title: Next-generation interaction screening to discover luciferase (construct #1) related protein-protein interactions regulating barley powdery mildew disease immunity and susceptibility

first: Roger

last: Wise

state: IA

summary: This luciferase (construct # 1) dataset was part of a larger Y2H screen (GEO GSE150396) to identify novel proteins involved in barley immune responses. Yeast-two-hybrid (Y2H) screening was coupled with next-generation sequencing to identify and quantify interacting proteins. The luciferase (construct # 1) bait was mated to a cDNA prey library derived from a 0-48 hour time course of infected leaf tissue. Screens were performed in batch liquid culture to enrich yeast populations for cells expressing positive interactions. After two rounds of enrichment under selective (Histidine absent) and non-selective (Histidine present) conditions, yeast cells were collected. Y2H plasmids were extracted and prey cDNA amplicons were generated via low-cycle PCR. Fragmented amplicons were used as input to generate sequencing libraries and processed on the HiSeq 3000 platform. Reads were mapped to the barley and Blumeria graminis genomes and read counts were analyzed using a custom data processing and scoring pipeline. Putative interactors were cloned and binary Y2H was used to confirm interactions.

overall_design: Y2H screening of a cDNA library was coupled to next-generation sequencing to identify and rank positive protein-protein interactions. Screening for the luciferase (contruct # 1) bait was conducted in triplicate with yeast cells grown in selective and non-selective conditions. NuGEN’s AnyDeplete technology was used to deplete HORVU3Hr1G072260 (zinc finger protein) from the samples. This dataset represents one of two luciferase constructs that were used as bait negative controls. Samples were run on three lanes of Illumina’s HiSeq 3000.

first: Sagnik

last: Banerjee

first: Valeria

last: Velasquez-Zapata

first: James

last: Elmore

first: Gregory

last: Fuerst


title: Bcl11b binding and transcriptome analysis of Bcl11b-deficient MAIT cells at steady state and infection

status: Public on Mar 22 2021

city: Tampa

first: Dorina

last: Avram

state: Fl

address: 12902 Magnolia Drive, MRC4

department: Department of Immunology

country: USA

institute: Moffitt Cancer Center

type: Expression profiling by high throughput sequencing

type: Genome binding/occupancy profiling by high throughput sequencing

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

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Refer to individual Series


title: Next-generation interaction screening to discover luciferase (construct #2) related protein-protein interactions regulating barley powdery mildew disease immunity and susceptibility

status: Public on Mar 23 2021

city: Ames

address: Department of Plant Pathology, Entomology, and Microbiology, Iowa State University, 1344 Advanced Teaching & Research Bldg., 2213 Pammel Dr.

department: CICGRU

institute: USDA/ARS

summary: This luciferase (construct # 2) dataset was part of a larger Y2H screen (GEO GSE150396) to identify novel proteins involved in barley immune responses. Yeast-two-hybrid (Y2H) screening was coupled with next-generation sequencing to identify and quantify interacting proteins. The luciferase (construct # 2) bait was mated to a cDNA prey library derived from a 0-48 hour time course of infected leaf tissue. Screens were performed in batch liquid culture to enrich yeast populations for cells expressing positive interactions. After two rounds of enrichment under selective (Histidine absent) and non-selective (Histidine present) conditions, yeast cells were collected. Y2H plasmids were extracted and prey cDNA amplicons were generated via low-cycle PCR. Fragmented amplicons were used as input to generate sequencing libraries and processed on the HiSeq 3000 platform. Reads were mapped to the barley and Blumeria graminis genomes and read counts were analyzed using a custom data processing and scoring pipeline. Putative interactors were cloned and binary Y2H was used to confirm interactions.

sample_organism:
Hordeum vulgare
Blumeria hordei

platform_organism:
Hordeum vulgare
Blumeria hordei

overall_design: Y2H screening of a cDNA library was coupled to next-generation sequencing to identify and rank positive protein-protein interactions. Screening for the luciferase (construct # 2) bait was conducted in duplicate with yeast cells grown in selective and non-selective conditions. NuGEN’s AnyDeplete technology was used to deplete HORVU3Hr1G072260 (zinc finger protein) from the samples. This dataset represents one of two luciferase constructs that were used as bait negative controls. Samples were run on two lanes of Illumina’s HiSeq 3000.

middle: M

middle: P


title: Langerhans cell RNAseq

city: Singapore

first: Christiane

last: Ruedl

state: Singapore

address: 60 Nanyang Drive

country: Singapore

institute: Nanyang Technological University

summary: CD45.1 bone marrow cells were transplanted into irradiated CD45.2 host to generate bone marrow chimera mice. Skin langerhans cells and other dendritic cell subsets were sorted  from chimera mice and sequenced.

overall_design: 3 donor, 3 host derived Langerhans cells, 3 LClike cells and 2 classical dendritic cells were compared.

first: Jianpeng

last: Sheng


title: Next-generation interaction screening to discover Mla6aa550-956 related protein-protein interactions regulating barley powdery mildew disease immunity and susceptibility.

summary: The barley R gene (Mla6aa550-956) dataset was part of a larger Y2H screen (GEO GSE150396) to identify novel proteins involved in barley immune responses. Yeast-two-hybrid (Y2H) screening was coupled with next-generation sequencing to identify and quantify interacting proteins. The Mla6aa550-956 bait was mated to a cDNA prey library derived from a 0-48 hour time course of infected leaf tissue. Screens were performed in batch liquid culture to enrich yeast populations for cells expressing positive interactions. After two rounds of enrichment under selective (Histidine absent) and non-selective (Histidine present) conditions, yeast cells were collected. Y2H plasmids were extracted and prey cDNA amplicons were generated via low-cycle PCR. Fragmented amplicons were used as input to generate sequencing libraries and processed on the HiSeq 3000 platform. Reads were mapped to the barley and Blumeria graminis genomes and read counts were analyzed using a custom data processing and scoring pipeline. Putative interactors were cloned and binary Y2H was used to confirm interactions.

overall_design: Y2H screening of a cDNA library was coupled to next-generation sequencing to identify and rank positive protein-protein interactions. Screening for the Mla6aa550-956 bait was conducted in triplicate with yeast cells grown in selective and non-selective conditions. NuGEN’s AnyDeplete technology was used to deplete HORVU3Hr1G072260 (zinc finger protein) from the samples. Two constructs of luciferase were used as bait negative controls. Samples were run on three lanes of Illumina’s HiSeq 3000.


title: Regulation of 3D genome organization, transcription and histone H3K9me2 by histone H3K9 methyltransferases [ChIP-Seq]

status: Public on Mar 18 2021

city: Saitama

first: Kei

last: Fukuda

state: Japan

address: 2-1 Hrosawa, Wakoshi

country: Japan

institute: RIKEN

summary: Histone H3 lysine 9 dimethylation (H3K9me2) is a highly conserved silencing epigenetic mark. Chromatin marked with H3K9me2 forms large domains in mammalian cells and overlaps well with lamina-associated domains and the B compartment defined by Hi-C. However, the role of H3K9me2 in 3-dimensional (3D) genome organization remains unclear. We investigated genome-wide H3K9me2 distribution, transcriptome, and 3D genome organization in mouse embryonic stem cells following the inhibition or depletion of five H3K9 methyltransferases (MTases): G9a, GLP, SETDB1, SUV39H1, and SUV39H2. H3K9me2 was regulated by all five MTases; however, H3K9me2 and transcription in the A and B compartments were regulated by different MTases. H3K9me2 in A compartments was primarily regulated by G9a/GLP and SETDB1, while H3K9me2 in the B compartments was regulated by all five MTases. Furthermore, decreased H3K9me2 correlated with changes to the more active compartmental state that accompanied transcriptional activation.

overall_design: Examination of H3K9me2, H3K9me3, transcriptome and 3D genome organization in H3K9 methyltransferase deficient mESCs and iMEFs

first: Yoichi

last: Shinkai


title: Regulation of 3D genome organization, transcription and histone H3K9me2 by histone H3K9 methyltransferases [HiC]


title: Regulation of 3D genome organization, transcription and histone H3K9me2 by histone H3K9 methyltransferases [RNA-Seq]


title: Deficiency of histone lysine methyltransferase SETDB2 in hematopoietic cells promotes  vascular inflammation and accelerates atherosclerosis

city: New Haven

first: Xinbo

last: Zhang

state: Connecticut

address: Rm320, 10 Amistad St

department: Comparative Medicine

institute: Yale University

summary: Epigenetic modifications of the genome, including DNA methylation, histone methylation/acetylation  and noncoding RNAs, have been reported to play a fundamental role in regulating immune response  during the progression of atherosclerosis. SETDB2 is a member of the KMT1 family of lysine  methyltransferases and members of this family typically methylate histone H3 Lys9 (H3K9), an epigenetic  mark associated with gene silencing and previous studies have shown SETDB2 is involved in innate and  adaptive immunity, the pro-inflammatory response and hepatic lipid metabolism. Here we report that the  expression of SETDB2 is markedly upregulated in human and murine atherosclerotic lesions. The  upregulation of SETDB2 is observed in pro-inflammatory M1, but not anti-inflammatory M2 macrophages  (MΦ). Notably, we found that genetic deletion of SETDB2 in hematopoietic cells promotes vascular  inflammation and enhances the progression of atherosclerosis in bone marrow transfer studies in LDLR  knockout mice. Single cell RNA-Seq analysis in isolated CD45+ cells from atherosclerotic plaques from  mice with SETDB2 deficient bone marrow revealed a significant increase in inflammatory macrophage  population and enhanced expression of genes involved in inflammation, myeloid cell recruitment and lipid  metabolism. Additionally, we found that loss of SETDB2 in hematopoietic cells is associated with  macrophage accumulation in atherosclerotic lesions, macrophage proliferation and attenuated  efferocytosis. Overall, these studies identify SETDB2 as an important inflammatory cell regulator that  controls macrophage activation in atherosclerotic plaques.

overall_design: scRNA-seq of CD45+ cells from atherosclerotic plaques of Ldlr-/- mouse transplanted with WT or Setdb2-/- bone marrow cells

first: Carlos

last: Fernández-Hernando


title: Regulation of 3D genome organization, transcription and histone H3K9me2 by histone H3K9 methyltransferases


title: let-7e downregulation characterizes early phase colonic adenoma in APCMin/+ mice and human FAP patients

status: Public on Mar 17 2021

city: Bari

first: Elena

last: Piccinin

state: Ba

address: Piazza Giulio Cesare, 11, Palazzo Morgagni - Lab. Clinica Medica Frugoni - II piano

country: Italy

institute: University of Bari "Aldo Moro"

summary: The crypt-villus axis represents the essential unit of the small intestine, which integrity and functions are fundamental to assure tissue and whole-body homeostasis. Disruption of pathways regulating the fine balance between proliferation and differentiation results in diseases development. Nowadays, it is well established that microRNAs (miRNAs) play a crucial role in the homeostasis maintenance and perturbation of their levels may promote tumor development. Here, by using microarray technology, we analysed the miRNAs differentially expressed between the crypt and the villus in mice ileum. The emerged miRNAs were further validated by Real Time qPCR in mouse model (ApcMin/+), human cell lines and human tissue samples (FAP) of colorectal cancer (CRC). Our results indicated that miRNAs more expressed in the villi compartment are negatively regulated in tumor specimens, thus suggesting a close association between these microRNAs and the differentiation process. Particularly, from our analysis let-7e appeared to be a promising target for possible future therapies and a valuable marker for tumor staging, being upregulated in differentiated cells and downregulated in early stage colonic adenoma samples.

overall_design: Analysis of microRNA expression along the ileal crypt-villus axis in four wild type mice perfomed with Illumina iScan System. From the crypt (C1) to the tip of the villus (V0), five different fractions of differently differentiated cells have been analyzed.

first: Annalisa

last: Contursi

first: Maria

last: Arconzo

first: Marica

last: Cariello

first: Marilidia

last: Piglionica

first: Simona

last: D'Amore

first: Giusi

last: Graziano

first: Michele

last: Vacca

first: Raffaella Maria

last: Gadaleta

first: Rosa

last: Valanzano

first: Renato

last: Mariani-Costantini

first: Gaetano

last: Villani

first: Antonio

last: Moschetta


title: Transciptomic profiling of the magnum of laying and non-laying hens using RNA-seq

city: Honolulu

first: Nirvay

last: Sah

state: HI

address: 1955 East-west raod

department: Human Nutrition, Food and Animal Sciences

institute: University of Hawaii at Manoa

summary:
Purpose: With the advent of Next-generation sequencing (NGS), several novel genes/proteins and cellular pathways in wide varitey of tissues has discovered. The aim of this study are to perform transcriptome profiling (RNA-seq) of magnum to determine differently expressed genes in laying and non-laying hens and to further validate the expression of candidate genes using real-time quantitative reverse transcription polymerase chain reaction (qRT–PCR)  in laying, non-laying and molting hens.

Methods: Magnum mRNA profiles of 35-60 weeks-old laying and non-laying hens, three each, were generated with NextSeq 500 sequencer in single-end mode with a read length of 1x76 bp. Raw sequencing reads were cleaned and trimmmed with Prinseq tool and good reads were aligned against the chicken reference gemone (Galgal 5.0) in Array Studio. Differential gene expression analysis was performed by the DESeq2 algorithm as implemented in Array Studio. The genes with at least three-fold change (FC) and Benjamini and Hochberg q-value < 0.05 were called differentially expressed.

Results: Using an optimized data analysis workflow, we mapped about 30.5 million reads from layers and 33.4 million reads from non-layers to the chicken genome. A total of 19,152 gene transcripts were annotated from Ensembl alignment which represents 50.24% of the chicken genome assembly. Differential gene expression analysis showed 540 were differentially expressed between layer and non-layer hens. 152 DEGs were significantly up-regulated and 388 were significantly down-regulated in the laying hens when compared to the non-laying hens.

Conclusions: Our study reports the expression of several pre-discovered and many novel genes that may be involved in the transport of precurosor molecules for biosynthesis and secretion of the egg-white proteins in the magnum. These genes can be used as quantitative basis of selecting hens with an improved egg quality.

overall_design: Magnum mRNA profile of three 35-60 weeks old laying and non-laying hens was genereted using Illumina NextSeq 500

first: Birendra

last: Mishra


title: Expression Profile of Long Noncoding RNAs and Circular RNAs in mouse C3H10T1/2 mesenchymal stem cells undergoing myogenic and cardiomyogenic differentiation

city: jiang su

first: MIngshan

last: Li

state: su zhou

address: Industrial park

country: China

institute: suzhou university

type: Non-coding RNA profiling by high throughput sequencing

summary: Currently, a heterogeneous category of non-coding RNAs (ncRNA) that directly regulate the expression or function of protein-coding genes is shown to have an effect on the fate decision of stem cells. However, the detailed regulatory roles of ncRNAs in myogenic and cardiomyogenic differentiation of mouse C3H10T1/2 mesenchymal stem cells (MSCs) are far from clear. Methods: In this study, 5-azacytidine (5-AZA) treated C3H10T1/2 cells were differentiated into myocyte-like and cardiomyocyte-like cells. Next, ncRNA associated with myogenic and cardiomyogenic differentiation were identified using high-throughput RNA sequencing (RNA-seq) data. Bioinformatics analysis was conducted to identify the differentially expressed ncRNAs and the related signaling pathways. Results: Myotube-like structure was formed after 5-AZA treatment of C3H10T1/2 cells. In addition, myogenic and cardiomyogenic differentiation related genes like GATA4, cTnt, MyoD, and Desmin were up-regulated significantly after 5-AZA treatment. Totally, 1538 differentially expressed lncRNAs and 3398 differentially expressed mRNAs were identified, including 1175 up-regulated and 363 downregulated lncRNAs, 2429 up-regulated and 969 down-regulated mRNAs. In addition, 46 differentially expressed circRNAs were identified, including 25 up-regulated and 21 downregulated circRNAs. Moreover, the differentially expressed mRNAs were enriched into 5 significant pathways, including those for Focal adhesion, ECM-receptor interaction, PI3K-AKT signaling pathway, PPAR signaling pathway and Tyrosine metabolism. Conclusions: A systematic view of the expression of ncRNAs in myogenic and cardiomyogenic differentiation of MSCs was provided in the study.

overall_design: There were 5-Aza treatment and two groups of untreated, and two replicates were performed in each group.

first: Mingshan

first: Zijie

last: Pei

first: Hongtao

last: Zhang

first: Jing

last: Qu


title: Epigenetic Silencing by SETDB1 Suppresses Tumor-cell Intrinsic Immunogenicity

city: Boston

first: Jingyi

last: Wu

state: MA

address: 360 Longwood Ave

department: Department of Cancer Biology

institute: Dana-Farber Cancer Institute

summary: Epigenetic dysregulation is a defining feature of tumorigenesis and has been implicated in immune escape. However, the epigenetic mechanisms that drive immune evasion in cancer are poorly understood. To systematically identify epigenetic factors that modulate the immune sensitivity of tumor cells, we performed in vivo loss of function screens targeting 936 chromatin regulators in mouse tumor models treated with immune checkpoint blockade. We identified the H3K9-methyltransferase SETDB1 and other members of the HUSH and KAP1 complexes as cell-intrinsic mediators of immune escape in tumor cells. We also found that amplification of SETDB1 (1q21) in human tumors is associated with reduced cytotoxic T-cell infiltration and resistance to immune checkpoint blockade. Mechanistically, we demonstrate that SETDB1 targets broad domains, hundreds of kilobases in size, many of which reside within the open genome compartment. These SETDB1 domains are enriched for transposable elements (TEs) and immune gene clusters associated with segmental duplication events, a central mechanism of mammalian genome evolution. SETDB1 loss derepresses latent TE-encoded regulatory elements and proximal immune genes within these repetitive regions, including canonical NKG2D ligands, and induces hundreds of putative TE-encoded viral antigens. Our study establishes SETDB1 as an epigenetic checkpoint that suppresses intrinsic immunogenicity in cancer cells, and thus represents a candidate target for immunotherapy.

overall_design: ChIP-seq, RNA-seq and ATAC-seq were performed on control and Setdb1 KO Lewis Lung Carcinoma (LLC) and B16 melanoma cell lines. RNA-seq was also performed on A375 human melanoma cell line.

first: Gabriel

middle: K

last: Griffin

first: Arvin

last: Iracheta-Vellve

first: Robert

middle: T

last: Manguso

first: Bradley

middle: E

last: Bernstein


title: Transcriptional and Immunologic Correlates of Response to Pandemic Influenza Vaccine in Aviremic, HIV-Infected Children

city: Cleveland

first: Mark

last: Cameron

state: OH

address: 10900 Euclid Ave

department: Population and Quantitative Health Sciences

institute: Case Western Reserve University

summary: People living with HIV (PWH) often exhibit poor responses to influenza vaccination despite effective combination anti-retroviral (ART) mediated viral suppression.  There exists a paucity of data in identifying immune correlates of influenza vaccine response in context of HIV infection that would be useful in improving its efficacy in PWH, especially in younger individuals.  Transcriptomic data were obtained by microarray from whole blood isolated from aviremic pediatric and adolescent HIV-infected individuals (4-25 yrs) given two doses of Novartis/H1N1 09 vaccine during the pandemic H1N1 influenza outbreak.  Supervised clustering and gene set enrichment identified contrasts between individuals exhibiting high and low antibody responses to vaccination. High responders exhibited hemagglutination inhibition antibody titers >1:40 post-first dose and 4-fold increase over baseline. Baseline molecular profiles indicated increased gene expression in metabolic stress pathways in low responders compared to high responders. Inflammation-related and interferon-inducible gene expression pathways were higher in low responders 3 wks post-vaccination. The broad age range and developmental stage of participants in this study prompted additional analysis by age group (e.g. <13yrs and >13yrs). This analysis revealed differential enrichment of gene pathways before and after vaccination in the two age groups. Notably, CXCR5, a homing marker expressed on T follicular helper (Tfh) cells, was enriched in high responders (>13yrs) following vaccination which was accompanied by peripheral Tfh expansion. Our results comprise a valuable resource of immune correlates of vaccine response to pandemic influenza in HIV infected children that may be used to identify favorable targets for improved vaccine design in different age groups.

overall_design: HIV+ individuals were administered pH1N109/10 monovalent vaccine. The objective was to compare gene expression at different timepoints in the context of the serum antibody response to the vaccination.

first: Lesley

middle: R

last: de Armas

first: Varghese

last: George

first: Abdelali

last: Filali-Mouhim

first: Courtney

last: Steel

first: Anita

last: Parmigiani

first: Coleen

last: Cunningham

first: Adriana

last: Weinberg

first: Lydie

last: Trautmann

first: Rafick-Pierre

last: Sekaly

middle: J

first: Savita

last: Pahwa


title: Gene expression profiling of MDA231 cells depleted of MRCKalpha (CDC42BPA) and MRCKbeta (CDC42BPB)

city: Copenhagen

first: Cord

middle: Herbert

last: Brakebusch

address: Ole Maaløes Vej 5, Biocentret, Building: 1-3-14

country: Denmark

institute: Biotech Research and Innovation Centre

summary:
Aim: The MRCK family of kinases (namely MRCKalpha and MRCKbeta) are downstream effectors of the Rho GTPase signaling pathways with roles in mediating cell migration via regulating the actin cytoskeleon. As such, it is thought that the MRCKs might also be involved in promoting cancer cell invasion or metastasis. However, crucial in vivo data and a general view of the signaling pathways regulated by the MRCKs are still lacking in order to substantiate the hypothesis. Therefore, the goal of this study is to perform transcriptome profiling (RNA-seq) of MDA-MB-231 cells depleted of either CDC42BPA, CDC42BPB, or both, in order to obtain insight into the differential genes expressed and whether CDC42BPA and CDC42BPB contribute as a causative factor of metastatic breast cancer.
Method: We performed CRIPSR-based knockout of MRCKalpha (CDC42BPA), MRCKbeta (CDC42BPB), either alone or in combination, in the invasive breast cancer cell line MDA-MB-231. The combinatory double knockout was performed twice as two biological replicates to increase confidence in our data. The knockout cells were then prepared and sent for RNA-seq using by BGISEQ-500, and differential gene expression profile was obtained.

overall_design: mRNA from vector control, MRCKalpha (CDC42BPA) knockout, MRCKbeta (CDC42BPB) knockout and MRCKalpha/MRCKbeta (CDC42BPA/CDC42BPB) double knockout MDA-MB-231 cells

first: Mei

middle: Q

last: Kwa


title: MED1 is a lipogenesis co-activator required for postnatal adipose expansion

city: Bethesda

first: Kai

last: Ge

state: MD

address: 50 South Dr Rm 4154

department: NIDDK

institute: NIH

summary: MED1 often serves as a surrogate of the general transcription coactivator complex Mediator for identifying active enhancers. MED1 is required for phenotypic conversion of fibroblasts to adipocytes in vitro but its role in adipose development and expansion in vivo has not been reported. Here we show that MED1 is not generally required for transcription during adipogenesis in culture and that MED1 is dispensable for adipose development in mice. Instead, MED1 is required for postnatal adipose expansion and the induction of fatty acid and triglyceride synthesis genes after pups switch diet from high-fat maternal milk to carbohydrate-based chow. During adipogenesis, MED1 is dispensable for induction of lineage-determining transcription factors (TFs) PPARγ and C/EBPα but is required for lipid accumulation in the late phase of differentiation. Mechanistically, MED1 controls the induction of lipogenesis genes by facilitating lipogenic TF ChREBP- and SREBP1a-dependent recruitment of Mediator to active enhancers. Together, our ﬁndings identify a cell- and gene-specific regulatory role of MED1 as a lipogenesis coactivator required for postnatal adipose expansion.

overall_design: Expression profiling by RNA-Seq and nascent RNA-Seq at D7 of adipogenesis in brown or white preadipocytes in culture, RNA-Seq and adipoq+ adipocyte specific TRAP RNA-Seq in interscapular brown adipose tissue and inguinal white adipose tissue and ChIP-Seq profiling of histone H3K27ac, S5P-Pol II, S2P-Pol II, T7 and MED12 by ChIP-Seq and profiling of chromatin accessibility by ATAC-Seq, at D7 of adipogenesis of brown preadipocytes in culture.

first: Younghoon

last: Jang

first: Young-Kwon

last: Park

first: Ji-Eun

last: Lee

first: Danyang

last: Wan


title: The dual function of JmjC domain-containing protein KDM5C in both gene transcriptional activation and repression promotes breast cancer cell growth and tumorigenesis [RNA-seq]

city: Xiamen

first: Guosheng

last: Hu

state: Fujian

address: Xiang'an South Road, Xiang'an District

department: School of Pharmaceutical Sciences

institute: xiamen University

summary: Emerging evidence suggested that epigenetic regulators can exhibit both co-activator and co-repressor activities in gene transcriptional regulation and disease development, such as cancer. However, how these dual activities are regulated and coordinated in cellular contexts remains elusive. Here, we reported that KDM5C, a repressive histone demethylase, is unexpectedly required for estrogen/estrogen receptor alpha (ERa)-induced gene transcriptional activation to promote cell proliferation, while it suppresses the expression of type I interferons (IFNs) and interferon-stimulated genes (ISGs) to escape from immuno-surveillance. KDM5C-interacting protein, ZMYND8, is found to be accompanied with KDM5C in regulation of both subsets of genes. Mechanistically, during estrogen/ERa-induced gene transcriptional activation, ERa interacts and recruits KDM5C/ZMYND8 to active enhancers, where ERa masks KDM5C’s demethylase activity towards H3K4me2/3, converting KDM5C from a co-repressor to a co-activator. Furthermore, KDM5C and ZMYND8 are found to recruit the P-TEFb complex in a cooperative manner to activate estrogen/ERa-target genes. In contrast, KDM5C/ZMYND8 represses type I IFNs and ISGs through directly interfering TBK1 phosphorylation in an enzymatic-dependent manner. The combinatory effects of KDM5C’s dual activities in regulation of genes involved in both cell proliferation and immuno-escape lead to breast cancer cell proliferation in vitro and xenograft growth in mice. Taken together, we revealed a mechanism by which a repressive epigenetic regulator can be converted to a co-activator under specific signal cues to regulate specific gene programs, and the dual nature as both a co-repressor and co-activator together contributes to cancer development.

overall_design: RNA-seq performed in this study was designed to understand the molecular mechanisms underlying KDM5C in gene transcription regulation.

first: Wen

last: Liu

first: Hai-feng

last: Shen

first: Wen-Juan

last: Zhang

first: Guo-sheng


title: The dual function of JmjC domain-containing protein KDM5C in both gene transcriptional activation and repression promotes breast cancer cell growth and tumorigenesis


title: Structural Variation at the Maize wuschel1 Locus Alters Stem Cell Organization in Inflorescences

city: Piscataway

first: Andrea

last: Gallavotti

state: NJ

address: 190 Frelinghysen Rd.

department: Waksman Institute of Microbiology

institute: Rutgers University

summary:
Purpose: The goal of this study is to identify differentially expressed genes (DEGs) in immature ears between Barren inflorescence3 (Bif3) mutant and its wild-type siblings in maize.
Methods: the 3-4 mm ear primordia of maize wild-type and Bif3 mutant were dissected from maize plant at the reproductive stage, and samples were snap freeze using liquid nitrogen. The total RNAs were prepared using RNeasy mini kit (Qiagen). All those RNA samples of wild-type and Bif3 mutant were submitted for RNA-Seq.
Conclusions: Our RNA-seq experiments identified 1380 up- and 242 down-regulated genes (P < 0.05, FDR < 0.1, fold-change > 1.5).

overall_design: RNA-seq was performed on 3-4mm WT and Bif3 mutant maize ear primordia

first: Zongliang

last: Chen


title: Transcriptomic analysis of bone marrow derived macrophages (BMDM) activated by conditioned media (CM) generated from control (CO-Fibs) and hypertensive pulmonary artery adventitial fibroblasts (PH-Fibs)

status: Public on Mar 16 2021

city: Aurora

first: Sushil

last: Kumar

institute: University of Colorado Anschutz medical campus

summary:
Purpose: The goals of this study are to compare transcriptomic profiling (RNA-seq) of BMDMs activated by CM generated from CO-Fibs and PH-Fibs and to evaluate the mechanisms how healthy and pulmonary hypertensive adventitial microenvironment maintain or activate macrophage phenotypes.
Methods: Mouse BMDMs were treated with media conditioned by CO-Fibs (CO-CM) or PH-Fibs (PH-CM) or left untreated (UNX). Each CM was pooled from 5 different fibroblast populations. Total RNA was purified using NucleoSpin RNA Isolation kit (Machery-Nagel, www.mn-net.com) according to the manufacturer’s instructions. RNA quantity and quality were evaluated using NanoDrop and Bioanalyzer. For Transcriptomics analysis, RNA-Seq library preparation and sequencing were conducted using a NuGen universal plus mRNA-Seq kit. Sequencing was performed on Illumina NovaSEQ6000 system, using the paired-read 2x150 cycle option. RNA-Seq reads were generated using Illumina NovaSEQ6000 analysis pipeline. The read quality of all the samples were checked using FastQC v0.11.5. Adapter trimming, quality control, and base correction were performed by AfterQC (Chen et al., 2017). The sequencing reads in the FASTQ files of all the samples were aligned to the Mus musculus reference genome (Mus_musculus. GRCm38v91) with Spliced Transcripts Alignment to a Reference (STAR) version 2.6.0c. We next applied featureCounts v1.6.2 to assign reads to genes using mouse transcript annotations as a guide. EdgeR in R was used for data normalization and differential expression analysis of RNA-Seq expression profiles with biological replication. The Benjamini-Hochberg procedure was used to control the false discovery rate (FDR), and a cut-off criterion of FDR ≤ 0.05 was applied to identify differentially expressed genes. Individual gene expression was calculated as ‘reads per kilobase per million mapped reads’ (RPKM).
Results: Using a standard data analysis workflow (STAR-featureounts-edgeR), our results showed that both CO-CM and PH-CM had significant, yet clearly different, effects on gene expression in BMDMs. Principal component analysis (PCA) of all 12,191 genes showed overall separation of CO-CM and PH-CM treated BMDMs from untreated BMDMs. To obtain insight into the underlying mechanisms through which microenvironments regulate macrophage phenotype and function in control and diseased conditions, we analyzed genes uniquely regulated by CO-CM or PH-CM vs. naïve BMDM using Ingenuity Pathway Analysis (IPA). Canonical signaling pathways identified by IPA demonstrated that CO-CM significantly inhibited inflammatory pathways (role of pattern recognition receptors), immune responses (PD-1, PD-L1 cancer immunotherapy pathway, Tec Kinase signal, and T cell exhaustion signal), lipid (3-phosphoinositide) metabolism and glycolysis suggesting a metabolically quiescent and anti-inflammatory phenotype. In contrast, when the microenvironment was dictated by disease (i.e., PH-CM), macrophages exhibited distinct upregulated canonical pathways, particularly those related to inflammation (iNOS, Toll-like receptor, NF-kB, TNFR, etc.) and activated immune responses (Th1 pathway, T cell exhaustion signaling pathway, dendritic cell maturation). Pathways involved in metabolism were also notably altered with increased NO and ROS production, and decreased oxidative phosphorylation indicating altered mitochondrial function.
Conclusions: Results from our study provide the basis for a more systematic mechanistic understanding of the cross-talk between adventitial fibroblasts and macrophages in health and disease by revealing a comprehensive view of the macrophage transcriptomic and metabolomic landscape.

overall_design: mRNA profiles of mice BMDM treated with control and pulmonary hypertensive adventitial fibroblast culture media

first: Min

last: Li

first: Suzette

last: Riddle

first: Kurt

last: Stenmark


title: A conserved BAH module within mammalian BAHD1 connects H3K27me3 to Polycomb gene silencing

city: Durham

first: Greg

last: Wang

state: NC

address: 3 Genome Court

department: Duke Department of Pharmacology and Cancer Biology  and Duke Cancer Institute

institute: Duke University School of Medicine, and Department of Biochemistry and Biophysics, UNC at Chapel Hill (adjunct)

summary: Trimethylation of histone H3 lysine 27 (H3K27me3) is important for gene silencing, (epi)genome organization and organismal development. In a prevalent model, the functional “readout” of H3K27me3 in mammalian cells is achieved through the H3K27me3-recognizing chromodomain harbored within the chromobox (CBX) component of canonical Polycomb repressive complex 1 (cPRC1), which induces chromatin compaction and gene repression. Here, we report that binding of H3K27me3 by a Bromo Adjacent Homology (BAH) domain harbored within BAH domain-containing protein 1 (BAHD1) is required for overall BAHD1 targeting to chromatin and for optimal repression of the H3K27me3-demarcated genes in mammalian cells. Disruption of direct interaction between BAHD1-BAH and H3K27me3 by point mutagenesis leads to chromatin remodeling, notably, increased histone acetylation, at its Polycomb gene targets. Mice carrying an H3K27me3-interaction-defective mutation of Bahd1BAH causes marked embryonic lethality, showing a requirement of this pathway for normal development. Altogether, this work demonstrates an H3K27me3-initiated signaling cascade that operates through a conserved BAH “reader” module within BAHD1 in mammals.

overall_design: To dissect the role for an H3K27me3-binding BAH module within Bahd1 in gene expression regulation, we carried out transcriptome profiling of the mouse embryonic fibroblast (MEF) cells that carry either WT Bahd1 allele or an engineered W659G homozygous mutation within Bahd1 BAH. W659G disrupts the binding of the Bahd1 BAH motif to H3K27me3.

first: Huitao

last: Fan

first: Yiran

last: Guo


title: The dual function of JmjC domain-containing protein KDM5C in both gene transcriptional activation and repression promotes breast cancer cell growth and tumorigenesis [ChIP-seq]

overall_design: ChIP-seq performed in this study was designed to understand the molecular mechanisms underlying KDM5C in gene transcription regulation.


title: BAHD1 couples H3K27me3 to gene silencing via a conserved BAH reader module and associated corepressor complex in mammals (ChIP-Seq)

summary: Trimethylation of histone H3 lysine 27 (H3K27me3) is crucially involved for gene silencing, (epi)genome organization, cell-fate decision-making and development. To date, functional readout of H3K27me3 is viewed to be achieved mainly through a class of H3K27me3-recognizing chromodomains harbored within the chromobox (CBX) subunit of Polycomb repressive complex 1 (PRC1), which causes chromatin compaction and gene repression partly through histone H2A lysine 119 mono-ubiquitination. We here report that engagement of H3K27me3 by an evolutionarily conserved bromo adjacent homology (BAH) domain harbored within BAH domain-containing protein 1 (BAHD1) significantly contributes to optimal repression of the H3K27me3-demarcated genes in mammalian cells. BAHD1 assembles a NurD-like transcriptional corepressor complex. Abolishing the BAHD1BAH:H3K27me3 interaction by point mutagenesis interferes with BAHD1 binding to chromatin, leading to chromatin remodeling and derepression of Polycomb target genes. Mice carrying an H3K27me3-binding-defective mutation of Bahd1BAH causes marked embryonic lethality, indicating a requirement of this pathway for development. Altogether, this work demonstrates an H3K27me3-initiated signaling cascade that operates through a conserved BAH ‘reader’ module class in mammals.

overall_design: Examination of the genome-wide distribution of H3K27me3, H3K9ac, H3K27ac and H3K9me3 in HEK293 cells with stable expression of an exogenous BAHD1, either wildtype (WT) or BAH-mutated (i.e., W667G), followed by knockdown (KD) of endogenous BAHD1.


title: BAHD1

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

overall_design: Refer to individual Series


title: BAHD1 couples H3K27me3 to gene silencing via a conserved BAH reader module and associated corepressor complex in mammals (RNA-Seq)

overall_design: To dissect the role for an H3K27me3-binding BAH module within BAHD1 in gene expression regulation, we carried out transcriptome profiling of the HEK293 cells with stable expression of an exogenous BAHD1, either wildtype (WT) or BAH-mutated (i.e., W667G), followed by knockdown (KD) of endogenous BAHD1. To determine requirement of H3K27me3 for gene silencing, we carried out transcriptome profiling of the HEK293 cells post-treatment with UNC1999, an EZH2/EZH1-specific small-molecule inhibitor, versus DMSO.


title: Mutations in distinct translation-dependent quality control pathways lead to convergent molecular pathogenesis and neurodevelopmental defects.

status: Public on Mar 15 2021

city: Farmington

first: Scott

last: Adamson

state: CT

address: 10 Discovery Drive

institute: The Jackson Laboratory

summary: Translation-dependent quality control pathways govern protein synthesis and proteostasis by degrading potentially toxic peptide products derived from aberrant mRNAs. However, little is known about the in vivo defects that arise in the absence of these pathways and how translation is altered in the absence of these pathways. Here, we demonstrate that the ribosome rescue factors Pelo/Hbs1l are both critical for embryonic and neuronal development but dispensable for neuronal survival in the adult brain. Loss of Pelo/Hbs1l in cerebellar progenitors impaired differentiation of these cells. Our analysis of Pelo/Hbs1l deficient fibroblasts revealed translational reprogramming of multiple pathways including mTORC1 signaling likely to restore cellular homeostasis. Interestingly, similar affects on the translatome, signaling pathways and neurogenesis were observed when we conditionally deleted Upf2 to inhibit NMD. These data reveal that translation-dependent quality control pathways, which set out to mitigate errors in translation and to clear defective peptide products and aberrant mRNAs can trigger similar cellular responses and neurodevelopmental abnormalities.

overall_design: 41 samples are analyzed in this study which include RNA-seq and Ribosome footprint profiling data from mouse tissues with knockouts of different translation-dependent quality control genes. Profiling datasets include 3 replicates of Hbs1l global knockout from whole mouse embryos with paired wildtype controls. Ribosome profiling datasets also include 3 replicates of Hbs1l conditional knockout in cerebellum with matched wildtype controls. There are also RNA-seq and ribosome footprint profiling for Cre-induced knockouts from mouse embryonic fibroblasts of Pelo, Hbs1l, and Upf2, as well as Cre control samples. There are 3 replicates each with the exception of a Pelo RNA-seq sample which was excluded due to being a biological outlier. The matched Cre controls for profiling datasets are included with knockouts in each batch.

first: Markus

last: Terrey

middle: I

first: Jeffrey

middle: H

last: Chuang

first: Susan

middle: L

last: Ackerman


title: PPARg marks splenic precursors of multiple nonlymphoid-tissue Treg compartments [scRNA-seq multiple tissues]

status: Public on Mar 11 2021

first: CBDM

last: Lab

address: 77 Avenue Louis Pasteur

department: Microbiology and Immunobiology

institute: Harvard Medical School

summary: Foxp3+CD4+ regulatory T cells (Tregs) regulate most types of immune response as well as several processes important for tissue homeostasis – for example, metabolism and repair. Dedicated Treg compartments – with distinct transcriptomes, T-cell-receptor repertoires, and growth/survival factor dependencies – have been identified in several nonlymphoid tissues. These Tregs are specifically adapted to function and operate in their home tissue – when, where and how do they take on their specialized characteristics? We recently reported that a splenic Treg population expressing low levels of the transcription factor, PPARg, contains precursors of Tregs residing in visceral adipose tissue. This finding made sense given that PPARg, the “master-regulator” of adipocyte differentiation, is required for the accumulation and function of Tregs in visceral adipose tissue but not in lymphoid tissues. Here we use single-cell RNA sequencing, single-cell Tcra and Tcrb sequencing, and adoptive-transfer experiments to show that, unexpectedly, the splenic PPARglo Treg population is transcriptionally heterogeneous and engenders Tregs in multiple nonlymphoid tissues beyond visceral adipose tissue, e.g. skin and liver. The existence of a general pool of splenic precursors for nonlymphoid-tissue Tregs opens new possibilities for regulating their emergence experimentally or therapeutically.

overall_design: scRNA-seq of Tregs from VAT, liver, skin and spleen of a 20-week-old Pparg-Tdt.Foxp3-Gfp mouse.

first: Chaoran

first: Andrés

last: Muñoz-Rojas

first: Gang

first: Alexander

middle: O

last: Mann

first: Christophe

last: Benoist

first: Diane

last: Mathis


title: PPARg marks splenic precursors of multiple nonlymphoid-tissue Treg compartments [scTCR-seq]

overall_design: scTCR-seq of Tregs from VAT, liver, skin and spleen of a 20-week-old Pparg-Tdt.Foxp3-Gfp mouse.


title: TCR beta sequencing of blood and tumor samples from EG7.OVA bearing wildtype mice with OT-1 T cell transfer

status: Public on Mar 13 2021

city: Cambridge

first: Anna

last: Salzberg

address: 399 Binney St UNIT 300

institute: Rubius Therapeutics

summary: We report the application of TCRβ sequencing to understand T cell repertoire of matched blood and tumor samples pre and post treatment in EG7.OVA tumor bearing mice with OT-1 cell transfer. TCRβ sequencing demonstrated that OT-1 (CASSRANYEQYF) was the most abundant T-cell clone in both blood and tumors of mRBC‑OVA-4-1BBL-IL-12 treated mice. To investigate the effects of mRBC‑OVA-4-1BBL-IL-12 on immune memory, the T cell repertoire was also analyzed before and after EG7.OVA and EL4 tumor rechallenge in cured and treatment-naive mice. TCRβ sequencing on T cells in peripheral blood showed that OT-1 clones increased in frequency after EG7.OVA challenge in previously cured mice. OT-1 frequency did not increase in treatment-naïve mice after tumor challenge, indicating that the tumor alone is insufficient to drive OT-1 cell expansion. We also evaluated the frequencies of unique TCRβ sequences in T cell clones that significantly expanded post EL4 challenge (EL4 responsive TCR). Increased frequency of EL4-responsive TCRs upon each tumor challenge (EG7.OVA and EL4) was associated with complete responders (mice that rejected EL4 challenge), suggesting that T-cell-mediated protection against parental tumor antigens was generated prior to EL4 challenge. Partial responders (delayed tumor growth compared with naïve mice) had increases in EL4-responsive TCR frequencies after EL4 challenge but not during the EG7.OVA rechallenge, whereas the non-responder (tumor growth similar to naïve mice) had minimal increases in TCR frequencies upon EL4 challenge. Overall, the ability to control EL4 tumors correlated with the expansion of EL4-responsive TCR clones.

overall_design:
TCR repertoire analyses of matched blood and tumor samples, pre and post mRBC-OVA-4-1BBL-IL-12 treatment as well as pre and post EG7.OVA and EL4 challenge in cured mice in a murine lymphoma model with OT-1 T cell transfer
-------------------------------------------------------
Submitter claims they are unable to provide raw fastq files:
The authors state "Raw data were processed using the standard immunoSEQ Analyzer from Adaptive Biotechnologies Corporation (third party CRO), and this third party does not release raw data."

first: Xuqing

last: Zhang

first: Tiffany

middle: F

last: Chen

first: Mellissa

last: Nixon


title: Mutations in GEMIN5 cause a neurodevelopmental ataxia syndrome by a loss-of-1function mechanism

city: Pittsburgh

first: Bernhard

last: Kuhn

state: Pennsylvania

address: 4401 Penn Ave

institute: Children's Hospital of Pittsburgh

summary: GEMIN5 is a critical component of snRNP assembly complex. Patients carrying novel autosomal recessive variants in the GEMIN5 gene showed symptoms of developmental delay, central hypotonia, and cerebellar ataxia which are distinct than classical spinal muscular atrophy. We performed RNA-seq analysis in iPSC-derived differentiated neurons with biallelic GEMIN5-H913R mutation to identify global alterations in various genes and pathways mediated by GEMIN5 mutations in patients.

overall_design:
Examination of mRNA profile of iPSC-derived differentiated neurons carrying biallelic H913R varaint of GEMIN5 with isogenic WT controls

*** Raw data not submitted to GEO due to patient privacy concerns ***

first: Sukhleen

last: Kour

first: Udai

middle: B

last: Pandey


title: UBE3A Deficiency Downregulates Retinoic Acid Signalling Pathway

city: shanghai

first: ronggui

last: hu

state: China

address: Shanghai Institute of Biochemistry and Cell Biology

institute: Center for Excellence in Molecular Cell Science

summary: Ubiquitin-protein ligase E3A (UBE3A) has dual functions as a E3 ubiquitin-protein ligase and coactivator of nuclear hormone receptors. Mutations or deletions of the maternally inherited UBE3A gene cause Angelman syndrome. Here, we performed transcriptome profiling in the hippocampus of Ube3am+/p+ and Ube3am-/p+ mice, and determined that the expression of the retinoic acid (RA) signalling pathway was downregulated in Ube3a-deficient mice compared to WT mice. Furthermore, we demonstrated that UBE3A directly interacts with RARα and may function as a coactivator of the nuclear receptor RARα to participate in the regulation of gene expression. Loss of UBE3A expression caused the downregulation of the expression of RA-related genes, including Erbb4, Dpysl3, Calb1, Pten and Arhgap5 in Ube3am-/p+ mice brain tissues. This work revealed a new role for UBE3A in regulating retinoic acid (RA) signalling downstream genes and hopefully to shed light on the potential drug target of AS.

overall_design: mRNA sequencing data from hippocampus of three Ube3am+/p+ male mice  and three Ube3am-/p+ male mice

first: Meimiao

last: Fang

first: Yali

last: Li

first: Jin

last: Ren

first: Xiaobo

last: Gao

first: Ronggui

last: Hui


title: BAHD1 couples H3K27me3 to gene silencing via a conserved BAH reader module and associated corepressor complex in mammals (CUT&RUN)

first: Greg

last: Wang

overall_design: Examination of the genome-wide distribution of BAHD1 in HEK293 cells with stable expression of an exogenous Flag-tagged BAHD1, either wildtype (WT) or BAH-mutated (i.e., W667G or Y669A), followed by knockdown (KD) of endogenous BAHD1. CUT&RUN with Flag antibodies were carried out in these cells, with cells with no expression of Flag-BAHD1 used as a negative control.

first: Huitao

last: Fan

first: Yiran

last: Guo


title: RAB protein controls Argonaute localization for proper miRNA function in C. elegans

city: Riverside

first: Weifeng

last: Gu

state: California

address: 900 University Ave

department: Molecular, Cell and Systems Biology

institute: University of California at Riverside

summary: MicroRNAs are critical regulators of gene expression in animals. To repress their target mRNAs, these small RNAs are first loaded onto Argonaute proteins to form the silencing complex called miRISC. The complex must then be transported to its target mRNA where it interacts mostly within the 3’UTR through base-pairing. After target repression, the constituents of the miRISC undergo degradation or recycling mediated through their accurate sorting and localization in the cell. While some reports have linked intracellular trafficking to miRNA activity, it is still unclear how these pathways coordinate to allow proper miRNA-mediated gene silencing and turnover. Through a forward genetic screen performed in the nematode Caenorhabditis elegans, we identified the RabGAP tbc-11 as an important factor for the miRNA pathway. We show that TBC-11 is likely a GAP for the small GTPase RAB-6 and that its regulation is required for miRNA function in animals. The absence of functional TBC-11 leads to the persistent activation of RAB-6, which increases the localization of miRNA-unloaded Argonaute ALG-1 on endomembranes. This inactive form of Argonaute accumulates on polysomes, leading to defective miRNA-mediated target mRNA repression. Together, our results establish the importance of intracellular trafficking for miRNA function and demonstrates the involvement of a small GTPase in proper Argonaute localization in vivo.

overall_design: Develop a novel protocol to clone small RNA and mRNA

first: Pascale

last: Michaud

first: Rachel

last: Rioux

first: Camille

last: Lavoie-Ouellet

first: Pauline

last: Adjabade

first: Rachid

last: Mazroui

first: Martin

last: Simard


title: PPARg marks splenic precursors of multiple nonlymphoid-tissue Treg compartments

first: CBDM

last: Lab


title: PPARg marks splenic precursors of multiple nonlymphoid-tissue Treg compartments [scRNA-seq spleen]

overall_design: scRNA-seq of Tregs from spleens of two or three 8-week-old Pparg-Tdt.Foxp3-Gfp mice.

first: Chaoran

first: Andrés

last: Muñoz-Rojas

first: Gang

first: Alexander

last: Mann

first: Christophe

last: Benoist

first: Diane

last: Mathis


title: DNA methylation and expression profiles of whole blood in Parkinson’s disease (RNA-Seq)

city: TEMPE

first: QI

last: WANG

state: AZ

institute: ARIZONA STATE UNIVERSITY-TEMPE CAMPUS

summary: Parkinson’s disease (PD) is the second most common age-related neurodegenerative disease. It is presently only accurately diagnosed at an advanced stage by a series of motor deficits, which are predated by a litany of non-motor symptoms manifesting over years or decades. Aberrant epigenetic modifications exist across a range of diseases and are non-invasively detectable in blood as potential markers of disease. We performed comparative analyses of the methylome and transcriptome in blood from PD patients and matched controls. Our aim was to characterize DNA methylation and gene expression patterns in whole blood from PD patients as a foundational step toward the future goal of identifying molecular markers that could predict, accurately diagnose, or track progression of PD. We found that differentially expressed genes (DEGs) were involved in the processes of transcription and mitochondrial function and that PD methylation profiles were readily distinguishable from healthy controls, even in whole blood DNA samples. Differentially methylated regions (DMRs) were functionally varied, including near transcription factor NFYA, receptor tyrosine kinase DDR1, ubiquitin ligase RNF5, acetyltransferase AGPAT1, and vault RNA VTRNA2-1. Expression quantitative trait methylation (eQTM) sites were found at long non-coding RNA PAX8-AS1 and transcription regulator ZFP57, among others. Functional epigenetic modules (FEM) were highlighted by IL18R1, PTPRC, and ITGB2. We identified patterns of altered disease-specific DNA methylation and associated gene expression in whole blood. Our combined analyses extended what we learned from the DEG or DMR results alone. These studies provide a foundation to support characterization of larger sample cohorts, with the goal of building a thorough, accurate and non-invasive molecular PD biomarker.

overall_design: Parkinson's disease (PD) and control subjects where analyzed for changes in gene expression and DNA methylation. Each group contained 15 age-matched subjects. Two blood samples were taken from each subject. One sample was used for isolation of total RNA and the other was used for extraction of DNA. The PD group was compared to controls for each data type and correlations between data types were assessed.

first: Adrienne

last: Henderson

first: Qi

first: Bessie

last: Meechoovet

first: Ashley

last: Siniard

first: Marcus

last: Naymik

first: Matthew

last: DeBoth

last: Huentelman

first: Richard

last: Caselli

first: Erika

last: Driver-Dunckley

first: Travis

last: Dunckley


title: Rac1b modulates intestinal tumourigenesis via modulation of Wnt and Egfr signalling pathways

city: Edinburgh

first: Kevin

last: Myant

address: Crewe Road

country: United Kingdom

institute: MRC Institute of Genetics and Molecular Medicine

summary: mRNAseq of Apc mutant and Apc Rac1b double mutant mouse intestinal tumours.

overall_design: RNA was extracted from tumours from Apc mutant and Apc Rac1b double mutant mouse intestines. mRNAseq was carried out to identify differentially expressed transcripts.

first: Victoria

last: Gudiño

first: Sebastian

last: Pohl

first: Caroline

middle: V

last: Billard

first: Patrizia

last: Cammareri

first: Alfonso

last: Bolado

first: Stuart

last: Aitken

first: David

last: Stevenson

first: Adam

last: Hall

first: Mark

last: Agostino

first: John

last: Cassidy

first: Colin

last: Nixon

first: Alex

last: von Kriegsheim

first: Paz

last: Friele

first: Linda

last: Popplewell

first: George

last: Dickson

first: Laura

last: Murphy

first: Ann

last: Wheeler

first: Malcolm

last: Dunlop

first: Farhat

last: Din

first: Douglas

last: Strathdee

first: Owen

last: Sansom


title: DNA methylation and expression profiles of whole blood in Parkinson’s disease

first: QI

last: WANG


title: Cardiac DNA Methylation Underlies Racial and Socioeconomic Disparities among Patients with End-Stage Heart Failure (RNA-seq)

city: Stanford

middle: Emile

last: Pepin

state: CA

address: 870 Quarry Road

department: Cardiovascular Medicine

institute: Stanford University

summary: African Americans (AA) are 70% more likely than Caucasian Americans (CA) to die from heart failure (HF) even after adjusting for known causes. Although the causal factors responsible for this racial disparity remain unknown, it is theorized that environmental stressors This alarming health disparity represents an important challenge to U.S. healthcare as global prevalence of heart failure has already exceeded epidemic levels with a disease burden that disproportionately impacts members of ethnic and racial minorities. The current multicohort study of cardiac DNA methylation identifies the cardiac epigenome as a previously unrecognized syntax that encodes race-based environmental differences in the failing human heart.

overall_design: The current study began as a pilot analysis of myocardial genome-wide DNA methylation via Illumina® HumanMethylation450 array with tandem RNA-sequencing analysis of 11 subjects with NYHA Stage IV heart failure. Unsupervised multidimensional scaling (MDS) of genome-wide cardiac DNA methylation identified patient race as the factor which accounted for the highest degree of methylation variability across the samples. The effects of race could not be explained by any other patient factors present in the electronic health record. This observation prompted a confirmatory analysis of 31 human HF biopsies using the Illumina Beadship MethylationEPIC array, wherein race-based separation was again seen as the principal determinant of patient heterogeneity via unsupervised deconvolution.

first: Chae-Myeong

last: Ha

last: Wende


title: Evolution of AML genome and epigenome with IDH inhibitors and their association with clinical response and resistance

city: Houston

first: FENG

state: TX

address: 1881 East Road

institute: UT MD Anderson Cancer Center


title: DNA methylation and expression profiles of whole blood in Parkinson’s disease (Methylation)

first: Adrienne

last: Henderson

first: Qi

last: Wang

first: Bessie

last: Meechoovet

first: Ashley

last: Siniard

first: Marcus

last: Naymik

first: Matthew

last: DeBoth

last: Huentelman

first: Richard

last: Caselli

first: Erika

last: Driver-Dunckley

first: Travis

last: Dunckley


title: Evolution of AML genome and epigenome with IDH inhibitors and their association with clinical response and resistance [methylation]

summary: Allosteric inhibitors of mutant IDH1 or IDH2 induce terminal differentiation in mutant leukemic blasts and may provide durable clinical responses in approximately 40% of acute myeloid leukemia (AML) patients with the mutations. However, most responders eventually relapse. To understand the molecular underpinnings of clinical resistance, we performed multipronged genomic analyses (DNA sequencing, RNA sequencing and cytosine methylation profiling) in longitudinally collected specimens from 68 IDH1/IDH2-mutant AML patients treated with IDH inhibitors (IDHi), and described the evolution of AML genome and epigenome during the therapy and its association with clinical response and relapse. Co-occurrence of mutations in RUNX1/CEBPA or RAS-RTK pathway genes were associated with poor response to IDHi. The same group of mutations were also frequently selected or acquired at relapse. In addition, acquired mutations in BCOR, reciprocal IDH gene, and TET2 were also implicated at relapse. DNA methylation changes largely mirrored plasma 2HG dynamics and had little association with clinical response to IDHi. The mapping of mutation dynamics and methylation changes in longitudinal samples revealed the interplay between AML genome and epigenome with IDHi therapy. While the alteration in RAS-RTK signaling genes and RUNX1/CEBPA were the key molecular pathways involved in clinical resistance to IDHi, diverse molecular pathways were involved in IDHi resistance. The data highlights the role of clonal heterogeneity in therapeutic resistance in AML and implicates opportunities for novel combination strategies.

overall_design: DNA methylation analysis was performed using Illumina’s Infinium MethylationEPIC assay (EPIC) on longitudinal samples during the IDH inhibitor therapy.

first: Feng

first: Koichi

last: Takahashi


title: Evolution of AML genome and epigenome with IDH inhibitors and their association with clinical response and resistance [RNA-seq]

overall_design:
RNA expression profiles were generated by RNA-Seq using HiSeq 2000 platform on longitudinal samples during the IDH inhibitor therapy.
BL = baseline
REL = relapse


title: Identification of the target genes of androgen in skeletal muscle using myofiber specific ARKO female mice

status: Public on Mar 10 2021

city: Toon

first: Yuuki

last: Imai

state: Ehime

address: Shitsukawa

department: Proteo-Science Center

institute: Ehime University

summary: Androgens have a strong effect against skeletal muscles to increase muscle mass and strength. However, a molecular mechanism of AR action on muscle strength is not clear. To identify the target genes of AR in skeletal muscle, we generated myofiber specific ARKO using HSA-Cre and AR flox mice (cARKO). Nine-week-old female control and cARKO mice were treated with or without DHT for 4 weeks. After euthenization, gastrocunemius muscle were collected and total RNA were extracted.

overall_design: mRNA profiles of gastrocunemius muscle obtained from control and cARKO mice treated with or without DHT for 4 weeks were generated by sequencing, in triplicates, using Illumina Myseq.

first: Iori

last: Sakakibara


title: Racial and Socioeconomic Disparity Associates with Differences in Cardiac DNA Methylation among Men with End-Stage Heart Failure

first: Mark

type: Methylation profiling by genome tiling array

summary: Heart Failure (HF) is a multifactorial syndrome and that remains a leading cause of worldwide morbidity. Despite its high prevalence, only half 50% of HF patients respond to guideline-directed medical management, prompting therapeutic efforts . Therapeutic avenues are forced to confront the molecular underpinnings of its heterogeneity of HF as a clinical syndrome to identify novel molecular targets. In both pilot (n = 11) and testing (n = 31) cohorts (n = 31), unsupervised multidimensional scaling of genome-wide myocardial DNA methylation exhibited a bimodal distribution of CpG methylation found largely to occur in the promoter regions of metabolic genes.. Among the available patient attributes, only categorical patient race could delineate this methylation signature, with African American (AA) and Caucasian American (CA) samples clustering separately. Because race is a social construct, and thus a poor proxy of human physiology, extensive review of medical records was conducted, but ultimately failed to identify co-variates of race at the time of LVAD surgery. By contrast, retrospective analysis exposed a higher all-cause mortality among AA (56.3%) relative to CA (16.7%) patients at 2 years following LVAD placement (P = 0.03). .G To mitigate concerns of genetic confounding factors, putative single-nucleotide polymorphisms (SNPs) were identified by proximity to CpG methylation sites. Nevertheless, a minor proportion of SNPs (0.15%) was identified, and no evidence of racial dissimilarity was seen among them. By contrast, geocoding-based approximation of patient demographics uncovered disparities in income levels among AA relative to CA patients. Therefore, Although although additional studies studies are warrantedneeded, our multicohort studythe current analysis implicates  identifies cardiac DNA methylation as as a previously unrecognized indicator of socioeconomic disparity in human heart failure outcomes.

overall_design: In the current study, we examined epigenetics as a yet unexplored mechanism source of heterogeneity in among patients with end-stage HF. Specifically, a multicohort-based study was designed to analyze quantify cardiac genome-wide cytosine-p-guanine (DNA CpG) methylation of cardiac biopsies from patients undergoing left ventricular assist device (LVAD) implantation.


title: Nicotamide riboside attenuates age-associated transcriptional changes in hemoatpoietics stem cells

status: Public on Mar 09 2021

city: Brisbane

first: Christian

last: Nefzger

state: Queensland

address: 306 Carmody Road

department: Institute for Molecular Biology

country: Australia

institute: The University of Queensland

overall_design: Transcriptional profile (RNA-seq) of hematopoietic stem cells  (HSCs) from young (6-12 weeks), aged(20-24 moths) and aged mice treated with Nicotamide Riboside (NR, a form of vitamin B3). Female C57BL/6 mice  were used for experiments. Animals received drinking water with or without NR (3mg/ml) ad libitum for 2 months.

first: Xuan

last: Sun

first: Benjamin

last: Cao

first: Marina

last: Naval-Sanchez

first: Tony

last: Pham

first: Yu

first: Brenda

last: Williams

first: Shen

middle: Y

last: Heazlewood

first: Nikita

last: Deshpande

first: Y

last: Li

first: Felix

last: Kraus

first: James

last: Rae

first: Quan

last: Nguyen

first: Hamed

last: Yari

first: Jan

last: Schröder

first: Chadn

last: Heazkewood

first: Madeline

last: Fulton

first: Jessica

last: Hatwell-Humble

first: Kaustav

middle: D

last: Gupta

first: Ronan

last: Kapetanovic

first: Xiaoli

last: Chen

last: Sweet

first: Robert

middle: G

last: Parton

first: Michael

last: Ryan

first: Jose

middle: M

last: Polo

first: Susan

last: Nilsson


title: RNA-seq analysis of A549 cells harboring Smad3 R104K A105P mutation.

city: Tokyo

first: Daizo

last: Koinuma

address: Hongo 7-3-1, Bunkyo-ku

department: Pathology

institute: University of Tokyo

summary: Transforming growth factor-beta (TGF-beta) transmits signals that facilitate cancer progression. Especially, epithelial-mesenchymal transition (EMT) induced by TGF-beta is considered to crucially contribute to the malignant phenotype of cancer cells. Here we report that the EMT-associated cellular responses induced by TGF-beta are mediated through distinct signaling pathways that diverge at Smad3; cell motility and epithelial marker downregulation are Smad3-dependent while mesenchymal marker induction is not. Furthermore, using a chimeric protein approach in SMAD3 knockout A549 cells, we found that the beta 4 region in the MH1 domain of Smad3 is indispensable for TGF-beta–induced cell motility, but not for epithelial marker downregulation. A transcriptome analysis was performed using A549 cells expressing Smad3 mutant of the MH1 domain.

overall_design: RNA-seq analyses were performed in wild type- and mutant Smad3 (R104K A105P)-expressing A549 lung adenocarcinoma cells stimulated with TGF-beta.

first: Mitsuyoshi

last: Motizuki

first: Takashi

last: Yokoyama

first: Yuka

last: Itoh

first: Chiho

last: Omata

first: Kohei

last: Miyazono

first: Masao

last: Saitoh

first: Keiji

last: Miyazawa


title: Population and single cell RNAseq analysis of CD4+ T cells in FOXP3 mutant mice ( scurfy") and IPEX patients

status: Public on Mar 08 2021

city: Boston

first: CBDM

last: Lab

overall_design: Refer to individual Series


title: Profiling of cultured primary stromal cells from human lung

city: South San Francisco

first: Jason

middle: Anthony

last: Vander Heiden

address: 1 DNA Way

department: Biological Research AI Development

institute: Genentech

summary: Recent publications have implicated senescent alveolar epithelial cells as the cause of lung remodeling and fibrosis. We sought to better understand the heterogeneity in the basal epithelial cell population, especially with regards to characterizing senescent alveolar epithelial cells. The basal cell cultures contain a portion of cells that display a senescent phenotype and stain for a marker of senescence. We also sought to understanding the heterogeneity in the fibroblast population. Primary fibroblast cultures stained with myofibroblast markers show that only a portion of the cells are positive for ACTA2, suggesting that there are at least two distinct populations in this particular cell type. The diversity in this cell type remains uncharacterized at this point.

overall_design: Primary lung basal epithelial cells (EPCAM+) and fibroblasts (CD90+) were isolated from a human lung explant and cultured in appropriate media to enhance expansion. Subsequent passaging enriched cultures for either epithelial or fibroblast cell types. Cells at passage 4 were processed for single-cell RNA-seq to investigate replicative senescence in epithelial cells and fibroblast subtypes in culture.

first: Daryle

last: DePianto

first: Katrina

last: Morshead

middle: A

first: Joseph

last: Arron


title: Characterizing senescent lung epithelial cells

summary: We characterized senescent alveolar epithelial cells in an effort to better understand how these cells contribute to lung remodeling and fibrosis. The transcriptional profiles of several different types of senescent lung epithelial cells was evaluated using various induction methods in order to better understand core characteristics associated with the senescent program in epithelial cells of the lung.

overall_design: Three separate cell types have been subjected to hydrogen peroxide or doxorubicin treatment in order to induce senescence, as evidenced by positive staining for senescent-associated beta-galactosidase activity. This gave us the opportunity to look at senescence in different cell types and with differing methods of induction.

middle: J

middle: R


title: Examining gene expression profile from varying preparations of fibrotic lung tissue

summary: Gene expression of human lung tissue bronchoalveolar lavage (BAL), whole tissue biopsy, and digested single cell suspension of lung tissue from control and ILD samples.

overall_design: Gene expression profiles were captured from various samples derived from human lung tissue explants. Bronchoalveolar lavage, whole tissue biopsy, and a single cell suspension derived from digested lung tissue were prepared and evaluated via RNA-seq. This allowed for evaluation of sample preparation, and retention of singal in digested samples, to ensure that there was not significant loss of signal due to incomplete digestion or bias of preparation protocol.

first: Paul

last: Wolters


title: Arginine effect on prostate cancer cell lines

status: Public on Mar 07 2021

city: Miaoli County

first: Chia-Lin

last: Chen

address: No.35 Keyan Rd, Zhunan

department: Molecular and Genomic Medicine

country: Taiwan

institute: National Health Research Institutes

summary: In this study, we performed the microarray analysis in two arginine auxotrophic prostate cancer cell lines, 22Rv1 and PC3, to detail the global gene expression with or without arginine condition.

overall_design: Two arginine-sensitive prostate cancer cell lines, 22Rv1 (highly-sensitive) and PC3 (sensitive), were treated with or without arginine and evaluated the global gene expression by Affymetrix (Clariom™ S Assay, human) microarray analysis.

first: Chia

first: Sheng

middle: C

last: Hsu

first: Hsing

last: Kung


title: Arginine acts as an epigenetic modulator to regulate mitochondrial function

summary: In this study we showed that arginine globaly induces metabolic genes expression via histone acetylation in prostate cancer cells

overall_design: Examination of the alteration of total histone H3 acetylation upon arginine stimulation.

first: Hsing-Jien


title: Molecular mapping of interstitial lung disease reveals a phenotypically distinct senescent basal epithelial cell population

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


title: Insights into inherited thrombocytopenia resulting from mono-allelic germline mutations in ETV6 or RUNX1 using a human pluripotent stem cell model

status: Public on Mar 06 2021

city: Philadelphia

last: Gadue

state: PA

address: 3501 Civic Center Blvd

department: Hematology

institute: University of Pennsylvania

summary: Inherited thrombocytopenia results in low platelet counts and increased bleeding. Subsets of these patients have monoallelic germline mutations in ETV6 or RUNX1 and a heightened risk of developing hematologic malignancies. Utilizing CRISPR/Cas9, we compared the in vitro phenotype of hematopoietic progenitor cells and megakaryocytes derived from induced pluripotent stem cell (iPSC) lines harboring mutations in either ETV6 or RUNX1. Both mutant lines display phenotypes consistent with a platelet-bleeding disorder. Surprisingly, these cellular phenotypes were largely distinct. The ETV6-mutant iPSCs yield more hematopoietic progenitor cells and megakaryocytes, but the megakaryocytes are immature and less responsive to agonist stimulation. On the contrary, RUNX1-mutant iPSCs yield fewer hematopoietic progenitor cells and megakaryocytes, but the megakaryocytes are more responsive to agonist stimulation. However, both mutant iPSC lines display defects in proplatelet formation. Our work highlights that while patients harboring germline ETV6 or RUNX1 mutations have similar clinical phenotypes, the molecular mechanisms may be distinct.

overall_design: Three biological replicates were analyzed for each of the four genotypes 6 conditions (four genotypes; three biological replicates)

first: Sara

last: Borst


title: Transcriptional analysis of cystic fibrosis airways at single-cell resolution reveals altered epithelial cell states and composition

city: Los Angeles

first: Brigitte

last: Gomperts

address: 10833 Le Conte Ave, A2-410 MDCC

institute: University of California, Los Angeles

summary: Single cell sequencing of human airway epithelium from normal and cystic fibrosis lungs.

overall_design: Single cell RNA-seq using dropSeq (UCLA) or 10x chromium system (CFF, CSMC), on freshly isolated (UCLA, CFF), FACS isolated (DAPI-,CD45-,CD31-,CD326+) (CSMC), or freshly isolated, expanded, and cultured on ALI for 28 days (CFF) epithelial cells  from control and CF lung explant tissue. The barcoded sequencing libraries were quantified by qPCR using the KAPA Library Quantification Kit (KAPA Biosystems). Sequencing libraries were loaded on a NovaSeq 6000 (Illumina) (CSMC), NextSeq 500 (Illumina) (CFF) or HiSeq4000 (Illumina) (UCLA).

first: Barry

last: Stripp

first: Gianni

last: Carraro

first: John

last: Mahoney


title: Compare transcriptomes of without or with BMP4 treatment in 2H11 cells

first: GUOYU

last: YU

address: 1515 Holcombe Blvd, Houston,

department: Translational Molecular Pathology

institute: MD Anderson Cancer Center

summary: In order to observe BMP4-induced transcriptome, 2H11 cells treated without or with BMP4 for 48 hours were subjected to RNAseq analyses

overall_design: mRNA profiles of without or with BMP4 treatment for 48 hours in 2H11 cells

first: Guoyu

last: Yu

first: Sue-Hwa

last: Lin

first: Yanggu

last: Shi


title: Cardiac DNA Methylation Underlies Racial and Socioeconomic Disparities among Patients with End-Stage Heart Failure (MethylationEPIC array)

first: Mark

last: Pepin

middle: E

first: Chae-Myeong

last: Ha

first: Adam

last: Wende


title: Yeast cell fate control by temporal redundancy modulation of transcription factor paralogs

city: Beijing

first: Yan

last: WU

state: Beijing

address: No.5 Yiheyuan Road Haidian District, Beijing, P.R.China

department: Academy for Advanced Interdisciplinary Studies

institute: Peking University

summary: Recent single-cell studies have revealed that yeast stress response involves multiple transcription factors that are temporally activated in pulses. However, it remains largely unclear whether and how these dynamic transcription factors temporally interact to regulate stress survival. Here we show that budding yeast cells can exploit the temporal relationship between paralogous general stress regulators, Msn2 and Msn4, during stress response. We found that individual pulses of Msn2 and Msn4 are largely redundant, and cells can enhance the expression of their shared target genes by increasing their temporal divergence. Thus, functional redundancy between these two paralogs is modulated in a dynamic manner to confer fitness advantages for yeast cells, which might feed back to promote the preservation of their functional redundancy. This evolutionary implication was supported by evidence from Msn2/Msn4 orthologs and analyses of other transcription factor paralogs. Together, we show a cell fate control mechanism through temporal redundancy modulation in yeast, which may represent an evolutionarily important strategy for maintaining functional redundancy between gene duplicates.

sample_organism:
Saccharomyces cerevisiae
Nakaseomyces glabratus

overall_design: First, for transient stimulation experiments, 7 genotypes (namely, 3 genotypes of S. cerevisiae: wild type sc, Msn2_sc deletion, Msn4_sc deletion and 4 genotypes of C. glabrata: wild type cg, Msn2_cg deletion, Msn4_cg deletion, Msn2_cg deletion Msn4_cg deletion) under two conditions (namely, stressed and control) were sequenced with 2 biological repeats. Second, for steady-state experiments, wild type sc strain was sequenced under 4 different carbon stresses with 2 biological repeats.

last: Wu

first: Jiaqi

first: Minghua

last: Deng

first: Yihan


title: RNA-seq for primary human bronchial epithelial cells, proliferating versus doxorubicin-induced senescence.

first: Jason

last: Vander Heiden

summary: Epithelial senescence, specifically in the bronchiolized regions of the fibrotic lung, is apparent in IPF but not control tissue. In order to better understand the phenotype of these senescent cells we generated a in vitro culture model of senescence utillizing primary normal human bronchial epithelial cells. Cultures were treated with doxorubicin to induce senescence and the transcriptional phenotype of the senescent cultures was compared to DMSO-treated control cultures.

overall_design: Primary human bronchial epithelial cells in culture were treated with DMSO, as a control, or doxorubicin (500nM) for a total of 5 days. RNA was then harvested and RNA-seq was performed to reveal the transcriptional profile enacted in senescent cells.

first: Daryle

last: DePianto

first: Katrina

last: Morshead

first: Joseph

last: Arron

first: Grace

last: Teng


title: Expression data for  cCD1 cell-sorted from the lungs of healthy or KP tumor-bearing mice

city: Padova

first: Silvio

last: Bicciato

address: via U. Bassi 59/b

department: Molecular Medicine

institute: University of Padova

summary: The goal of this study was to identify mechanism of suppression of cDC1 in a model of genetically driven lung adenocarcinoma (KP). As part of the study we analyzed the transcriptional differences between cDC1 cell-sorted from healthy lung and those cell-sorted from KP bearing lungs.

overall_design: C57/BL6 mice were injected with the KP tumor cell line and cDC1 were isolated by cell sorting from lung tissues. cDC1 from healthy lung were used as controls. Each experimental replica includes cDC1 isolated from a pool of lung tissues from 2-3 mice/condition.

first: Federica

last: Benvenuti

first: Nicoletta

last: Caronni


title: Next Generation Sequencing Facilitates Quantitative Analysis of healthy indiviual and HUWE1-promoted XLID  Transcriptomes

city: Trondheim

first: Sarah

middle: Louise Fordyce

last: Martin

address: Erling Skjalgssons gt 1

department: Department of Clinical and Molecular Medicine (IKOM)

country: Norway

institute: Norwegian University of Science and Technology

overall_design:
mRNA profiles of healthy vs HUWE1 mutant cells were generated by deep sequencing, in triplicate, using Illumina HiSeq 4000 (single read, 75bp).
Note from submitter: We will not be submitting fastq or any sequence files due to Norwegian law for human data of these individuals.

first: Rossana

last: Aprigliano

first: Barbara

last: vanLoon


title: p38α plays differential roles in hematopoietic stem cell activity dependent on aging contexts II

status: Public on Mar 03 2021

first: Noriko

last: Sorimachi

address: 2-11-16 Yayoi, Bunkyo-ku

department: Department of Biophysics and Biochemistry

institute: Graduate School of Science, The University of Tokyo

summary: Hematopoietic stem cells (HSCs) and their progeny sustain lifetime hematopoiesis. Aging alters HSC function, number, and composition and increases risk of hematological malignancies, but how these changes occur in HSCs remains unclear. Signaling via p38MAPK has been proposed as a candidate mechanism underlying induction of HSC aging. Here, using genetic models of both chronological and premature aging, we describe a multimodal role for p38α, the major p38MAPK isozyme in hematopoiesis, in HSC aging. We report that p38α regulates differentiation bias and sustains transplantation capacity of HSCs in the early phase of chronological aging (from young to 1-year-old). However, p38α decreased HSC transplantation capacity in the late progression phase of chronological aging (from 1- to 2-years-old). Furthermore, co-deletion of p38α in mice deficient in Ataxia-telangiectasia mutated (Atm), a model of premature aging, exacerbated aging-related HSC phenotypes seen in Atm single mutant mice. Mechanistically, p38α makes a positive contribution to inflammation during the late phase aging, resulting in defects in 2-year-old HSCs. Overall, we propose multiple functions of p38MAPK, which both promotes and suppresses HSC aging context-dependently.

overall_design: We injected CAG-CreERT2::Atmflox/flox (Atmfl/fl) and CAG-CreERT2::Atmflox/floxp38αflox/flox (Atmfl/flp38αfl/fl) mice aged approximately 5 weeks with TAM, and then analyzed those mice and corresponding controls 4 weeks later (WT, AtmsKO, Atmp38dKO). lineage-Sca-1+c-Kit+CD34-Flt3- cells (LT-HSC) in the bone marrow (femur and tibia) from WT, AtmsKO, Atmp38dKO C57BL/6J mice were sorted into SF-O3 medium. Then, sorted cells were centrifuged at 340 x g for 5 min at 4°C and lysed with 75µL RLT buffer + 0.75µL 2-ME. RNA extraction, cDNA synthesis, microarray analysis, and data normalization were outsourced to DNA Chip Research Inc.

first: Yuriko

first: Daiki

last: Karigane

first: Yukako

last: Ootomo

first: Hiroshi

last: Kobayashi

first: Keiyo

last: Takubo


title: Modified Hi-C reveals distinct chromatin architecture in endosperm and leaf of Arabidopsis

status: Public on Mar 04 2021

city: Uppsala

first: Juan

last: Santos-González

state: Uppsala

address: Almas Allé 5

department: Department of Plant Biology and Forest Genetics

country: Sweden

institute: SLU

summary: Higher-order chromatin structure undergoes dramatic changes in response to various developmental and environmental signals, wherein distinct cell types posses specific chromatin organization. High throughput chromatin conformation capture assays (Hi-C) allows study of higher-order chromatin structure; however, it requires a large number of cells. This requirement has so far limited the establishment of cell type-specific higher-order chromatin structure in the plant. To overcome this limitation, we modified the Hi-C protocol (mHi-C) to be applicable to a limited amount of starting material. For this, mHi-C libraries were generated from INTACT-purified endosperm and leaf nuclei. Correlation plots showed that our mHi-C data from INTACT-leaf accurately reiterate chromatin interaction patterns derived from conventional leaf Hi-C data. We further identified compacted structural domains (CSDs) and loose structural domains (LSDs) in leaf and endosperm and their differential patterns revealed distinct higher-order chromatin organization in leaf and endosperm. Our analysis revealed that DNA methylation and repressive histone marks positively correlate with the chromatin compaction level. We discovered increased chromatin interactions frequencies in the endosperm in comparison to leaf tissue. Using cell-specific Hi-C and INTACT-RNA-seq, on a genome-wide scale higher expression of self-looped interacting genes was observed. It was further identified that interacting intergenic regions act as a negative regulator and influence the gene expression in a specific cells in the plant. Our study provides evidence that the higher-order chromatin structure differs between cell types in plants and these interactions could influence the transcription activity positively as well as negatively.

overall_design: Two independent biological replicates of the mHi-C library from Arabidopsis endosperm were generated in order to examine the tissue-specific spatial organization of chromatin. Two additional independent biological replicates from 4-week-old leaves of Arabidopsis were also processed with the purpose of corroborating the data generated by our mHi-C method and comparing them with previously published conventional Hi-C data from leaf.

first: Vikash

last: Kumar Yadav

first: Claudia

last: Köhler


title: p38α plays differential roles in hematopoietic stem cell activity dependent on aging contexts

overall_design: we injected CAG-CreERT2: p38αflox/flox (p38αfl/fl) mice aged approximately 1 year (50–60 weeks) with TAM to delete p38α, and then analyzed those mice and corresponding controls 1 year later (i.e., at 2 years-of-age) (p38αcKO). lineage-Sca-1+c-Kit+CD34-Flt3- cells (LT-HSC) in the bone marrow (femur and tibia) from 2-year-old wild type (WT) or p38αcKO C57BL/6J mice (Ly5.2) were sorted into SF-O3 medium. Then, sorted cells were centrifuged at 340 x g for 5 min at 4°C and lysed with 75µL RLT buffer + 0.75µL 2-ME. RNA extraction, cDNA synthesis, microarray analysis, and data normalization were outsourced to DNA Chip Research Inc.


title: Transcriptomic profiles of seven M. abscessus isolates during log growth

city: Pittsburgh

first: Rebekah

last: Dedrick

address: 4249 Fifth Ave

department: Biological Sciences

institute: University of Pittsburgh

summary: The goal of this sequencing project was to measure transcription levels of the bacterial genes, prophages and plasmids

overall_design: Total RNA was isolated from cultures grown in log phase, depleted of rRNA and prepared for sequencing using a strand-specific library kit during log growth.

first: Graham

last: Hatfull

first: Rebecca

last: Garlena


title: Assessing the plasticity of the senescent phenotype

first: Jason

last: Vander Heiden

summary: The senescent phenotype was evaluated to observe potential changes over time and on differing culture substrates.

overall_design: While numerous studies have characterized the in vitro-derived senescent phenotype of cells in culture, all of these studies observed cells within a short time-frame and while cultured on tissue culture plastic. We investigated whether the senescent phenotype changes over time or is affected by the substrate/context in which the cells are cultured. To this end, we worked with primary small airway epithelial cells where senescence was induced while cells were cultured on Matrigel. In addition, we also cultured senescent cells for a short, 5 days, and long period, 12 days, of time to observe if the phenotype of these cells is altered.

first: Daryle

last: DePianto

first: Katrina

last: Morshead

first: Joseph

last: Arron


title: GAF is essential for zygotic genome activation and chromatin accessibility in the early Drosophila embryo

city: Madison

first: Melissa

last: Harrison

email: mharrison3@wisc.edu, ykaur@wisc.edu, torreszelada@wisc.edu, mmfreund2@wisc.edu

state: WI

address: 420 Henry Mall

department: Biomolecular Chemistry

institute: University of Wisconsin Madison

overall_design: Refer to individual Series


title: DNA methylation analysis of acute myeloid leukemia (AML)

city: Columbus

first: Christopher

last: OAKES

state: OH

address: 400 W. 12th Avenue, Wiseman Hall, Suite 455

institute: The Ohio State University

summary: Acute myeloid leukemia (AML) is a molecularly complex disease characterized by heterogeneous tumor genetic profiles and involving numerous pathogenic mechanisms and pathways. Integration of molecular data types across multiple patient cohorts may advance current genetic approaches for improved sub-classification and understanding of the biology of the disease. Here we analyzed genome-wide DNA methylation in 649 AML patients using Illumina arrays and identified a configuration of 13 subtypes (termed ‘epitypes’) using unbiased clustering. Integration of genetic data revealed that most epitypes were associated with a certain recurrent mutation (or combination) in a majority of patients, yet other epitypes were largely independent. Epitypes demonstrated developmental blockage at discrete stages of myeloid differentiation, revealing epitypes that retain arrested hematopoietic stem cell-like phenotypes. Detailed analyses of DNA methylation patterns identified unique patterns of aberrant hyper- and hypomethylation among epitypes, with variable involvement of transcription factors influencing promoter, enhancer and repressed regions. Patients in epitypes with stem cell-like methylation features showed inferior overall survival along with upregulated stem cell gene expression signatures. We further identified a DNA methylation signature involving STAT motifs associated with FLT3-ITD mutations. Finally, DNA methylation signatures were remarkably stable at relapse for the large majority of patients, and rare epitype switching accompanied loss of the dominant epitype mutations and reversion to stem cell-like methylation patterns. These results demonstrate that DNA methylation-based classification integrates important molecular features of AML to reveal the diverse pathogenic and biological aspects of the disease.

overall_design: Genomic DNA was extracted from from bone marrow derived Mononuclear cells collectected from 272 patients with AML at diagnosis and a matched relapse sample from 28 patients. Following bisulfite conversion the samples were hybridised to the Infinium methylationEPIC assay following standard protocol. Additional samples are matched diagnosis and relapse samples from an additional 8 patients.

first: Brian

last: Giacopelli

last: Oakes


title: GAF is essential for zygotic genome activation and chromatin accessibility in the early Drosophila embryo [RNA-seq]

summary: Following fertilization, the genomes of the germ cells are reprogrammed to form the totipotent embryo. Pioneer transcription factors are essential for remodeling the chromatin and driving the initial wave of zygotic gene expression. In Drosophila melanogaster, the pioneer factor Zelda is essential for development through this dramatic period of reprogramming, known as the maternal- to-zygotic transition (MZT). However, it was unknown whether additional pioneer factors were required for this transition. We identified an additional maternally encoded factor required for development through the MZT, GAGA Factor (GAF). GAF is necessary to activate widespread zygotic transcription and to remodel the chromatin accessibility landscape. We demonstrated that Zelda preferentially controls expression of the earliest transcribed genes, while genes expressed during widespread activation are predominantly dependent on GAF. Thus, progression through the MZT requires coordination of multiple pioneer-like factors, and we propose that as development proceeds control is gradually transferred from Zelda to GAF.

overall_design: RNA-seq transcriptomic profiling was performed in triplicate on pools of 150-200 GAFdeGradFP embryos lacking GAF protein, or control embryos.

first: Marissa

last: Gaskill

first: Tyler

last: Gibson

first: Elizabeth

last: Larson


title: Tissue-dependent regulation of miRNA activity

city: Tuebingen

first: Marja

last: Timmermans

address: Morgenstelle 32

country: Germany

institute: University of Tuebingen

summary:
miRNAs play important roles in every aspect of plant development. Small RNA regulation confers sensitivity and robustness onto gene regulatory networks, and the morphogen-like readout of small RNA mobility gradients
yields sharply delineated domains of target gene expression. However, how the spatiotemporal patterns of miRNA activity are attained is less well understood.
We used the high-resolution maize shoot apex transcriptome atlas (Knauer et al., 2019) together with data from small RNA-Seq, PARE analysis, small RNA in-situ hybridization, and RNAseq of weak dcl1 mutant tissues
to demonstrate that miRNA accumulation and function during development is regulated in a tiissue dependent manner at the transcriptional and post-transcriptional level.

overall_design: small RNA-seq, 1 replicate, of maize shoot apices (SAM and up to four leaf primordia); Bulk RNA-seq, 2x replicates, of the SAM center zone of wild type or dcl1-2 captured by LCM (Knauer et al., 2019); Bulk RNA-seq, 2x replicates, of leaf primordia (P3) of  wild type or dcl1-2 captured by LCM (Knauer et al., 2019).


title: Human iris pigment epithelial cell (HIPE) response to ZIKV infection

city: Adelaide

first: Feargal

last: Ryan

state: Adelaide

address: North Terrace

institute: SAHMRI

summary: Pathogenesis of anterior uveitis (ocular manifestation of ZIKV infection in adults, distinct from children).

overall_design: 20 HIPE donors with mock and Zika Virus infected. Collected at one timepoint at 24 hours.

first: David

last: Lynn

first: Justine

last: Smith


title: GAF is essential for zygotic genome activation and chromatin accessibility in the early Drosophila embryo [ChIP-seq]

summary: Following fertilization, the genomes of the germ cells are reprogrammed to form the totipotent embryo. Pioneer transcription factors are essential for remodeling the chromatin and driving the initial wave of zygotic gene expression. In Drosophila melanogaster, the pioneer factor Zelda is essential for development through this dramatic period of reprogramming, known as the maternal-to-zygotic transition (MZT). However, it was unknown whether additional pioneer factors were required for this transition. We identified an additional maternally encoded factor required for development through the MZT, GAGA Factor (GAF). GAF is necessary to activate widespread zygotic transcription and to remodel the chromatin accessibility landscape. We demonstrated that Zelda preferentially controls expression of the earliest transcribed genes, while genes expressed during widespread activation are predominantly dependent on GAF. Thus, progression through the MZT requires coordination of multiple pioneer-like factors, and we propose that as development proceeds control is gradually transferred from Zelda to GAF.

overall_design: GAF ChIP-seq was performed in duplicate using anti-GFP antibody (Abcam #ab290) on embryos in which endogenous GAF protein was tagged with super folder GFP (sfGFP). Sequencing was performed on immunoprecipitated samles as well as input controls. Additional  immunoprecipitations were performed using the anti-GFP antibody in w1118 to control for nonspecific pulldown. Zelda ChIP was performed in duplicate with the anti-Zelda antibody (Harrison et al., PLoS Genetics 2011) with input controls.


title: The lncRNA MIR31HG regulates the senescence associated secretory phenotype

city: Copenhagen

first: Anders

middle: H.

last: Lund

address: Ole Maaløes Vej 5

department: Biotech Research & Innovation Centre (BRIC)

institute: University of Copenhagen

summary: Oncogene-induced senescence provides a barrier against malignant transformation. Senescent cells secrete pro-inflammatory cytokines, chemokines and growth factors collectively known as the senescence-associated secretory phenotype (SASP). Paradoxically, the SASP can also negatively impact the neighbouring tissues through inducing an inflammatory environment that promotes tumorigenesis and aged-related pathologies. We have previously shown that the lncRNA MIR31HG is overexpressed and located in the cytoplasm during BRAF-induced senescence. In young proliferating cells, nuclear MIR31HG inhibits p16/CDKN2A expression through interaction with polycomb repressor complexes (PRC1/2). Here, we show that MIR31HG regulates the expression and secretion of a subset of SASP components during BRAF-induced senescence. The SASP secreted from senescent cells depleted for MIR31HG fails to induce paracrine invasion without affecting the growth inhibitory effect. Mechanistically, MIR31HG interacts with the translation factor YBX1 facilitating its phosphorylation at serine 102 (p-YBX1S102) by the kinase RSK.  p-YBX1S102 induces IL1A translation which acts as upstream regulator inducing the transcription of the other SASP mRNAs. Our results suggest a dual role for MIR31HG in senescence depending on its cellular localization and place the lncRNA as a potential therapeutic target in the treatment of senescence-related pathologies.

overall_design: Poly-A selected RNA-seq from fibroblast BJ cells, Oncogene-induced senescent (OIS) BJ cells, OIS BJ cells with knockdown of MIR31HG and OIS BJ cells with knockdown of YBX-1

first: Marta

last: Montes

first: Michal

last: Lubas

first: Frederic

middle: S

last: Arendrup

first: Bettina

last: Mentz

first: Sarunas

last: Tumas

first: Lea

last: Harder

first: Jens

last: Andersen

middle: H


status: Public on Mar 02 2021

first: CBDM

last: Lab

state: MA

address: 77 Avenue Louis Pasteur

department: Microbiology and Immunobiology

institute: Harvard Medical School

summary: Single-cell RNAseq (10x Genomics) analysis of human CD4+ T cells in IPEX patients, healthy donors and heterozygous mothers (blood). Human CD4+T cells from IPEX, HD and mothers were isolated from frozen peripheral blood mononuclear cells by flow cytometry as DAPI–CD3+CD4+ cells. In cohort 1, cells from separate donor were encapsulated in separate channel following 10x Genomics Single Cell 3′ Reagent Kit (V2 chemistry). In cohort 2, samples were tagged using a different Hashtags, pooled and encapsulared following 10x Genomics Single Cell 3′ Reagent Kit (V3 chemistry).

last: Zemmour

first: Louis

last: Charbonnier

first: Juliette

last: Leon

first: Tala

last: Chatila

first: Diane

last: Mathis

first: Christophe

last: Benoist


title: The role of Phc1 in organizing the chromatin interactions of the Nanog locus in ESCs

city: Hangzhou

first: Li

last: Chen

address: 866 Yuhangtang Rd

department: School of Medicine

institute: Zhejiang University

summary: To look into whether Phc1 is involved in organizing the chromatin interactions of the Nanog locus in ESCs. We performed circularized chromosome conformation capture sequencing (4C-seq) of the genome-wide regions interacting with the Nanog locus in both Phc1+/+ and Phc1-/- mESCs. By doing so, we showed that ablation of Phc1 impaired the genome-wide chromatin interactions of the Nanog locus in mESCs.

overall_design: 4C-seq of the genome-wide regions interacting with the Nanog locus in both Phc1+/+ and Phc1-/- mESCs.

first: Penglei

last: Jiang

first: Junfeng

last: Ji