the hippocampus and alters memory processes [ChIP-seq]

status: Public on Dec 31 2024

city: Strasbourg

first: Anne-Laurence

last: Boutillier

address: 12 rue Goethe

department: LNCA

country: France

type: Genome binding/occupancy profiling by high throughput sequencing

summary: This project aims to identify  epigenomic signatures (histone methylation- and acetylation-related) of FUS∆NLS/+ (FUS) vs FUS+/+ (WT) mice, using ChIP-sequencing performed on FACS-sorted nuclei. Cytoplasmic mislocalization of the nuclear Fused in Sarcoma (FUS) protein is associated to amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Cytoplasmic FUS accumulation is recapitulated in the frontal cortex and spinal cord of heterozygous Fus∆NLS/+ mice. We show that in these mice, the hippocampus, a critical structure involved in learning and memory, paradoxically displays nuclear FUS accumulation. FUS binds to a set of genes characterized by the presence of an ETS/ELK-binding motifs, and involved in RNA metabolism, transcription, ribosome/mitochondria and chromatin organization. Importantly, hippocampal nuclei showed a decompaction of the neuronal chromatin at highly expressed genes and an inappropriate transcriptomic response was observed after spatial training of Fus∆NLS/+ mice. Furthermore, these mice lacked precision in hippocampal-dependent spatial memory task. These studies shows that mutated FUS affects epigenetic regulation of the chromatin landscape in hippocampal neurons, which could participate in FTD/ALS pathogenic events.

overall_design: Total hippocampus from 5 month-old FUS∆NSL/+ (FUS) and FUS+/+ (WT) mice were sorted by FACS to isolate neuronal nuclei and about 3-400000 sorted NeuN+ nuclei were then subjected to ChIP-sequencing using antibodies against 3 histone marks:  H3K4me3, H3K27ac and H4K12ac. For this experiment, we have biological replicates (2 independent lots of mice).  We will compare here, the epigenetic profiles of the 3 marks in WT vs FUS mice in NeuN+ FACS-sorted nuclei. We have INPUT DNA For each of the 4 conditions (2 WT and 2 FUS). Overall, this represents 16 samples.


title: m6A RNA suquencing of cholestatic liver fibrosis

city: Taiyuan

first: Tingjuan

last: Huang

address: NO.56 Road Jiefang

country: China

institute: Shanxi Medical University

summary: To investigate the differential gene expression of m6A RNA methylation in cholestatic liver fibrosis, we extablished a mouse model of liver fibrosis by common bile duct ligation.

overall_design: We performed m6A RNA sequencing for BDL-induced cholestatic hepatic fibrosis.

first: Chunhong

last: Zhang

first: Xuewei

last: Li

first: Xiaoling

last: Wang


title: TonEBP increases TRIM37

city: JINJU

first: GU SEOB

last: ROH

address: 15, 816 Beon-gil, Jinju-daero

country: South Korea

institute: Gyeongsang National University

type: Expression profiling by high throughput sequencing

summary: Aging-related microglial activation is associated with dendritic regression and spine loss in the aged brain. However, the microglia-mediated reductions of spine densities by Tonicity-responsive enhancer-binding protein (TonEBP) in the aged brain is yet unknown. We began to address this question by examining the effect of age on microglial activation and the TonEBP expression in mice. by using molecular and morphologic approaches, the roles of TonEBP in microglial activation and dendritic spines were examined in 12-month-old mice and Alzheimer's diseases (AD) mouse models. Here, we found the increased TonEBP in the hippocampus of aged mice and frontal cortex of AD patients. TonEBP haploinsufficiency reduced microglial activation and dendritic spine regression in 12-month aged mice compared to wild-type (WT) mice. We performed electron microscopy to supply interactions with microglial and synapses. We analyzed microglial processes and extracellular space in areas contacting spines and axon terminal, synaptic cleft, among the hippocampus CA1 of 12-month-old TonEBP haploinsufficient mice versus wild-type littermates. Furthermore, in amyloid-ꞵ oligomer (AꞵO)-injected AD mouse model, TonEBP haploinsufficiency inhibited the dendritic spine loss and improved memory deficits in AβO-treated mice compared to WT mice. These findings indicate that TonEBP may play an important role in aging-induced microglial activation and memory deficits.

overall_design: TonEBP (+/-) mice and their Wild-type (WT) (TonEBP+/+) littermates were created by crossing TonEBP (+/-) mice. WT and TonEBP (+/−) mice were back-crossed on the C57BL/6J background for 5 to 6 generations to produce homogeneous animals, with no background effects on phenotypes.


title: ChIP-Seq of mouse mammary gland tissues

city: Macau

first: Lihua

last: Mo

address: Taipa

institute: University of Macau

summary: To sort out the factors that might be negatively regulated by Brca1 but potentiated by ERα in the proliferating mammary epithelial cells, we investigate transcriptional activities of promoters genome-wide using antibodies against H3K4me3, H3K27Ac, Brca1, and estrogen alpha (ERα) by performing chromatin immunoprecipitation (ChIP) sequencing in primary mammary epithelial cells isolated from MTP12 and WTP12 mice

overall_design: ChIP sequence for mammary gland tissues isolated from both Brca1 wild type (3XFlag-Brca1+/+) and mutant mice (Brca1co/co Trp53+/- MMTV-Cre mice) at 12.5 (P12) days of pregnancy by using Flag-Brca1, ERα, H3K4me3, and H3K27Ac.

first: Ragini

last: Adhav


title: Effect of artemisinin on the expression of lung cancer related genes in mice with lung cancer

city: Nanyang

first: Zhenzhen

last: Guo

address: No. 163Wolong Road, Wolong District

department: College of Chemistry and Pharmaceutical Engineering

institute: Nanyang Normal University

summary:
In order to study the mechanism of artemisinin in the treatment of lung cancer, we established a mouse model of lung cancer, treated it with artemisinin by gavage.
Then, we analyzed the gene expression profile of the data obtained from the RNA sequences of the lung tissues of mice in the model group and the artemisinin treated group.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for lung cancer mice (MF1, MF2, and MF3) and artemisinin treated mice (GF1, GF2, and GF3).

first: Yanmin


title: ALKBH5 impacts malignancy of colorectal cancer via m6A-mediated epigenetic modulation [meRIP-seq]

city: Guangzhou

first: Dingcheng

last: Shen

address: 26 Yuancun Erheng Road

institute: The Sixth Affiliated Hospital, Sun Yat-sen University

type: Methylation profiling by high throughput sequencing

summary: ALKBH5 is one of the important m6A demethylases and is reported to be closely associated with tumorigenesis.In this study, we report gene expression profiles of HCT116 cells treated with ALKBH5 shRNA. This study reveals a novel role of ALKBH5 in downstream gene regulation and provides a new explanation ofALKBH5’ s function in colorectal cancer.

overall_design: MeRNA-seq after knockdown of ALKBH5 in HCT116 cells  to explore the features and distribution dynamics of ALKBH5-mediated RNA m6A modification.

first: Yanxin

last: Luo

first: Huichuan

last: Yu


title: Nematode-induced morphological differentiation regulated by the histone H3K4 methyltransferase AoSET1 in nematode-trapping fungus Arthrobotrys oligospora

city: Kunming

first: Juan

state: Yunnan

address: North Cuihu Road #@

institute: Yunnan University

summary: AobZip129 is a bZip transcription factor gene，it affects the formation of adhesive networks and pathogenicity in Arthrobotrys ligospora.We analyzed global gene expression in the WT and AobZip129 strains grown in WA medium and further induced by NE for 24h when the traps began to form. Compared with the AobZip129 and WT strains grown in WA, we detected 1,542 and 1,271 genes were up- and down-regulated (1.5-fold change), respectively. Similarly, 256 and 327 genes (1.5-fold change) were up- and down-regulated in the AobZip129 NE-induced culture in comparison to their expression in WT NE-induced culture. KEGG enrichment analysis revealed that some of the AobZip129-dependent up-regulated genes were overrepresented in the categories of “proteasome”, “starch and sucrose metabolism”, “peroxisome”, “starch and sucrose metabolism”, and “phagosome”. By contrast, the putative AobZip129-dependent down-regulated genes were involved in “Ribosome biogenesis in eukaryotes”, “purine metabolism”, “citrate cycle (TCA cycle)”, “pyrimidine metabolism”, and “protein proceeding in endoplasmic reticulum”.

overall_design: RNA-seq analysis between wild-type samples and AobZip129 mutant samples, with three biological replicates per sample, and prediction of multiple cytological processes that may affect trap formation in Arthrobotrys oligospora.

first: Qiao

last: Miao

first: Ziyu

last: Yin

first: Ran

first: Ke-Qin

first: Juanl


title: ChIP-seq studies of Th17 cells in Malat1 KO and WT mice

city: San Diego

first: Wendy

address: 9500 Gilman Drive #0651

department: Cellular and Molecular Medicine

country: USA

institute: University of California San Diego

summary: Chromatin immunoprecipitation (ChIP) was performed in biological replicates. For H3K27me3 ChIP assays, cells were cross-linked with 1% (vol/vol) formaldehyde. For SUZ12 and EZH2 ChIP assays, cells were cross-linked with 2 mM disuccinimidyl glutarate (DSG) and 1% (vol/vol) formaldehyde.

overall_design: Two independent biological replicates were performed.

first: Shengyun

last: Ma

first: Bing

last: Zhou

first: Yohei

last: Abe

middle: J


title: A BNC1-IRF6-AP1 transcriptional network in keratinocytes and Squamous Cell Carcinoma

city: Boston

first: Matthew

last: Ramsey

state: MA

address: 77 Ave Louis Pasteur, HIM 668

department: Dermatology

institute: Brigham and Women's Hospital

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

overall_design: Refer to individual Series


title: A BNC1-IRF6-AP1 transcriptional network in keratinocytes and Squamous Cell Carcinoma [RNA-Seq]

summary: Analysis of transcriptional regulation by BNC1, IRF6, and FRA1 in keratinocyes and Squamous Cell Carcinoma cell lines

overall_design: We performed RNA-seq on keratinocytes or SCC cells 72 hours after doxycycline treatment to induce expression of shRNAs targeting BNC1, FRA1, IRF6, BNC1 and FRA1, or BNC1 and IRF6


title: neuroendocrine control of planarian regeneration

city: Shanghai

first: Yuxiaofei

address: 320 Yue-Yang Road

institute: University of Chinese academy of sciences

summary: Effective tissue regeneration requires cells to perceive the surrounding context and make internal state adaptations that enable context-dependent fate changes in time and space, but the mechanisms underlying this complex process remain unclear. Here, transcriptom sequencing was used for study neuroendocrine control of planarian regneration

overall_design: Examine gene expression profiles of 10dpf adult planarian after EGFP RNAi(control) and nsf RNAi.

first: Yue

last: Chen

first: An

last: Zeng


title: Proliferation is a driver of quorum sensing in an in vitro model of tissue resident macrophages

city: Melbourne

first: Jarny

last: Choi

state: Victoria

address: The University of Melbourne

department: Anatomy and Neuroscience

country: Australia

institute: Centre for Stem Cell Systems

summary: Macrophages coordinate the initial host inflammatory response to tissue infection as well as mediating the reparative phase by producing growth factors that promote tissue repair. One model of this functional dichotomy is that peripherally recruited monocyte-derived macrophages drive acute inflammatory responses to infection, whereas tissue resident macrophages are responsible for tissue repair. Alternatively, inflammation and repair may be inter-dependent molecular programs, such that both recruited and resident cells have equivalent capacity to contribute. In this study, induced pluripotent stem cells or peripheral blood monocytes were used to generate models of tissue-resident and recruited macrophages, respectively. Using single-cell sequencing, we assessed whether lipopolysaccharide activation drives heterogeneous responses in these two macrophage models. We observe heterogeneity at population and gene expression level in the activation programs adopted by either macrophage population. We further identify proliferation to be a driver of quorum sensing in LPS responsiveness. Our findings highlight the usefulness of this model system to explore the differences between recruited and resident macrophages in response to tissue infection.

overall_design: Monocytes(from 3 donors) /progenitor cells (from 3 iPSC lines) were cultured in tissue-culture treated 6 well plates (blood cells from all 3 donors were cultured together. Progenitors from all 3 iPSC lines were cultured together). Cells were cultured in RPMI-1640 medium containing L-Glutamine with 10% Fetal Bovine Serum and 100ng/ml recombinant Human M-CSF for 4 days. On day 4, media was changed and cells left to settle for either 2 hours before stimulation series. Stimulation consisted of either a 2 hour acute stimulation with 10ng/ml LPS, 18 hour stimulation with 10ng/ml LPS, or a 18 hour stimulation, 2 hour rest, and restimulation (tolerance) with 10ng/ml LPS.  Cells were collected and processed for 5' single cell sequencing.

first: Nadia

last: Rajab

first: Linden

last: Gearing

first: Yair

middle: D

last: Prawer

first: Paul

middle: W

last: Angel

first: Sean

middle: M

last: Grimmond

first: Andrew

middle: L

last: Laslett

first: Christine

middle: A

last: Wells


title: Proteogenomic profiling of thymic and pulmonary neuroendocrine neoplasia.

city: Göttingen

first: Hanibal

last: Bohnenberger

address: Robert-Koch-Straße 40

department: Institute of Pathology

country: Germany

institute: University medical center Göttingen

summary: This study represents a proteogenomic resource for thymic and pulmonary neuroendocrine neoplasia (NEN) and provides a proteomgenomic diagnostic signature for classification of low and high grade NEN.

overall_design: In this study, we characterized a cohort of 54 and 22 low and high grade NEN, respectively of both thymic and pulmonary origin by NanoString RNA analysis, NGS DNA Panel Sequencing analysis and MS-based proteomic analysis.

first: Stefan

last: Küffer

first: Philipp

last: Ströbel


title: Dynamics of gene expression profiling by microarrays and identification of high-risk patients for severe COVID-19 [Array]

city: Barcelona

first: Júlia

last: Perera-Bel

address: Doctor Aiguader, 88

department: MARData-BU

country: Spain

institute: Hospital del Mar Research Institute

summary: The clinical manifestations of SARS-Co-2 infection vary widely, from asymptomatic infection to the development of acute respiratory distress syndrome (ARDS) and death. The host response elicited by SARS-CoV-2 plays a key role in determining the clinical outcome. We hypothesized that determining the dynamic whole blood transcriptomic profile of adult patients hospitalized for COVID-19 and characterizing the subgroup that develops severe disease and ARDS would broaden our understanding of the heterogeneity in clinical outcomes. We recruited 60 hospitalized patients with microbiology-confirmed COVID-19, among whom 19 developed ARDS. Peripheral blood was collected using PAXGene RNA tubes within 24 hours of admission and at day 7. There were 2150 differently expressed genes in patients with ARDS at baseline, and 1963 at day 7. We found a dysregulated inflammatory response in COVID-19 ARDS patients, with an increased expression of genes related to pro-inflammatory molecules and neutrophil and macrophage activation at admission, in addition to the loss of immune regulation. This led in turn to a higher expression of genes related to reactive oxygen species, protein polyubiquitination, and metalloproteinases in latter stages. Some of the most significant differences in gene expression found between patients with and without ARDS corresponded to long non-coding RNA involved in epigenetic control.

overall_design: 137 samples were analyzed (Control=51, Covid19=52, Covid19_SDRA=34)

first: Alexander

last: Rombauts

first: Lara

last: Nonell


title: Investigation of spontaneous pelvic organ prolapse in the rhesus macaque model

first: Jian

last: Liu

address: Zhiyuan Road,Chenggong

institute: Institute of Primate Translational Medicine, KUST

summary: An appropriate model is urgently required for pathological mechanism investigation of pelvic organ prolapse and the development of new therapeutic strategies. It is generally thought that damage to the vagina and its supportive tissues due to vaginal birth, menopause, and so on can result in loss of vaginal support and prolapse of the pelvic organs. Currently, ovariectomy and simulated vaginal delivery in animal models are applied to mimic the status of menopause and induce vaginal injury, however, the complex biochemical and biomechanical changes induced by vaginal delivery and prolapse is difficultly achieved by these methods. The rhesus macaque shares many features similarly to women, such as pelvic floor anatomy and parturition, and makes an excellent experimental model for the study of prolapse. Besides, the rhesus macaque develops prolapse spontaneously. Thus, there has great value to investigate the changes of spontaneous prolapse and make them animal models for the study of prolapse. In this study, we will assess the histomorphology, biochemical property and biomechanical property of prolapsed vagina. And then, cell atlas of normal and prolapsed vagina will be constructed by single-cell transcriptome sequencing. We will reveal the cell type specific molecular mechanisms. This study will improve the understanding of spontaneous prolapse in the rhesus macaque and provide fundamental data for the application of spontaneous prolapse as animal models to study POP.

first: Zhu

last: Lan

first: Li

last: Yaqian

first: Tan

last: Tao

first: Liu

last: Jian


title: Deconvolution of cell types from bulk gene expression profiling in a nonalcoholic steatohepatitis model of mice liver reveals global alteration of macrophages and hepatic stellate cells

city: Indianapolis

first: Sarath

middle: Chandra

last: Janga

state: Indiana

address: 535 West Michigan Street

department: Informatics and Computing

institute: Indiana University-Purdue University Indianapolis

summary:
Nonalcoholic fatty liver disease (NAFLD) is a common cause of chronic liver disease due to the accumulation of excess fat in the liver cells. The two histological categories of NAFLD are NASH (nonalcoholic steatohepatitis) and NAFL (nonalcoholic fatty liver). While NASH is typically considered a disorder of the hepatocytes, the role of other liver cell types in its pathophysiology and progression is becoming more established. Bioinformatics analytic approaches can now incorporate single cell RNA transcriptomics (sc-RNA-Seq) as a reference to deconvolute cell type profiles and proportions in bulk /RNA-Seq data. This technique can be inferred to investigate the changes in cellular compositions in tissues across multiple disease states.
This study performed deconvolution analysis using CIBERSORTx, by integrating the bulk/RNA-Seq data with single-cell transcriptomic data to identify changes in cell composition associated with NASH. This study aimed to observe the changes in cell types among healthy and NASH mouse liver. The result of the analyses revealed a general increase in the fraction of HSC (Hepatic Stellate cells), liver macrophages, and cholangiocytes in NASH samples. Hepatocytes had a higher proportion in healthy samples compared with NASH, however, no significant observable difference was established in the endothelial cell proportions. We gained insight on the cell-type proportions profiles of NASH mice models, which highly mimics the human NASH.

overall_design: Comparative gene expression analysis between wild-type whole liver homogenate, and NASH whole liver homogenate

first: Omoru

last: Okiemute

first: Mansi

last: Srivastava

first: Anthony

last: Butterfield

last: Krohannon

first: Jim

last: Parfield

first: Rebecca

last: Miles


title: Effects of rFGF4 on gene expression during wound healing.

city: Wenzhou

last: Sun

address: chashan

institute: wenzhou medical university

summary: To investigate the roles of FGF4 in the regulation of skin wound healing, we established wound healing models on mice and the wound were topically treated by rFGF4 or PBS control for 6 days, and wound edges were collected for RNA-seq analysis.

overall_design:
We then performed gene expression profiling analysis using data obtained from RNA-seq.
Comparative gene expression profiling analysis of RNA-seq data for wounds of mice treated with rFGF4 or PBS controls.

first: Jie

last: Zhou


title: Optimised, frameshift-driven design for single-cell CRISPR perturbation screens

city: Cambridge

first: Alisdair

last: Russell

address: Robinson Way

country: United Kingdom

institute: Cancer Research UK Cambridge Institute

summary: CRISPR-Cas9 has been widely used to functionally interrogate multiple aspects of cellular physiology and pathophysiology from single gene studies to genome-wide screens. Proper design of highly efficient guide RNAs directing the CRISPR genome editing process is critical for success in these types of experiments. Here, we present a pipeline for designing highly efficient loss-of-function guide RNA (gRNA) libraries with improved rates of knock-out efficiency compared to previous guide RNA library designs. We provide pre-computed and triaged gRNAs from our pipeline for all human and mouse transcripts through a fully searchable online portal as a resource to the community.

overall_design: 13 Key regulators of the Oestrogen Receptor (ER) complex in MCF7 breast cancer cells were perturbed using CRISPR in duplicate experiments and analysed using scRNAseq

first: Abigail

middle: R

last: Edwards

last: Baker

first: Paula

last: D'Santos

first: Ashley

last: Sawle

first: Matthew

last: Eldridge

first: Florian

last: Markowetz

first: Rasmus

last: Siersbaek

first: Alasdair


title: A Single-Cell Atlas of HR+ Breast  Cancer Reveals the Shifting of the Tumor Microenvironment in Response to Neoadjuvant Chemotherapy

city: Hangzhou

first: Chao

last: Ni

state: Zhejiang

address: 88 Jiefang Road,Shangcheng District

department: Second Affiliated Hospital

institute: The Second Affiliated Hospital, Zhejiang University School of Medicine

summary: Neoadjuvant chemotherapy (NAC) is the standard presurgery treatment for HR+ breast cancer (HR+ BC), but its response rate is frequently unsatisfactory, and the mechanism of chemoresistance and how NAC reshapes the immune microenvironment are still poorly understood. To investigate this question, the transcriptomes and spatial distribution patterns of cancer cells and immune cells from paired HR+ BC samples pre- and post-NAC were characterized with single-cell RNA sequencing (N=8) and multiplexed immunofluorescence staining (N=21).

overall_design: Single-cell expression profiling of the  HR+ breast  cancer in pre- and post-treatment patients.  For data usage terms and conditions, please contact the principle investigator.

first: Fang

last: Jia

first: Jiaxin

last: Li


title: Claudins family as potential prognostic indicators in ovarian cancer by bioinformatics analysis

city: Shenyang

first: Qian

address: Liaoning

institute: Cancer Hospital of China Medical University/ Lianning Cancer Hospital&Institute

summary: In this paper, the expression and possible function of CLDNs in ovarian cancer were explored by bioinformatics analysis, and verified by mRNA high-throughput sequencing of ovarian cancer and cancer-free ovary.

overall_design: Comparative gene expression profiling analysis of mRNA-seq data for 4 pairs of ovarian cancer and cancer-free ovary and their KEGG Pathway Enrichment Analysis.

first: Xin

last: Zhang


title: Gene expression data of human bone marrow mesenchymal stem cells

city: Chengdu

first: Yu

last: Shen

address: NO.2025 Chengluo Avenue

institute: Chengdu University

summary:
Extracellular matrix (ECM) acts as an intrinsic and natural microenvironment of cells, has been shown a pivotal effect on numerous biological processes of MSCs. Especially, MSCs are inclined to differentiate into specific cells lineage commitment when grow on their adaptive and tissue-like microenvironment elasticity. The mechanism still remains elusive on mechanical transduction processes in the microenvironment.
In this experiment, human bone marrow mesenchymal stem cells were seeded on substates  with different ECM rigidity, and the changes of gene expression were detected by chips, so as to explore the effect of extracellular microenvironment on stem cell differentiation.

overall_design: Human bone marrow mesenchymal stem cells were seeded on substrates of 32Kp and 0.5Kpa, and the total RNA samples were collected on 3 and 7 days respectively for chip detection.Therefore, the test samples were divided into 4 groups according to different culture time and matrix hardness (3H-represents the experimental group cultured on 32kpa substrate for 3 days, 7H-cultured on 32kpa substrate for 7 days, 3S-cultured on 0.5kpa substrate for 3 days, and 7S-cultured on 32kpa substrate for 7 days)

first: Dian

last: Jing


title: RNA sequencing of Naive CD8 T cells isolated from Brca1 wildtype and Brca1 het splenocytes

city: Washington

first: Leilei

last: Qi

state: DC

address: 2300 I St NW

institute: The George Washington University

summary: To determine the impact of Brca1 gene heterozygosity on transcriptomics of CD8+ T cells, we carried out bulk RNA-sequencing (RNA-seq) using naive CD8+ T cells isolated from WT and Het mice.

overall_design: Naive CD8+ T cells were directly isolated from mouse splenocytes

first: Bogang

last: Wu

first: Li-Ju

last: Wang

first: Rong


title: Transcriptomic studies of mouse naïve and Th17 cell from wildtype or Malat1 deficient background

first: Wendy

last: Huang

summary: Long non-coding RNAs (lncRNAs) are critical regulators of mammalian gene programs. Metastasis Associated Lung Adenocarcinoma Transcript 1 (Malat1) is the most abundant lncRNA expressed in intestinal Th17 cells critical for maintaining tissue homeostasis and regulating local inflammation. Here, we report that Malat1 negatively regulates IL-17A and IL-17F productions in intestinal Th17 cells during colitis and contributes to local inflammation. Global RNA interactions with DNA by deep sequencing (GRID-seq) coupled with transcriptomic studies revealed Malat1 is recruited to the Il17a-Il17f super enhancer in Th17 cells, and regulates Il17a-Il17f transcription.

overall_design: Two independent biological replicates of Th17 cells were performed. The wild type samples are GSM4770961 and GSM4770962.

first: Shengyun

last: Ma

first: Bing

last: Zhou


title: Investigating the transcriptomics of MCL-1 knockout Nasopharyngeal carcinoma cell lines using next generation microarray technology

city: Gelugor

first: Nethia

last: Mohana Kumaran

state: Penang

address: Persiaran Sains

department: School of Biological Sciences

country: Malaysia

institute: Universiti Sains Malaysia

summary: To investigate the changes of gene expression in the NPC MCL-1 knockout cell lines when compared to wildtype, in order to find out the precise target for the co-inhibition with MCL-1 in NPC treatment.

overall_design: The MCL-1 knockout cell lines, HK-1 MCL-1 knockout cell line and C666-1 MCL-1 knockout cell line, were established from their respective wildtype cell lines, HK-1 cell line and C666-1 cell line, by using the CRISPR-Cas9 technology. Total RNA were extracted from these cell lines: HK-1 MCL-1 knockout cell line, C666-1 MCL-1 knockout cell line,  HK-1 cell line and C666-1 cell line for the microarray.

last: Mohana-Kumaran

first: Jia Lin

last: Teh


title: Human In-vitro Inflammatory Liver Model Recapitulates Immune-Associated Drug Efects with High Predictivity

city: Singapore

first: Farah

last: Tasnim

state: Singaore

address: 31 Biopolis Way, #05-01, Nanos Building

country: Singapore

institute: Institute of Bioengineering and Bioimaging

summary: Immune-mediated drug-induced liver injury is often undermined or not well-studied due to lack of appropriate in vitro models comprised of immune cells and suitable endpoints. Current advances in co-cultures deploying primary human Kupffer cells (PHKCs) or alternative cells such as THP-1 macrophages are limited by their inability of recapitulating drug-induced cytokine responses which are important immune-mediated drug effects. Since human iPSC-derived Kupffer cells (iKCs, PMID: 30497023) functionally resemble PHKCs, we investigated the use of iKCs to mimic drug-specific cytokine responses in vitro. iKCs were co-cultured with hiPSC-derived hepatocytes, which were then subjected to treatment of drugs with known immune-mediated effects. Out of the cytokine measurements including interleukin-6 and tumor necrosis factor α obtained from 19 compounds, 17 showed good in vitro-in vivo correlation (89.5%). Furthermore, co-cultures using THP-1 macrophages instead of iKCs could not recapitulate IL-6 responses upon treatment with paradigm compounds. RNA-Seq was used to explore the interaction between cytokines and drug-induced hepatotoxicity in greater detail for one pair of drugs - trovafloxacin and its non-hepatotoxic analogue, levofloxacin. Our study demonstrates for the first time, the use of iKCs in a physiologically relevant model that captures cytokine responses upon treatment with a wide range of immune-mediated drugs and hence can be very useful in warranting further investigation of new drugs that elicit drug-induced cytokine imbalance.

overall_design: Human pluripotent stem cells were differentiated into hepatocytes (iHeps) and Kupffer cells (iKCs). On the first day, iKCs were seeded, followed by iHeps 24h later. On the third day, the co-culture was subjected to drug treatment for 48h. RNAseq was performed on 12 sample types with 3 biological replicates each (except mTVX+ which only has 2 replicates). The sample types are named as <mono-or co-culture (m or c)> <vehicle or trovafloxacin or levofloxacin (Veh or TVX or LVX)> <absent or including 100ng/ml lipopolysaccharide (- or +)>. E.g. mTVX+ indicates an iHep monoculture treated with trovafloxacin and LPS.

first: Xiaozhong

first: Yun-Ting

last: Soong

first: Claire

last: Ng

first: Jiahao

first: Hanry

last: Yu


title: GRID-seq studies of mouse naïve and culture Th17 cells from wildtype mice

overall_design: Two independent biological replicates of naïve and Th17 cells were performed.

first: Xiang-Dong

last: Fu


title: Characterization of a transcriptional factor FevR, involved in improving folding in the periplasm of Escherichia coli.

city: Ipswich

first: Augusto

middle: Emanuel

last: Garcia

address: 240 County Rd

department: Research

institute: New England Biolabs

summary: DegP (HtrA: High Temperature Requirement A) is a periplasmic protease with minor chaperone activity that plays a key role in the quality control of protein folding in the envelope of Escherichia coli. Periplasmic and outer membrane proteins that fail to fold in the periplasm can be proteolysed, while others are chaperoned to their native folded state by DegP. In a ΔdegP strain, E. coli is unable to survive the protein folding stress induced at 42º C. Utilizing this phenotype, we developed a plasmid-based selection of suppression of heat-induced lethality in a ΔdegP strain. Plasmid libraries of various prokaryotic genomes were screened for proteins that overcame heat-induced lethality.   We identified FevR, a putative transcription factor from Citrobacter amalonaticus having close homologs in environmental E. coli capable of overcoming envelope stress. Through genetic characterization, FevR is shown to induce expression of a periplasmic chaperone-proline isomerase fkpA. Over-expression of FkpA alone is sufficient to suppress heat-induced lethality of a ΔdegP strain. This study demonstrates the use of genetic selections to uncover the hidden potential of E. coli to improve its protein folding capacity.

overall_design: Differential expression of genes by one of the two homolog transcriptional factor, FevRCit or FevREco105, compared to empty vector, in different conditions: background strain E. coli K-12 DHB4 wild type or ΔdegP, grown at 30°C, with or without a heat shock at 42°C during log phase, or grown at 42°C

first: Eliz

last: Dikener

first: Iris

last: Walker

last: Ke

first: James

last: Eaglesham

first: Javier

last: Santos

first: Mehmet

last: Berkmen


title: A BNC1-IRF6-AP1 transcriptional network in keratinocytes and Squamous Cell Carcinoma [ChIP-Seq]

first: Matthew

last: Ramsey

summary: Analysis of BNC1, IRF6, FRA1, and JUNB genome binding/occupancy in epidermal keratinocyes and cutaneous Squamous Cell Carcinoma cell lines

overall_design: ChIP-seq of BNC1, IRF6, FRA1, and JUNB in SCC-13 and G5-Ep/TERT cell lines


title: Expression of Frankia inefficax EuI1c under selenite stress

city: Durham

first: Louis

middle: S

last: Tisa

state: NH

address: 46 College Rd

department: MCBS

institute: University of New Hampshire

summary: Purpose: To compare RNASeq data of  Frankia inefficax EuI1c under selenite stress.   Frankia cultures were grown for 4 days under 0.1 mM selenite stress  or without selenite (Control).  RNA-seq analysis provided insight into how the the bacteria responds to selenite stress and exhibits selenite tolerance.

overall_design:
mRNA profiles of bacteria from Frankia inefficax EuI1c cultures grown under selenite stress and no added selienite conditions were generated by deep sequencing and mapped to F. inefficax EuI1c genome by CLC Gemonic Workstation. The transcriptome profiles of the selenite stressed cells were compared to the control (no selenite) condition.

USDA NIFA Hatch project NH00686 "Functional Genomic Approaches toward Understanding the Frankia-Actinorhizal Plant Association and their Responses to Harsh Environments"

first: Medhat

last: Rehan


title: Transcriptome analysis of immortalized OPLL ligament cells

first: Chen

last: Xu

address: No, 800, Rd. Xiangyin

institute: The Second Military Medical University

summary: Ossification of the posterior longitudinal ligament (OPLL) is formed by heterogeneous ossification of posterior longitudinal ligament. The patho-mechanism of OPLL is still largely unknown. For the rarity of the primary cell samples, here we immortalized the OPLL patient's posterior longitudinal ligament primary cells and normal patient's posterior longitudinal ligament primary cells. The transcriptome of the cells were compared and we found that the osteogenic ability of the OPLL ligament cells were inherited in the immortalized cells.

overall_design: mRNA profiles were done using primary cells of human ossification of the posterior longitudinal ligament (OPLL) tissue and normal posterior longitudinal ligament (PLL) tissue. The cells were immortalized using SV40 and hTERT insertion. The mRNA profiles of immortalized cells using either SV40 and hTERT insertion were also analyzed using sequencing technologies.

first: Yang

last: Liu

first: Wen

last: Yuan


title: Evaluation of potential impact of anti-LIF therapy on the tumor microenvironment in the immune-component mice by scRNA-seq analysis.

city: Beijing

first: Rong

last: Huang

address: 5 Dong Dan San Tiao

institute: Chinese Academy of Medical Sciences and Peking Union Medical College

summary: We report the application of single-molecule-based sequencing technology for anti-LIF therapy on the tumor microenvironment in the immune-component mice.C57 mice were treated with isotype control IgG, anti-LIF, anti-PD-1, or the combination of anti-LIF with anti-PD-1 (combo) individually after subcutaneously injected with PAN02 cells for 5 days. we explored the therapeutic potentials in blocking the LIF-LIFR signaling pathway and confirmed that it can effectively enhance the tumor responsiveness to PD-1 immunotherapy by shifting the T-cells population composition in addition to the inhibition of tumor growth.

overall_design: Anti-PD-1, anti-LIF and combo groups were analyzed in comparison to the control, 3 samples per group.

first: Yanhong

last: Wang


title: Transcriptome analysis of EGR1 knock-down eutopic endometrial mesenchymal stem cells by RNA-sequencing in patients with adenomyosis.

city: Nanjing

first: Tao

last: Gui

state: Jiangsu

address: NO.100 Shizi Street, Hongshan Road

institute: Affiliated Hospital of Integrated Traditional Chinese and Western Medicine, Nanjing University of Chinese Medicine

summary: To identify differentially expressed genes (DEGs) and molecular pathways in EGR1 knock-down endometrial mesenchymal stem cells (eMSCs) from adenomyosis patients and provide a new insight into disease mechanisms at transcriptome level.

overall_design: Gene expression profiling of EGR1 knock-down endometrial mesenchymal stem cells (EGR1-KD) and scrambled-sequence control (Scr) were analyzed by using RNA-sequencing.

first: Guiping

last: Wan

first: Meiling

last: Cao


title: Expression data of Frankia inefficax strain EuI1c under biphenyl stress

summary: Purpose: To compare RNASeq data of Frankia inefficax EuI1c in normal growth conditions to biphenyl-stressed Frankia inefficax EuI1c. Frankia cultures were grown for 3 days under one of three conditions before harvesting: 1.) 1 mM biphenyl stress with glucose as an alternative carbon and energy source, 2.) 1mM biphenyl with no alternative carbon or energy source, or 3.) media with glucose as a carbon and energy source (Control). RNA-seq analysis provided insight into how the the bacteria responds to biphenyl stress and exhibits biphenyl tolerance.

overall_design: mRNA profiles of bacteria from Frankia inefficax EuI1c cultures grown under biphenyl stress and no added biphenyl conditions were generated by deep sequencing and mapped to F. inefficax EuI1c genome by CLC Gemonic Workstation. The transcriptome profiles of the biphenyl stressed cells were compared to the control (no biphenyl) condition.

first: Erik

last: Swanson


title: Sex-specific transcriptomic response of Anopheles gambiae to O'nyong Nyong virus infection

city: University Park

first: Cory

last: Henderson

state: PA

address: Room W127, Millennium Science Complex

department: molecular cellular and integrative biosciences

institute: Pennsylvania State University

summary: O’nyong’nyong virus (ONNV) is an arbovirus of the genus Alphavirus which is  transmitted to humans primarily by mosquitoes of the genus Anopheles. Here, An. stephensi  males and females have been infected with ONNV to investigate their differential  transcriptional response to infection. Males and female An. gambiae were injected with either  a suspension of ONNV or sterile medium as a control and underwent pooled RNA extraction at  2- and 7-days post infection (DPI). Differential expression will be performed on pooled RNA  samples to determine which genes are differentially regulated between males and females in  response to infection, and which genes are activated or repressed in each sex as infection  progresses. Further, genes identified as being regulated by infection or that differ in expression  between males and females will be used to perform gene ontology (GO) analysis to determine  what pre-identified gene sets are regulated under the scenarios being examined.

overall_design: 24 total samples have total RNA sequenced from 2 different time points, (2 and 7 days post infection). 3 replicates are sequenced from each time point, infection status (O’nyong’nyong infected or control), and sex (male or female). Differential expression on read counts mapping to known genes in the AgamP4.12 genome is conducted to determine genes regulated in response to infection at different time points and how this response differs by sex.

first: Karen

last: Kemirembe

first: Alexandra

last: Beck

first: Christina

last: Bergey

first: Jason

last: Rasgon


title: Integrative analysis of altered genes expressed in human Alzheimer’s disease brain

city: Fukuoka

first: Yusaku

last: Nakabeppu

state: Fukuoka

address: 3-1-1 Maidashi Higashi-ku

department: Department of Immunobiology and Neuroscience

country: Japan

institute: Medical Institute of Bioregulation, Kyushu University

summary: We had previously performed comparative gene expression profiling of human postmortem Alzheimer’s disease (AD) brains donated for the Hisayama study. The hippocampi from AD brains showed the most significant alteration in gene expression profile [PMID: 23595620]. In the present study, in order to identify molecular pathological alterations in AD hippocampus, we applied human transcriptome array and RNA-sequencing to the same samples and reanalyzed the gene array data, and performed integrative analysis of all 3 methods. We also applied gene array to hippocampus of 6-month-old AppNL-G-F/NL-G-F knock-in AD model mice as a preclinical AD stage model, and at last we performed the interspecies comparison. Functional annotation clustering with the results of all 3 or at least 2 methods in human, showed synapse, cell junction, calmodulin binding, ion transport and glycoproteins as the top affected terms. In hippocampi of AppNL-G-F/NL-G-F mice which exhibit significant amyloidosis but not neuronal degeneration, we found that expression of genes categorized in signaling specially signaling peptides with disulfide bond or glycoproteins, together with the innate immune response and lysosome of cells are upregulated. Comparing the extended human data with mouse data, we found and confirmed by qRT-PCR, 2 common up-regulated genes, Clec7a and C4b, which induce innate immune response and synapse elimination. We also found and confirmed by qRT-PCR, 2 common down-regulated genes, one is Slc17a6, which can affect synaptic transmission of glutamate, and second one is Rxfp1, which is involved in gene transcription. Amyloid-beta accumulation is the major hallmark of AD at 6 month-old AppNL-G-F/NL-G-F  mice, therefore our data suggests that amyloid-beta accumulation induces innate immune response, synapse elimination, besides it can decreases glutamate transmission, and affect gene transcription in early stage of hippocampal AD brain, prior to neuronal degeneration.

overall_design: We analyzed postmortem human hippocampus brains from 8 AD and 10 non-AD subjects using the Affymetrix Human Transcriptome Array (HTA) 2.0. Array data was processed by Affymetrix Transcriptome Analysis Console (TAC) software 4.0.

first: Nona

last: Abolhassani


title: Blood gene expression changes in response to exposure to dust containing silica

city: Morgantown

first: Pius

last: Joseph

state: WV

address: 1095 Willowdale Road

department: Health Effects Laboratory Division/Toxicology and Molecular Biology Branch

institute: National Institute for Occupational Safety and Health (NIOSh)

summary: Occupational exposure to dust containing crystalline silica may result in serious adverse health effects including silicosis and cancer. Previous studies which employed animal models for inhalation exposure to crystalline silica revealed changes in blood gene expression profiles in association with the silica-induced lung toxicity. Currently, global gene expression profiles were determined in the whole blood samples obtained from control (not exposed to dust) and dust containing silica exposed individuals with or without clinically identified silicosis. Differences in the blood gene expression profiles were detected in the blood samples obtained from the control and the silica-containing dust exposed individuals. Between the two groups of the silica-containing dust exposed individuals, the number of significantly differentially expressed genes was more in the blood samples obtained from those with silicosis.

overall_design: Whole blood samples collected from individuals who were exposed to dust containing silica were used in the study. Blood samples collected from an unexposed, matching cohort were used as the controls (CTl2 to CTl60) (n = 23). The lung function and silicosis status of the study participants were determined by spirometry and chest x-ray, respectively. Based on the results of chest x-ray, the silica exposed subjects were classified into two groups – dust exposed with silicosis (TESC_02 to TESC_40) (n = 30) and dust exposed without silicosis (TESC_42 to TESC_89) (n = 33). Gene expression profiles were determined by next generation sequence analysis in the blood samples collected from the subjects.

first: Elijah

last: Lowe


title: RNA-seq analysis of differentially expressed genes between primary cells and recurrent cells in a B-prolymphocytic leukemia (B-PLL).

city: Kobe

first: Shinichiro

last: Kawamoto

state: Hyogo

address: Chuoh-ku Kusunokicho 7-5-2

department: Department of Transfusion Medicine and Cell Therapy

institute: Kobe University Hospital

summary: We confirmed that transcription of FAS was  upregulated about 30 fold highly in the recurrent cells that lost CD20 expression and showed tumor lysis syndrome compared to the cells at diagnosis in a B-prolymphocytic leukemia patient.

overall_design: Examination of mRNA expression in leukemia cells between the primary cells and the recurrent cells.


title: The effect of microglial STAT3 signaling on early postnatal brain development

city: College Station

first: Jianrong

last: Li

state: TX

address: VBEC Receiving, VICI Bldg Rm127, 4458 TAMUS

department: Department of Veterinary Integrative Biosciences

institute: Texas A&M University

summary: Microglia play critical roles in the maturation and refinement of neural networks of the developing brain.  The goal of this study was to investigate the role of microglial STAT3 in postnatal brain development. This experiment was aimed to determine gene profiles (RNA-seq) of wild-type and Stat3 mutant microglia.  Cx3cr1CreERT2/+:Stat3loxP mice were generated to achieve tamoxifen-inducible deletion of exon 22 of the Stat3 gene in microglia. mRNA was prepared from acutely isolated microglia from postnatal day 8 (P8) pups that received tamoxifen from postnatal day 1 to 3.  Overall, we found that Stat3 mutant produced signficantly higher levels of inlfammatory cytokines and had compromised DNA damage checkpoints.

overall_design: Cx3cr1CreERT2/+:Stat3fl/fl mice received daily tamoxifen from P1-P3. CD11b+ cells were isolated by percoll gradient centrifugation and FACS sorting from P8 forebrains of Stat3 conditional knockout (Cx3cr1-CreERT/+: Stat3 fl/fl) and littermate controls (Stat3fl/fl). Each sample represents a pool of 2-4 pups (both sexes) with the same genotype.  TruSeqRNA Sample Prep Kit (Illumina) was used for poly A mRNA selection and library preparation. Fourty base-pair reads were generated for each sample on an Illumina HiSeq 2500 instrument at the Whitehead Institute Genome Technology core facility. A total of 286 million reads were mapped against GRCm38/mm10 assembly using TopHat version 2.0.13 with default settings, out of which a total of 274 million reads (approximately 95%) were aligned to the reference genome.

first: H

last: Lu

first: S

last: Kim

first: J


title: Transcriptomic and miRNA analysis of control versus treated HCT-116 cells with vitamins A and D

city: Chicago

first: Gail

middle: B

last: Mahady

state: IL

address: 833 S Wood St

department: Pharmacy Practice

institute: University of Illinois

type: Non-coding RNA profiling by high throughput sequencing

summary:
mRNA-seq:
HCT-116 colon cancer cells were grown and maintained and treated with a combination of Vitamin D2 and D3 and all trans-retinoic acid (ATRA) at 1 µM for four hours. The cells were harvested, RNA was isolated and analyzed for integrity using Agilent 4200 TapeStation and RNA Screen Tape. RNA-seq libraries were prepared using a Universal Plus mRNASeq kit, and library fragment size distribution was confirmed by electrophoresis using a 2200 TapeStation system with D1000 ScreenTape. Sequencing was carried out on NovaSeq 6000, SP flowcell, 2x50 nt reads. General quality-control metrics were obtained using FastQC and raw reads were aligned to human reference genome hg38 using STAR and BWA MEM. ENSEMBL genes were quantified using FeatureCounts, and differential expression statistics were computed using edgeR. Specific gene expression was measured and verified using qPCR.
miRNA-seq:
HCT-116 cells were cultured in McCoy’s 5a Medium (Gibco, Life Technology, Grand Island, NY, USA) supplemented with 10% FBS and 1% Penicillin/Streptomycin and incubated at 37°C in a humidified atmosphere of 5% CO2 and 95% air. At 80% confluence, cells were harvested by adding 0.25% trypsin/EDTA and counted by means of Trypan blue and hemocytometer. The cells were then re-suspended at appropriate concentration and plated in 96 or 6 well plates for cellular assays. Cell were treated with ATRA+D2+D3 at the IC50 concentration (1mM), then harvested and RNA isolated using Trizol

overall_design: Control samples in triplicate versus treated samples in triplicate

first: Pinal

last: Kanabar


title: Genome-wide maps of chromatin state in mouse Th17 cell [ChIP-seq]

city: San Diego

first: Wendy

last: Huang

state: CA

address: 9500 Gilman Drive #0651

institute: University of California San Diego

summary: We applied ChIP-seq technology for mapping chromatin modification in Th17 cells. We find that acetylations on histone H3 lysine 27, an active enhancer mark, significantly overlap with open chromatin as defined by ATAC-seq. These results were used to define putative super enhancer loci in Th17 cells. Our study shows that super-enhancer mediated gene transcription is important  for Th17 effector functions.

overall_design: Examination of histone H3 lysine 27 acetylation in two independent biological replicates of cultured Th17 cells. Input genomic DNA from each biological replicate were used for MACS peak call analysis.

first: Shengyun

last: Ma

first: Yuxin

first: Bing

last: Zhou

first: Xiang-dong

last: Fu


title: Transcriptomic studies of mouse naïve and Th17 cell from wildtype or DDX5 deficient background [RNA-seq]

summary: Gene expression in naïve and Th17 cell were characterized. We report a notable dynamic change in the expression of genes encoding RNA editing enzymes. Birefly, the transcript encoding ADAR2 (Adarb1) was significantly upregulated in Th17 cells, and the transcript encoding ADAR1 (Adar) was downregulated during the polarization from naive to Th17 cells. The altered expression of RNA editing genes results in the dynamic A-to-I editing in naive and Th17 cells. In addtion, loss of DDX5 dysregulated ADAR2 expression, resulting in altered A-to-I editing on a subset of Th17 transcripts.

overall_design: Two independent biological replicates in each condition were performed. In wildtype cells, Th17 results were compared to control naïve conditions. Results from DDX5-deficient Th17 cells were compared to wildtype Th17 cells.

first: Benjamin

last: Cho


title: Chromatin accessibility timecourse of Naïve to Th17 polarization in wildtype mouse CD4 T cells [ATAC-seq]

summary: We applied ATACseq technology for mapping chromatin accessibility during CD4+ T cells activation. Briefly, we generated genome-wide chromatin-state maps of naive, Th17 at 48 hours and 72 hours. Using a series of ATAC-seq timecourse experiment, we revealed that chromatin accessibility is dynamic  between naive and Th17, especially at the Th17 singature gene loci. These open chromatin sites may function as  important cis-regulate elements for cell-specific genes expression.

overall_design: Two independent biological replicates in each condition were performed.


title: Systematic mapping of 2’-O-methylation sites in rRNAs implicates new strategy for ribose methylation in Arabidopsis

city: Shanghai

first: Chenjiang

last: You

state: Shanghai

address: 2005 Songhu Rd

institute: Fudan University

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

overall_design: Refer to individual Series


title: PHF6 Interacts with LMO2 During Normal Haematopoiesis and in Leukaemia and Regulates the Level of Gene Expression and Genome Integrity

city: Birmingham

first: Maarten

last: Hoogenkamp

state: West Midlands

address: Vincent Drive

department: Institute of Cancer and Genomic Sciences

institute: University of Birmingham

summary: The transcriptional mediator LIM domain only 2 (LMO2) forms a large multi-protein complex together with TAL1/LYL1, HEB/E2A, LDB1 and GATA. This complex regulates transcription from the onset of haematopoietic development and during differentiation. Chromosomal re-arrangements involving LMO and other members of the complex are causative for T-cell lymphoblastic leukaemia (T-ALL). We have identified Plant Homeodomain (PHD)-like Finger 6 (PHF6) as a new LMO2 interacting factor. Somatic mutations in PHF6 have been found to occur in several types of leukaemia. We show that PHF6 interacts with LMO2 during the initial stages of the haematopoietic development, myeloid differentiation and in T-ALL. The LMO2/PHF6 complex binds the DNA and regulates linage-specific gene expression. Additionally, a loss or reduction of LMO2 and PHF6 leads to chromosomal instability. PHF6 and LMO2 are required for maintaining levels of γH2AX and 53BP1, where PHF6 is important for γH2AX accumulation and LMO2 has a role in recruiting 53BP1 to γH2AX foci.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: ChIPseq (single end), using (1) aLMO2, aTAL1, aLDB1, aGATA2, aPHF6 antibodies, in T-ALL cell lines (ARR, DU.528, HSB2, CCRF-CEM), (2) aPHF6 antibody in WT Flk-1+ haemangioblasts, (3) aLMO2, aTAL1, aLDB1, aPHF6 antibodies in the myeloid cell line PUER and its derivative shPHF6 PUER. RNAseq (paired end) of the myeloid cell line PUER and its derivative shPHF6 PUER at day 0, day 1, and day 3 of differentiation.

first: Vesna

last: Stanulovic


title: Systematic mapping of 2’-O-methylation sites in rRNAs implicates new strategy for ribose methylation in Arabidopsis [20ME]

summary: 2’-O-methylation (2’-O-me) is one of the most abundant covalent modifications in ribosomal RNAs (rRNAs) vital for the functionality of the ribosome, in particular, translational regulation. Aberrant 2’-O-me causes developmental defects in animals and is associated with human cancer. However, a lack of systematic delineation of 2’-O-me sites significantly impedes our understanding of 2’-O-me in plants. Here we report the global profiling of 2’-O-me in Arabidopsis rRNAs, leading to the identification of 221 2’-O-me sites, including 137 novel ones compared to the Arabidopsis database that documents predicted rRNA 2-O-me sites. We show that 65% of yeast and 53% of human 2’-O-me sites are conserved in Arabidopsis. We discover not only 2’-O-me guided by box C/D snoRNAs, but also unprecedentedly a prevalent snoRNA-independent mechanism in Arabidopsis: 48% of the 2’-O-me sites are devoid of snoRNAs, including some conserved sites where snoRNAs are indispensable in human and yeast. Through HITS-CLIP (high-throughput sequencing of RNAs isolated by cross-linking immunoprecipitation), we show that Fibrillarin (FIB1) and FIB2 predominantly associate with box C/D snoRNAs in vivo and uncover previously unknown snoRNAs. By generating and analyzing mutants in the FIB genes, we demonstrate that they are responsible for the ribose methylation at many rRNA sites regardless of their snoRNA-dependent or independent nature. We report, for the first time, extensive 2’-O-me of chloroplast rRNAs and, intriguingly, the dependence of FIB1/2 at some sites. Our studies reveal the landscape of 2’-O-me in nuclear and chloroplast rRNAs and provide novel insights into the deposition of rRNA 2’-O-me.

overall_design: Total RNAs were extracted from 14-day-old Arabidopsis seedlings. Eight samples including 2 biological replicates of wild type, fib1-1, and fib1-2-/- fib2-4+/-, one replicate of fib2-3 and  fib1-3-/- fib2-4+/-

first: Yong

last: Zhang

first: Runlai

last: Hang

first: Shaofang

first: Xinbei

last: Mo

first: Xiaofeng

last: Cao

first: Xuemei

last: Chen


title: Role of tobacco smoke on lung response to crystalline silica exposure in rats

first: Pius

last: Joseph

summary: Previous studies have shown that smoking induces oxidative stress and inflammation, known factors that coincide with the development and progression of lung toxicity in response to crystalline silica exposure.  Nevertheless, the precise role of tobacco smoke exposure on the lung response to tobacco smoke exposure and the underlying mechanisms remain largely elusive. Therefore, the objective of the present study was to determine the effect of smoking, if any, on silica-induced pulmonary toxicity and the underlying molecular mechanisms. Pulmonary toxicity and lung gene expression profiles were determined in rats exposed to air, crystalline silica, tobacco smoke, or crystalline silica plus tobacco smoke. Silica exposure resulted in significant pulmonary toxicity which was further exacerbated by tobacco smoke exposure in the rats. Significant differences in the gene expression profiles were detected in the lungs of the rats exposed to tobacco smoke, silica or a combination of both compared with the air exposed control rats.

overall_design: Approximately 3 months old male Fischer rats (Charles River Laboratories, Wilmington, MA) were used in this study which was conducted in an Association for Assessment and Accreditation of Laboratory Animal Care International approved animal facility (NIOSH, Morgantown, WV) following a protocol approved by the Animal Care and Use Committee. The rats were housed in groups of 3 rats/cage and maintained on a 12-hour light-dark cycle in a temperature (68-72oF) and humidity (30-70%) controlled room with free access to filtered tap water and Teklad rodent diet (Envigo, Indianapolis, IN). The rats were divided into 4 groups (n=6) and exposed to air (control) tobacco smoke (80 mg/m3, twice weekly, 12 months), crystalline silica (15 mg/m3, 6 hours/day, 5 days) or crystalline silica (15 mg/m3, 6 hours/day, 5 days) followed by tobacco smoke (80 mg/m3, twice weekly, 12 months) by whole-body inhalation. Approximately 12 months following initiation of the exposures, the rats were euthanized, and lung gene expression profiles determined by RNA sequencing. Total RNA isolated using the miRNeasy Kit (Qiagen, Inc, Valencia, CA) and next generation sequencing libraries prepared using TruSeq Stranded mRNA Library Prep Kit (Illumina, Inc, San Diego, CA) were used in the sequencing.

first: Tina

last: Sager

first: Elijah

last: Lowe

first: Naveena

last: Yanamala


title: Distribution of CHD7 and H3K27ac in cardiac nerual crest cells

city: Augusta

first: Kai

last: Jiao

state: GA

address: 1462 Laney Walker Blvd.

department: Center for Biotechnology and Genomic Medicine

institute: Medical College of Georgia at Augusta University

summary: We isolated cardiac neural crest cells from E10.5 embryos and performed CHD7- and H3K27ac- ChIP-Seq. From this study, we determined the localization of CHD7 and H3K27ac along the chromosomes in cardiac nerual crest cells.

overall_design: CHD7- and H3K27ac- ChIP seq using E10.5 cardiac nerual crest cells

first: Shun

last: Yan

first: David

last: Crossman


title: CHD7 downstream genes in mouse cardiac neural crest cells

summary: CHD7 encodes an ATP-dependent chromatin remodeling factor. Mutation of this gene causes multiple developmental disorders including CHARGE syndrome, in which conotruncal anomalies are the most prevalent form of heart defects. How CHD7 regulates conotruncal development remains unclear. In this study, we establish that deletion of Chd7 in neural crest cells (NCCs) causes severe conotruncal defects and perinatal lethality. We thus provide the first mouse genetic evidence demonstrating that CHD7 cell-autonomously regulates cardiac NCC development, thereby clarifying a long-standing controversy in the literature. Using transcriptomic analyses, we show that CHD7 fine-tunes the expression of a gene network that is critical for cardiac NCC development. To gain further molecular insights into gene regulation by CHD7, we performed a protein-protein interaction screen by incubating recombinant CHD7 on a protein array. We find that CHD7 directly interacts with several developmental disorder-mutated proteins including WDR5, a core component of H3K4 methyltransferase complexes. This direct interaction suggested that CHD7 may recruit histone modifying enzymes to target loci, independently of its remodeling functions. We therefore generated a mouse model that harbors an ATPase-deficient allele and demonstrate that mutant CHD7 retains the ability to recruit H3K4 methyltransferase activity to its targets. Thus, our data uncover that CHD7 regulates cardiovascular development through ATP-dependent and -independent activities, shedding light on the etiology of CHD7-related congenital disorders. Importantly, our data also imply that patients carrying a premature stop codon versus missense mutations will likely display different molecular alterations; these patients might therefore require personalized therapeutic interventions.

overall_design: mRNA-Seq using Chd7 wild type or Chd7-deletion cardiac neural crest cells.

first: Dongquang


title: Pathway-Associated Gene Expression in Mouse GI Tissues Under Western Diet and Fetal Antibiotic Exposure.

city: Rockville

first: Jeffrey

last: Robinson

state: United States

address: 9630 Gudelsky Dr

department: Translational Life Sciences Technology

institute: UMBC

summary: Pregnant mice were treated with control, interperitoneal, or orally administered antibiotics. Offspring were raised on normal or western diet. Organs were harvested at 30 weeks of age, flash-frozen, and stored at -80C until RNA extraction.

overall_design: The custom probe panel is a mouse version of platform GPL26764, with pathway enriched selection of KEGG pathway genes for tight junction, regulation of actin cytoskeleton, and colorectal cancer.

first: Simon

last: Turkington

first: Stephanie

last: Prescott

first: Wendy

last: Henderson

first: Giorgio

last: Trinchieri


title: Systematic mapping of 2’-O-methylation sites in rRNAs implicates new strategy for ribose methylation in Arabidopsis [FIB CLIP]

first: Chenjiang

last: You

overall_design: RNAs associated FIB1/2 were immunoprecipitated and extracted from 14-day-old Arabidopsis seedlings expressing GFP-FIB1 and CFP-FIB2. GFP was used as negative control. Four replicates were included.

first: Yong

last: Zhang

first: Runlai

first: Shaofang

last: Li


title: Next Generation Sequencing Facilitates Quantitative Analysis of uterine NK cell transcriptomes from Pbx1f/f and Pbx1f/f; Ncr1Cre mice

city: Hefei

first: Zhou

last: Yonggang

state: Anhui

address: 443 HuangShan Road

department: School of Life Science and Medical Center

institute: University of Science and Technology of China

summary:
Purpose: Next-generation sequencing (NGS) has revolutionized systems-based analysis of cellular pathways. The goals of this study are to investigatethe effect of the Pbx1 gene on the gene expression program of mature uterine NK cells.
Methods: The differential analysis of gene expression profiles in uterine NK cells from Pbx1f/f and Pbx1f/f; Ncr1Cre mice at gd11.5 by RNA-Seq .
Results: Using an optimized data analysis workflow, we found that the inactivation of PBX1 altered the NK cell gene expression program, and many gene expression changes, including significant down-regulation of Ptn and Ogn expression.
Conclusions: PBX1 not only regulates the proliferative capacity of NK cells but also promotes the expression of GPFs in NK cells.

overall_design: Uterine NK cells mRNA profiles of Pbx1f/f and Pbx1f/f; Ncr1Cre mice at gd11.5 were generated by deep sequencing, in duplicate, using Illumina HiSeq X Ten.

first: Yonggang

last: Zhou

first: Binqing

last: Fu

first: Xiuxiu

last: Xu

first: Haiming

last: Wei


title: Transcriptional profiling analysis of C.albicans mnn9D/D null mutant [K060]

city: Salamanca

first: Angel

last: Domínguez

state: Salamanca

address: Plaza de los Doctores de la Reina

department: Microbiology and Genetics/CIETUS/IBSAL

institute: University of Salamanca

summary: Whole genome microarray were used to generated the transcriptional profile of Candida albicans mnn9D/D null mutant vs. isogenic strain CAI4 (WT)

overall_design: Comparisons of expression profiles of mnn9D/D mutant and isogenic wild-type were performed in exponential growth at 28ºC (grown in yeast form) and under filamentation-inducing conditions at 37ºC for 15, 60 and 180 min. Three replicates were performed, corresponding to 2 biological independent replicates (Rep1 and Rep 3) and one dye swap experiment (Rep 2). Eurogentec arrays, bach K060H.

first: Carlos

last: Cívicos

first: M

middle: C

last: López


title: Transcriptional profiling analysis of C.albicans mnn1D/D and mnn9D/D null mutants


title: Transcriptional profiling analysis of C.albicans mnn9D/D null mutant [B200H]

overall_design: Comparisons of expression profiles of mnn9D/D mutant and isogenic wild-type were performed in exponential growth at 28ºC (grown in yeast form). Two samples were performed, corresponding to clon 1 of mutant (Rep1 and Rep 2) and one dye swap experiment (Rep 2). Eurogentec arrays, bach B200H.


title: Transcriptional profiling analysis of C.albicans mnn1D/D null mutant

summary: Whole genome microarray were used to generated the transcriptional profile of Candida albicans mnn1D/D null mutant vs. isogenic strain CAI4 (WT)

overall_design: Comparisons of expression profiles of mnn1D/D mutant and isogenic wild-type were performed in exponential growth at 28ºC (grown in yeast form) and under hypha-inducing conditions at 37ºC for 15, 60 and 180 min. Three replicates were performed, corresponding to 2 biological independent replicates (Rep1 and Rep 3) and one dye swap experiment (Rep 2). Eurogentec arrays, bach I180H.


title: Deep sequencing of NPKO-1 cells with NRDE2 knockdown

first: Hong

last: Cheng

address: 320 YueYang Road

department: Shanghai Institute of Biochemistry and Cell Biology

institute: Shanghai Institutes for Biological Sciences

summary: To study whether NRDE2 downregulation affects the expression of mitotic genes, we carried out NRDE2 knockdown in NPKO-1 cells. The mitotic cells were shaken off, and the rest of cells were harvested for RNA extract and deep sequencing.

overall_design: NPKO-1 cells were treated with control or NRDE2 shRNA for 72 hrs, and then the mitotic cells were shaken off, and the rest cells were harvested for RNA deep sequencting

first: Guifen

last: Wu

first: Xingya

last: Chang

first: Xudong


title: Transcriptional profiling analysis of C.albicans sin3 null mutant

summary: Whole genome microarray were used to generated the transcriptional profile of Candida albicans sin3 null mutant vs. isogenic strain CAI4

overall_design: Comparisons of expression profiles of sin3 mutant and isogenic wild-type were performed in exponential growth at 28ºC (grown in yeast form) and under hyphae-inducing conditions at 37ºC for 15, 60 and 180 min. Four replicates were performed, corresponding to 2 biological independent replicates (Rep1 and Rep 3, Rep 2 and Rep 4) and two dye swap experiments for each one (Rep 1 and Rep 2, Rep 3 and Rep 4). Eurogentec arrays, bach L040B, bach I010C, bach K220D, bach G070D and bach I270D.

first: Nuria

last: Laso


title: NRDE2 acts as a negative regulator of the exosome complex by disrupting hMTR4 interactome

summary: The RNA exosome complex is the most versatile RNA-degradation machine in eukaryotes that requires MTR4 for every aspect of its nuclear functions.  To date, how exosome functions are negatively regulated remains largely unknown.  Here, we report the identification of NRDE2 as an inhibitory regulator of exosome functions.  NRDE2 tightly and directly interacts with human MTR4 (hMTR4) via its N-terminal domain, resulting in their mutual stabilization.  Overexpression of NRDE2 attenuates associations of hMTR4 with the exosome, CBC, TRAMP and NEXT components, resulting in accumulation of exosome target RNAs as well as 3’ extended 5.8S rRNAs.  Conversely, in NRDE2 knockdown cells, the association of hMTR4 with CBC, TRAMP or NEXT components are significantly enhanced, accompanied by promoted RNA degradation.  Together, our work uncovers NRDE2 as a direct negative regulator of exosome functions that prevents hMTR4 from associating with its interacting partners.

overall_design: RNA levels were compared when NRDE2 is over expressed or knocked down by deep sequencing.

first: Xundong

first: Guohui


title: Transcriptiome sequence of Agaricus blazei mycelium and fruit body primodium

city: Fuzhou

first: Meiyuan

last: Chen

state: Fujian Province

address: No.10.95th Lane, Qianheng Rd

department: Institute of Edible Fungi

institute: Fujian Academy of Agricultural Sciences

summary: To investigate gene expression at different development stages of Agaricus blazei, we utilized Illumina Hiseq PE150 sequencing technology to obtain the transcriptome of mycelium and fruit body primodium

first: Yuanping

last: Lu

first: Jianhua

last: Liao


title: Reseveratrol and Rosiglitazone regulation of retroperitoneal tissue gene expression in ZDF rats

city: Guelph

first: David

last: Mutch

state: Ontario

address: 50 Stone Road East

department: Human Health & Nutritional Sciences

country: Canada

institute: University of Guelph

summary: The goal of this work was to examine if reserveratrol or rosiglitazone treatment could improve the metabolic status of obese male ZDF rats after 6 weeks. Gene expression was analyzed in several key metabolic tissues, including liver, various white adipose tissue depots, red tibalus muscle, and peripheral blood mononuclear cells.

overall_design: A total of 5 microarrays were run for each group, for a total of n=20 samples. For each sample, 100 ng of total RNA was prepared for hybridization to Affymetrix Rat Gene 2.1 ST array strips, according to the manufacturer’s instructions (Affymetrix Inc., Fremont, CA, USA).

first: Monica

middle: H

last: Wong

first: Brittany

last: MacIntyre


title: Reseveratrol and Rosiglitazone regulation of subcutaneous tissue gene expression in ZDF rats


title: Apoptosis, autophagy and senescence protect BRAFV600E nevi from malignant transformation

city: Biella

first: Giovanna

last: Chiorino

state: Biella

address: via malta 3

country: Italy

institute: fondazione edo ed elvo tempia onlus

summary:
The oncogenic mutation BRAFV600E is a common event in nevi and melanomas, which  are aggressive skin tumors characterized by MAPK signaling pathway activation.  It has been observed that mutated benign melanocytic lesions can remain unchanged for decades, but also proliferate or undergo oncogene-induced senescence.
The purpose of this study was to investigate the presence of a gene expression signature in BRAFV600E mutated nevi compared to wild-type ones, all derived from sun exposed sites.
Microdissected tissues from excisional biopsies of acquired nevi were analyzed to detect the presence of BRAF and NRAS mutations and to profile whole genome expression by means of oligonucleotide microarrays. BRAFV600E mutation was evidenced in 64% of nevi, while no NRAS mutations were detected.
Functional analysis of genes differentially expressed between wild-type and mutated lesions pointed out the role of oxidative stress in causing the oncogenic BRAF mutation and that the direct consequence of constitutive activation of BRAF-MEK-ERK pathway, in a benign contest, is the activation of apoptosis, autophagy and senescence. Our data also indicate that in mutated lesions, p53 plays a crucial role in the maintenance of the benign status.

overall_design:
In our study, in order to understand the role of BRAF mutation in acquired nevi, we performed gene expression profiling analysis on fourteen lesions derived from intermittently exposed sites, using whole genome oligonucleotide microarrays. Expression data were coupled with sequencing results on BRAF and NRAS genes and class comparison algorithms were applied to detect sequences differentially expressed between BRAFV600E and BRAFwt-NRASwt lesions. Some of the obtained results were further validated by Real-Time RT-PCR on an independent cohort of samples.
Mutation analysis evidenced BRAF mutations in 64% of the common acquired nevi.
All the mutations consisted in the substitution V600E (T1799A). No alteration was found on exon 11 of the BRAF gene nor on exons 1 and 2 of the NRAS gene, as expected since no lesion came from chronically exposed sites.
Transcriptome analysis was carried out using oligo-microarrays and differential expression was tested applying the Linear Models package LIMMA available at www.bioconductor.org that uses Bayesian statistics to compute the probability of a gene being differentially expressed. Only 25 transcripts had B value greater then 2.
We then performed a one-way ANOVA, dividing samples into three classes according to their BRAF mutational status (wt, ++, +++) as assessed by pyrosequencing. We found 2882 known transcripts by selecting those with ANOVA p-value<0.01 and increasing or decreasing expression trends along the three classes.
Functional analysis was performed using the MetaCoreTM software package to identify  overrepresented functional processes.
BRAF signature, acquired nevi, gene expression profiling

"++ and +++" refer to the percentage of alleles carrying the mutation in the
*BRAF*V600E lesions. This has been assessed by pyrosequencing analysis
(Venesio et al, 2008. PMID= 18408659).

first: Maria

last: Scatolini

first: Paola

last: Ostano

first: Maurizia

last: Mello-Grand

first: Enrico

last: Grosso

first: Chiara

last: Ghimenti

first: Antonella

last: Balsamo

first: Tiziana

last: Venesio

first: Mauro

last: Risio


title: Identification of distinct molecular signatures in BRAF V600E and BRAF wt cutaneous melanomas

summary: The purpose of this study was to investigate the presence of a gene expression signature in BRAF V600E melanomas compared to wild type ones, all derived from sun-exposed sites. Microdissected tissues from excisional biopsies of 18 cutaneous melanomas were analyzed to detect the presence of BRAF and NRAS mutations and to profile whole genome expression by means of oligonucleotide microarrays. Class comparison methods were used to select differentially expressed genes between wild type and mutated lesions. Real Time RT-PCR and immunohistochemistry were applied to validate differences at the mRNA and protein levels on an independent cohort of samples. BRAF mutations were evidenced in 67% of melanomas. All of them consisted of the oncogenic change V600E (and the mutation event was independent of Clark's level). Data indicate that in V600E melanomas there is an over-expression of cancer stem cell markers and an upregulation of important oncogenes, like KRAS. This, together with the downregulation of genes involved in oxidative UV stress response and immuno system defense, confers an advantage to V600E melanomas compared to wt lesions. Moreover, the downregulation of topoisomerase I and CDKN2A results in an increased replicative potential associated with a decrease in senescence markers and a diminished DNA damage response. As far as the wild-type lesions, we interestingly pointed out the overexpression of PML, PIK3CA and the downregulation of two tumour suppressor genes (BRCA1 and TP73) relevant to DNA repair.

overall_design: A total of 18 excisional biopsies from primary radial growth phase malignant melanomas (n = 5) and primary vertical growth phase malignant melanomas (n = 13) were examined. Fresh melanoma specimens were examined after removal by a pathologist and a representative portion of the lesion was immediately put in RNA later (RNA Stabilization Reagent – Ambion Inc, Austin TX) for further molecular  processing. Lesions were classified according to TNM. All tumor samples and clinical data were collected according to the IRCC ethical commitee’ s approval. Informed consent was also obtained for each patient. All the samples were homogenized (Ultra – TurraxT8, Ika – Werke, Staufen, Germany) with 600 ul of RNA lysis buffer plus 1% 2-mercaptoethanol, and total RNA was isolated by using RNeasy Mini Kit (Qiagen, Dusseldorf, Germany). TotRNA quality was checked by means of RNA 6000 pico chip assays (Agilent Technologies, Palo Alto, CA) run on the Agilent 2100 bioanalyzer. Paraffin-embedded tissues derived from each lesion were subjected to laser microdissection by means of optical microscope or laser microdissector (Leica AS-LMS, Leica Microsystems, Wetzlar, Germany) and  DNA was extracted from microdissected cells by using QIAamp Kit (Qiagen) as described by Venesio et al. (2008). BRAF exon 11 and 15 and NRAS exons 1 and 2 were amplified by PCR using primers and conditions as previously reported by Venesio et al. (2008). PCR products were purified by ExoSAP-IT (USB Corporation, Cleveland, Ohio) and sequenced using Big Dye Terminator V3.1 Cycle Sequencing Kit and Prism Model 3730 DNA Analyzer (Applied Biosystems, Foster City, CA). Sequence analysis was based on BRAF and NRAS sequences (GenBank accession nos. NM_00433 and NM_002524 for BRAF and NRAS, respectively). RNA from each sample and from the human universal reference (BD™ Human Universal Reference Total RNA, Clontech, Palo Alto, CA) was amplified by means of the Amino Allyl MessageAmp I aRNA Kit (Ambion) to obtain amino allyl antisense RNA (aaRNA) following the method developed by Eberwine and coworkers (1990). Two rounds of amplification were carried out to obtain the necessary quantity of aaRNA for labeling. Briefly: mRNA was reverse transcribed into cDNA single strand; after the second strand synthesis (in the second round of amplification), cDNA was in vitro transcribed in aaRNA including amino allyl modified nucleotides (aaUTP). Both dsDNA and aaRNA underwent a purification step using columns provided with the kit. Labeling was performed using NHS ester Cy3 or Cy5 dyes (Amersham Biosciences, Buckinghamshire UK) which are able to react with the modified RNA. mRNA quality was checked by means of RNA 6000 nano chip assays (Agilent Technologies). At least 5 ug of mRNA for each sample were labeled and purified with columns. Equal amounts (0.75 ug) of labeled specimens from sample and reference were put together, fragmented and hybridized to oligonucleotide glass arrays representing 41K human unique genes and transcripts (Human Whole Genome Oligo Microarray Array, Agilent Technologies). All steps were performed using the In Situ Hybridization kit-plus (Agilent Technologies) and following the 60-mer oligo microarray processing protocol (Agilent Technologies). Then, slides were washed with the SSPE wash procedure and scanned with the dual-laser microarray scanner Agilent G2505B. For each sample, a dye-swap replicate was performed. Images were analyzed using Feature Extraction software (Agilent Technologies) version 7.6. Output files containing feature and background intensities and the related statistical parameters for red and green signals were then loaded into the Resolver SE System (Rosetta Biosoftware, Seattle, WA) together with the scan images and the Agilent Human Whole Genome pattern file. Data processing and normalization were performed as previously described (Scatolini et al. 2010).  To compare class expression patterns between BRAF mutated samples and wild-type ones t-test and empirical Bayes statistics (LIMMA package) for two class comparison were applied. Transcripts with significant modulation, B value greater than 2 for empirical Bayes statistic, were further considered. A B value greater than 2 means that the gene has more than 88% probability of being differentially expressed by assuming that 1% of the genes are expected to be differentially expressed.  Moreover, we applied to the ratio experiment a one-way ANOVA available within the Resolver System with Multiple Test Correction method (Benjamini Hochberg FDR). Three classes were considered for the ANOVA (wt, ++, +++). For all the transcripts with significant modulation (p-value<0.01), we looked for those with increasing or decreasing expression and, in particular, with an absolute log ratio difference >0.7 between the wt class and the +++ one. In order to underline the enrichment of Biological  Processes-level 5 (BP5) of the Gene Ontology (GO), we used the functional annotation tool available within DAVID website, using the Entrez Gene ID of modulated transcripts (p-value< 0.01). Several databases were interrogated to find out and compare functional information on single genes and on groups of differentially expressed genes. In particular, NCBI database (http://www.ncbi.nlm.nih.gov/) and UCSC Genome Bioinformatics website were used (http://genome.ucsc.edu/). Quantitative RT polymerase chain reaction (qPCR) was performed, as previously described (Scatolini M et al. 2010), to validate a subset of differently expressed transcripts between BRAF mutated and wt samples, identified by microarray analysis.


title: Lysine 5 acetylation of histone H2A associates with cell-type specific regulatory networks in human chromatin

city: Vienna

first: Iros

last: Barozzi

address: Borschkegasse 8a

country: Austria

institute: Medical University Vienna

type: Genome binding/occupancy profiling by genome tiling array

summary:
The identification of structural determinants of chromatin predicting cell-type, stage- and disease-specific functional networks is a major task of epigenomic studies. Histone post-translational modifications (PTMs) are critical determinants of chromatin accessibility at active cis-regulatory DNA, but they have been found distinctively associated with functional elements only in a few cases.
In this study, we have exploited the availability of a validated dataset of promoter and non-promoter Nuclease Accessible Sites (NAS), to dissect through quantitative chromatin immunoprecipitation (qChIP) assays the correlation between 25 histone PTMs and cis-regulatory DNA. Our data show that a combinatorial pattern of histone PTMs is associated mainly with human promoters, and that lysine 5 acetylation of histone H2A (H2AK5ac) is found at functional elements independently from their genomic position. Genome-wide experiments (both ChIP-chip and ChIP-seq were employed) revealed a preferential association of H2AK5ac, and not of its tri-acetylated variant H2A.Z (K4+K7+K11Ac), with CpG islands, DNaseI-hypersensitive sites (DHS) and sites of mono-methylation of lysine 4 of histone H3 (H3K4me1). We demonstrate that H2AK5ac distribution well correlates with active gene expression, in a cell-type and stage-specific manner. Modulation of H2AK5ac during cell differentiation often occurs at sites of H3K4me1, and allowed the localization of developmentally regulated enhancers as well as large-scale chromosomal transitions. Genomic distribution and functional studies showed that H2AK5ac is partially dependent on, but not limited to the enhancer-binding acetyl-transferase p300. Analysis of inducible H2AK5ac sites allowed the computational prediction of binding to enhancer regions by transcription factors regulating cell fate and tumor suppression, which was then experimentally validated.
Our findings provide further insights into the functional links between chromatin accessibility and histone PTMs at cis-regulatory elements during biological processes, and pinpoint H2AK5ac as a novel marker for the functional annotation of the epigenome, in a simple and cost-effective manner.

overall_design: ChIP against H2AK5ac was performed in a CD4+ (one biological replicate) and a PR9 (two biological replicates) cell lines. ChIP against H2A.ZK(4-7-11)ac was performed in a CD4+ cell line (one biological replicate). ChIP products were then PCR amplified, labeled with Cy3/Cy5 fluorescent dyes and hybridized to the NimbleGen custom made NGS_HG17_Chr.19Array. Non-immunoprecipitetaed genomic DNA from the same cell lines served as control for non-specific antibody binding.

first: Gaetano

last: Gargiulo

first: Gianmaria

last: Frigè

first: Matteo

last: Cesaroni

last: Romanenghi

last: Biancotto

first: Pier

middle: G

last: Pelicci

first: Hendrik

last: Stunnenberg

first: Saverio

last: Minucci


title: ERα governs a specific gene expression program and metabolic function in white adipose tissue.

city: Sherbrooke

first: Nicolas

last: Gévry

state: Québec

address: 2500, boulevard de l'Université

department: Biologie

institute: Université de Sherbrooke

summary: The White Adipose Tissue (WAT) has a major influence on energy metabolic functions and expresses the estrogen receptor ERα. Ovariectomized mice or mice carrying an ERα knock-out gene (ERKO), have a higher risk of becoming obese and developing metabolic disorders such as diabetes or dyslipidemia. To identify the direct target genes of ERα in visceral WAT, we treated ovariectomized mice with estradiol (E2) for 2 hours and proceeded to a genome-wide analysis of ERα binding sites to systemically identify them. We demonstrate that the E2/ERα pathway increases the expression of genes involved in the balance between lipids and glucose catabolism, the energy expenditure through the TCA cycle and the respiratory membrane chain. ERα binding sites and gene expression analysis in visceral and subcutaneous WAT, brown adipose tissue and liver show a specific program for each tissue. Our results are consistent with E2/ERα signaling protecting against metabolic diseases such as obesity by regulating an important gene regulatory network that helps maintaining normal metabolic functions in WAT.

overall_design: ChIP-seq and RNA-seq

first: Martin

last: Morin


title: Spinal neuronal ferroptosis aggravates peripheral nerve injury-induced neuropathic pain via modulating competing endogenous RNA axis

status: Public on Dec 30 2024

city: jining

first: gao

last: xu

state: Shandong

address: 89 Guhuai Road

institute: Affiliated Hospital of Jining Medical University

summary: Peripheral nerve injury-induced neuropathic pain presents a significant clinical challenge and the pathogenesis remains unclear. Recent findings have implicated spinal neuronal ferroptosis in the modulation of neuropathic pain progression, but the molecular mechanism is inadequately understood. This work constructed competitive endogenous RNA (ceRNA) regulatory network in neuronal ferroptosis model through total transcriptome sequencing. Bioinformatic analyses elucidated the biological roles and signaling pathways of differentially expressed genes associated with neuronal ferroptosis. In vitro experiments substantiated the involvement of two critical regulatory axes, namely lncRNA-Sh3bp5l/miR-92b-5p/Pgd and lncRNA-Scrn2/miR-133a-3p/Srxn1, in the regulation of neuronal ferroptosis. The knockdown of PGD and SRXN1 was observed to significantly inhibit the ferroptosis phenotype. Utilizing an in vivo chronic constriction injury (CCI) pain model, we demonstrated that the expression of ceRNA-mediated downstream targets, PGD and SRXN1, was localized within spinal neurons.  Furthermore, lncRNA-Sh3bp5l and lncRNA-Scrn2 were found to be upregulated in the ipsilateral spinal cord following CCI injury, accompanied by increased levels of PGD and SRXN1. In brief, this study explored the key ceRNA axis that regulates spinal neuronal ferroptosis, which might serve as the potential therapeutic targets for neuropathic pain.

overall_design: miRNA sequencing of retinoic acid-induced N2a cells with or without RSL3 stimulation

first: Xu

last: Gao

first: Xiao

last: Liang

first: Longfei

first: Chunyang

last: Meng


title: Iron regulatory protein 1 deficient mice exhibit hypospermatogenesis

city: Giessen

first: Marek

last: Bartkuhn

state: Hessen

address: Aulweg 132

department: Biomedical Informatics and Systems Medicine

institute: Justus-Liebig-University Giessen

summary: Imbalances in testicular iron levels are linked to compromised sperm production and male infertility. Iron regulatory proteins (IRP) 1 and 2 play crucial roles in cellular iron regulation. We investigated the role of IRP1 on spermatogenesis using Irp1-deficient mice (Irp1-/-). Histological analysis of the testis of Irp1-/- mice revealed hypospermatogenesis with a significant reduction in the number of elongated spermatids and daily sperm production compared to wild-type (WT) mice. Flow cytometry of germ cells from WT and Irp1-/- mice showed reduction in spermatocytes, round and elongated spermatids, which was confirmed by histological and immunofluorescence quantification. Finally, stage VIII of spermatogenesis, crucial for spermatid maturation, was less frequent in Irp1-/- testicular cross-sections. Hypospermatogenesis worsened with age despite unchanged intratesticular iron levels. Mechanistically, this was due to increased oxidative stress indicated by elevated 8-Oxoguanine (8-OxoG) levels, a DNA lesion resulting from reactive oxygen species (ROS). Furthermore, bulk RNA-seq data indicated compromised DNA damage repair and cell cycle processes, including mitosis and meiosis, which may explain hypospermatogenesis.Our results suggest that IRP1 deletion leads to hypospermatogenesis due to impaired cell cycle progression, decreased DNA damage repair capacity and oxidative damage. Altogether, this study uncovers a role for IRP1, independent of traditional iron regulation mechanisms.

overall_design: Postpubertal mice (20 weeks old) were euthanized by 5% isoflurane inhalation following cervical dislocation. Afterwards, testes were harvested, decapsulated and total RNA was isolated. After obtaining the RNAseq data, comparative gene expression analyses between Irp1 deficient and wild type were performed. To identify changes in gene expression related to spermatogenesis.

first: Aileen

last: Harrer

first: Niraj

last: Ghatpande

last: Grimaldini

first: Daniela

last: Fietz

first: Vishnu

last: Kumar

first: Christiane

last: Pleuger

first: Monika

last: Fijak

first: Dankward

last: Föppl

first: Lennart

last: Rynio

first: Hans-Christian

last: Schuppe

first: Adrian

last: Pilatz

first: Tara

last: Procida-Kowalski

first: Noga

last: Guttmann-Raviv

first: Sudhanshu

last: Bhushan

first: Esther

last: Meyron-Holtz

first: Andreas

last: Meinhardt


title: Dehydration induced Ae-Aper50 regulates midgut infection in Aedes aegypti mosquitoes

city: Galveston

first: Haiping

last: Hao

state: Texas

address: 301 University Blvd

institute: UTMB NGS Core

summary: Identify genes expressed in response to dehydration

overall_design: Female Aedes aegypti whole bodies from the THI and PK10 lines from Senegal were harvested 24 hours post exposure to acute desiccation stress or control conditions and homogenized in trizol for RNA extraction and sequencing.

first: Anastasia

last: Accoti

first: Margaret

last: Becker

first: Angel Elma I.

last: Abu

first: Julia

last: Vulcan

first: Ruimei

last: Yun

first: Steven G.

last: Widen

first: Massamba

last: Sylla

first: Vsevolod L.

last: Popov

first: Laura B.

last: Dickson


title: Inhibition of tumor-intrinsic NAT10 enhances antitumor immunity by triggering type I interferon responses via MYC/CDK2/DNMT1 pathway.

city: Suzhou

first: Wancheng

last: Liu

state: jiangsu

address: 100 Chongwen Road, Industrial Park

institute: Chinese Academy of Medical Sciences and Peking Union Medical College

summary: Purpose:Posttranscriptional modifications are deeply involved in cancer progression; however, there remains knowledge gap regarding the function and immune regulatory mechanism of newly discovered mRNA acetylation modification.Methods: The expression of NAT10 in human tumor tissues was analyzed based on TCGA data. NAT10 knockout murine cell lines were generated to analyze the NAT10-associated tumor progression and anti-tumor immune response in tumor xenograft models. Immune cells and cytokines in TME were quantified by immunofluorescence, flow cytometry, quantitative reverse transcription-PCR, and ELISPOT assay. NAT10-associated differentially expressed genes were investigated in cancer cells and tissues with RNA-Seq. Results:Loss of tumoral NAT10 significantly stimulated tumor-specific cellular immune responses and suppressed tumor growth. Mechanistically, we identified MYC as a key downstream target of NAT10 via enhancing mRNA stability and translation efficiency. Inhibition of NAT10 blocked the MYC/CDK2/DNMT1 pathway, subsequently enhancing double-stranded RNAs (dsRNA) formation, which triggered type I interferon responses to enhance the in vivo response of tumor specific CD8+ T cells. Conclusions: Inhibition of NAT10 using either small molecule inhibitor (Remodelin) or PEI/PC7A/siNAT10 nanoparticles combined with PD-1 blockade synergistically enhanced the anti-tumor immune response and repressed tumor progression. Our findings uncovered the crucial role of tumor-intrinsic NAT10 in tumor immune microenvironment, representing a promising target for enhancing cancer immunotherapy.

overall_design: mRNA profiles of TC1 wild type (WT) and sgNAT10 or sgMyc cell lines; acRIP-seq of TC1 wild type (WT) and sgNAT10 cell line; Ribo-seq of TC1 wild type (WT) and sgNAT10 cell line.

first: W

first: Y

last: Wei

first: J

last: Chen

first: D

last: Ma


title: Leptin signaling maintains autonomic stability during severe influenza infection

city: Boston

first: CBDM

last: Lab

state: MA

address: 77 Avenue Louis Pasteur

department: Microbiology and Immunobiology

institute: Harvard Medical School

summary: Leptin is a pleiotropic hormone that is released by adipocytes and tightly controls energy balance by regulating a range of physiological processes. While leptin is best known for signaling through the hypothalamic arcuate nucleus to regulate feeding, it also supports the immune system by binding its cognate receptor Ob-R to activate various immunocyte populations. Indeed, patients with congenital leptin or leptin receptor deficiency not only display severe obesity but also exhibit increased mortality from infections. Leptin supplementation to leptin deficient mice and humans increases CD4+ cell numbers and augments proinflammatory Th1 responses ex vivo. However, whether this leptin-induced immune response is necessary for viral clearance in vivo is unknown. To better delineate the role of leptin in infection, we inoculated leptin deficient ob/ob mice with influenza and acutely supplemented leptin in physiologically relevant doses prior to infection. Although ob/ob mice and diet-induced obese (DIO) mice have comparable levels of insulin resistance and adiposity, a sublethal dose of influenza virus in DIO mice causes mortality in approximately 80% of ob/ob mice, which was completely reversed by leptin supplementation. Serum cytokine profiling and whole tissue RNA sequencing of spleen, lung, and visceral adipose tissue demonstrated increased inflammatory and anti-viral responses in leptin-supplemented mice and no signs of excessive immunopathology in either group. Further flow cytometric analysis indicated an elevated Th1 immune response locally and systemically in leptin-supplemented mice, along with a two-fold increase in spleen sizes and an expansion in the proportion of splenic B cells. Despite augmented anti-viral responses in leptin-supplemented mice, leptin deficient ob/ob mice were equally capable of clearing the virus. Moreover, pulse oximetry showed diminished oxygen saturation in both groups, a finding further corroborated by similar levels of histopathologic lung damage. Interestingly, leptin-supplementation in ob/ob mice prevented profound hypothermia and bradycardia, which occurred prior to death in control ob/ob mice. Taken together, we provide evidence that leptin is dispensable for immunologic control of viral infection and may instead confer a survival benefit by maintaining autonomic stability. These findings open new avenues for research on the role of leptin signaling in infection.

overall_design: RNAseq on lung and spleen of ob/ob mice supplemented with leptin or saline controls and infected with influenza

first: Andrés

last: Muñoz-Rojas

first: Adam

last: Wang

first: Lisa

middle: E

last: Pomeranz

first: Jeffrey

last: Friedman

first: Diane

last: Mathis

first: Kartik

middle: N

last: Rajagopalan