title: Associations between IFI44L gene variants and rates of respiratory tract infections during early childhood

status: Public on Aug 18 2020

city: Dallas

first: Nicole

last: Baldwin

address: 3434 Live Oak St

institute: Baylor Research Institute

summary: Background: Genetic heterogeneity in the innate immune system may account for variable susceptibility to respiratory tract infections (RTIs) in children.  Objective: We aimed to assess the impact of polymorphisms rs273259 and rs1333969 in type I interferon related gene IFI44L on susceptibility to RTIs and acute otitis media in children.  Methods: In two prospective, population-based birth cohorts, the FinnBrain Birth Cohort Study and the STEPS Study, IFI44L genotypes for rs273259 and rs1333969 were determined in relation to the development of RTIs until one and two years of age, respectively, and adjusted incidence rate ratios (aIRR) or odds ratios (OR) were calculated. At age 3 months, whole blood transcriptional profiles were analyzed and nasal samples were tested for respiratory viruses in a subset of children.   Results: In respiratory virus-positive children at 3 months of age, IFI44L gene variants were associated with decreased expression levels of IFI44L and several other interferon related genes.  Conclusions: Variant forms of IFI44L gene were protective against early-childhood RTIs or acute otitis media in two independent birth cohorts, and they attenuated interferon pathway activation by respiratory viruses.

overall_design: 71 samples, one each from 71 different subjects; no replicates; no controls; subjects at time of collection were afebrile and without symptoms of an RTI

first: Johanna

last: Lempainen

first: Laura

last: Korhonen

first: Katri

last: Kantojärvi

first: Santtu

last: Heinonen

last: Toivonen

first: Panu

last: Räty

first: Octavio

last: Ramilo

first: Asuncion

last: Mejias

first: Antti-Pekka

last: Laine

first: Tytti

last: Vuorinen

first: Matti

last: Waris

first: Linnea

last: Karlsson

first: Hasse

first: Tiina

last: Paunio

first: Ville

last: Peltola


title: Transcriptomes of several organs of lean and obese mice infected with avian influenza

status: Public on Apr 14 2020

city: Parkville

first: Boris

last: Novakovic

state: Victoria

address: Murdoch Children's Research Institute, Royal Children's Hospital, Flemington Road

country: Australia

institute: Murdoch Children's Research Institute

type: Expression profiling by high throughput sequencing

summary: Influenza A virus (IAV) causes a wide range of extra-respiratory complications, ranging from central nervous system disease to cardiac disease. Obesity/overweight has previously been identified as a susceptibility factor for both severe influenza and cardiac disease. We therefore sought to investigate the systemic pathogenesis of influenza in an obese mouse model with a focus on cardiac complications of influenza.

overall_design: C57BL/6 mice were fed either a high-fat diet consisting of 40% calories from fat or a low-fat diet consisting of 12% calories from fat for 10 weeks and supplied with water ad libitum. The mice were then inoculated with influenza A/Auckland/09(H1N1). Lung, heart and brain was collected from naive or infected mice and subjected to RNA sequencing.

first: Jurre

last: Siegers

first: Debby

last: van Riel

first: Kirsty

last: Short


title: Blood genome expression Profiles in Infants with Congenital Cytomegalovirus Infection: Is the asymptomatic infant truly asymptomatic?

status: Public on May 26 2020

first: Nicole

last: Baldwin

country: USA

summary: Background: Congenital cytomegalovirus (cCMV) infection represents the most frequent non-genetic cause of sensorineural hearing loss (SNHL). The disease spectrum varies from clinically apparent (“symptomatic”) to clinically inapparent (“asymptomatic”). Analysis of host genome expression profiles has provided new insights into disease pathogenesis and assessment of clinical severity. Its value as a biomarker in infants with cCMV has not been explored.  Methods: Infants with cCMV infection, both symptomatic and asymptomatic, were enrolled at a median age of 17 days of age and followed up for 3 years to assess the development of SNHL. Healthy age matched controls were also enrolled as a reference. Blood samples were obtained at enrollment and follow-up to assess host responses by RNA transcriptional profiling. Symptomatic CMV was defined as the presence of any clinical, laboratory, ophthalmologic, audiologic or neuroimaging abnormalities.

overall_design: 158 total samples; 56 Asymptomatic; 92 Symptomatic; 10 Control

first: Christopher

last: Ouellette

first: Pablo

middle: J

last: Sanchez

first: Fiker

last: Zeray

first: Andrea

last: Ronchi

first: Lisa

last: Jaramillo

first: Masako

last: Shimamura


title: GH directly inhibits steatosis and liver injury in a sex-dependent and IGF1-independent manner

status: Public on Jul 11 2020

city: Chicago

first: Rhonda

middle: D

last: Kineman

state: Illinois

address: 1819 West Polk St

department: Medicine

institute: University of Illinois at Chicago

summary: A reduction in hepatocyte growth hormone (GH)-signaling promotes non-alcoholic fatty liver disease (NAFLD). However, debate remains as to the relative contribution of the direct effects of GH on hepatocyte function vs indirect effects, via alterations in insulin-like growth factor 1 (IGF1). To isolate the role of hepatocyte GH receptor (GHR) signaling, independent of changes in IGF1, mice with adult-onset, hepatocyte-specific GHR knockdown (aHepGHRkd) were treated with a vector expressing rat IGF1 targeted specifically to hepatocytes. Compared to GHR-intact mice, aHepGHRkd reduced circulating IGF1 and elevated GH. In male aHepGHRkd, the shift in IGF1/GH did not alter plasma glucose or non-esterified fatty acids (NEFA), but was associated with increased insulin, enhanced systemic lipid oxidation and reduced white adipose tissue (WAT) mass. Livers of male aHepGHRkd exhibited steatosis associated with increased de novo lipogenesis, hepatocyte ballooning and inflammation. In female aHepGHRkd, hepatic GHR protein levels were not detectable, but moderate levels of IGF1 were maintained, with minimal alterations in systemic metabolism and no evidence of steatosis. Reconstitution of hepatocyte IGF1 in male aHepGHRkd lowered GH and normalized insulin, whole body lipid utilization and WAT mass. However, IGF1 reconstitution did not reduce steatosis or eliminate liver injury. RNAseq analysis showed IGF1 reconstitution did not impact aHepGHRkd-induced changes in liver gene expression, despite changes in systemic metabolism. These results demonstrate the impact of aHepGHRkd is sexually dimorphic and the steatosis and liver injury observed in male aHepGHRkd mice is autonomous of IGF1, suggesting GH acts directly on the adult hepatocyte to control NAFLD progression.

overall_design: We analyzed total RNA from male and female livers (6 mice/sex) from GHR-intact controls mice and adult-onset, hepatocyte-specific GHR knockdown mice, without (aHepGHRkd) and with hepatic reconstitution of IGF1 (aHepGHRkd+2.0*1011 AAV8-TBGp-rIGF1).

first: R


title: Single-cell RNA-seq of the developing C57/BL6 mouse back skin at E13.5, E16.5 and P0

status: Public on Jul 01 2020

city: Qingdao

first: Wei

last: Ge

state: Shandong Province

address: Changcheng Road

department: College of life sciences

country: China

institute: Qingdao Agricultural University

summary: We performed single-cell RNA seq on C57/BL6 mouse back skin at E13.5, E16.5, and P0 to study embryonic hair follicle development. We analyzed 15,086 single cell transcriptome profiles from E13.5, E16.5 and newborn mice (postnatal day 0, P0) dorsal skin cells across hair follicle induction, organogenesis, cytodifferentiation stage. Based on t-distributed Stochastic Neighbor Embedding (tSNE) clustering, we identified 14 cell clusters from skin cells and delineated their cell identity gene expression profile. By using Monocle pseudotime ordering analysis, we constructed epithelium/dermal cell lineage differentiation trajectory and revealed sequential activation of key regulons involved during embryonic hair follicle morphogenesis. Our findings here provide molecular landscape during hair follicle epithelium/dermal cell lineage fate decisions.

first: Ge

last: Wei


title: Mast cells elicit protection against colorectal tumors by modulating the immune response during early tumorigenic steps

status: Public on Mar 13 2020

city: Ribeirao Preto

first: Juliana

middle: Yumi

last: Sakita

state: Sao Paulo

address: Av. do Café, s/ nº

department: Genetics

country: Brazil

institute: Ribeirao Preto Medical School - University of Sao Paulo

summary: Mast cells (MC) alter the development of colorectal cancer (CRC), although their precise role remains poorly defined. Here we show that MC numbers vary in CRC patients, which directly impacts on lymphocytes population. Mouse-based mechanistic experiments reveal that an intricate MC-lymphocyte axis can either promote or inhibit the CRC development, as well as be pharmacologically explored. Therefore, MC activity can be modulated and inhibit the early steps of CRC development.

overall_design:
Colon tumor mRNA profiles of AOM-exposed Kit-B6 and Kit-W/sh mice were generated by RNA sequencing using QIAseq Targeted RNAseq Mouse Immuno-Oncology panel and QIAseq Targeted RNA 96-Index I (#333117) in a in MiSeq Sequencing System (Illumina; Illumina MiSeq Reagent Kit v3 - 150 cycles, #15043894).

Please note that the records have been updated with additional RNA-seq data from allographic tumor samples on Mar 23, 2023.

first: Vinicius

last: Kannen

middle: Y

first: Luciana

last: Yamamoto Almeida

first: Emerson

last: de Souza Santos

first: Tathiane

middle: M

last: Malta

first: Mariângela

middle: O

last: Brunaldi

first: Sergio

last: Albuquerque

first: Cleide

middle: L

last: Araújo Silva

first: Daniela

last: Carlos

first: Marcus

middle: V

last: Andrade

last: Britto Garcia

first: Fernando

middle: Q

last: Cunha

first: Guilherme

middle: C

last: Martelossi Cebinelli

first: Madhuri

last: Koti

first: Jeremy

middle: A

last: Squire

last: Uyemura


title: RSV disease severity: Insights into Protective Immune Responses

status: Public on Apr 22 2020

first: Nicole

last: Baldwin

summary: This was a prospective observational cohort study involving a convenience sample of previously healthy children <2 years of age with acute RSV infection as well as healthy non-infected age matched controls. The study was conducted at Nationwide Children’s Hospital (NCH; Columbus, OH) during four respiratory seasons, from 2011 to 2015.  Children were enrolled at the NCH urgent care clinics, the emergency department or in the inpatient ward or intensive care unit (PICU). For the inpatients enrolled at ward or PICU the median time from admission to sample collection was 21.3h (interquartile range [IQR] 16.7 -37.6h). The majority of children had a confirmatory RSV test (either rapid antigen testing or PCR based) at enrollment per standard of care. In addition, the presence of RSV was confirmed by quantitative real time (qRT)-PCR in all study subjects.  Healthy age-matched controls were enrolled during well-child visits or minor elective surgical procedures not involving the respiratory tract as described [Mejias 2013].  Exclusion criteria were as follows: documented bacterial co-infections, premature birth (<36 weeks of gestation), chronic or congenital medical conditions, and immunodeficiency. For healthy controls, additional exclusion criteria included: presence of fever or symptoms of respiratory tract infection within two weeks of enrollment.

overall_design: 127 total samples. 5 technical replicates of Healthy samples (noted in description column). 38 Healthy controls (including technical replicates), 33 RSV Outpatient, 56 RSV Inpatient

first: Santtu

last: Heinonen

first: Victoria

last: Velazquez

first: Fang

last: Ye

first: Sara

last: Mertz

first: Bennett

last: Smith

first: Cristina

last: Garcia-Maurino

first: Silvia

last: Oliva

first: Daniel

last: Cohen

first: Octavio

last: Ramilo

first: Asuncion

last: Mejias


title: ChIP-seq Profiling of Histone H3K27ac, BRD4-NUT, and RNA Polymerase II Chromatin Localization During Megadomain Formation/Removal and RNA Polymerase II Inhibition

status: Public on Apr 01 2020

city: Houston

first: Kyle

last: Eagen

state: TX

address: 1 Baylor Plaza

department: Department of Molecular and Cellular Biology

institute: Baylor College of Medicine

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Delineating how chromosomes fold at length scales beyond one megabase remains obscure relative to smaller-scale folding into TADs, loops, and nucleosomes. We find that rather than simply unfolding chromatin, histone hyperacetylation results in interactions between distant genomic loci separated by tens to hundreds of megabases, even in the absence of transcription. These hyperacetylated “megadomains” are formed by the BRD4-NUT fusion oncoprotein, interact both within and between chromosomes, and form a specific nuclear subcompartment that has elevated gene activity with respect to other subcompartments. Pharmacological degradation of BRD4-NUT results in collapse of megadomains and attenuation of the interactions between them. In contrast, these interactions persist and contacts between newly acetylated regions are formed after inhibiting RNA polymerase II initiation. Our structure-function approach thus reveals that broad chromatin domains of identical biochemical composition, independent of transcription, form nuclear subcompartments, and also indicates the potential of altering chromosome structure for treating human disease.

overall_design: ChIP-seq to assess histone H3K27ac, BRD4-NUT, and RNA polymerase II localization after induction of BRD4-NUT expression in 293TRex cells and after MZ1 treatment to degrade BRD4-NUT, after MZ1 treatment followed by drug washout to partially restore BRD4-NUT protein levels, and after triptolide treatment to inhibit RNA polymerase II transcription in TC-797 cells.

first: Celeste

last: Rosencrance

first: Haneen

middle: N

last: Ammouri

first: Qi

last: Yu

first: Tiffany

last: Ge

middle: P


title: Transcriptomes of human trained macrophages before and after Mtb exposure

status: Public on Dec 10 2019

first: Boris

last: Novakovic

summary: Trained Immunity (TRIM) is the phenomenon whereby innate immune cells such as monocytes or macrophages undergo functional reprogramming after exposure to specific microbial or metabolic components. In this study, monocytes were trained with beta-glucan and then re-exposed to Mycobacterium tuberculosis.

overall_design: In this study human monocytes were exposed to beta-glucan for 24 hours in vitro, a stimulus that induced TRIM, and were then differentiated to macrophages for another 5 days. At day 6, trained and not-trained macrophages were exposed to Mycobacterium tuberculosis (Mtb) for 4 hours and their transcriptomes were profiled .

first: Simone

last: Moorlag

first: Mihai

last: Netea


title: Mycobacterium tuberculosis cording in lymphatic endothelial cells (hLEC)

status: Public on Jun 04 2020

city: London

first: Rachel

last: Lai

address: Du Cane Road

department: Infectious Disease

country: United Kingdom

institute: Imperial College London

summary: Mycobacterium tuberculosis (Mtb) displayed cording phenotypes during intracellular infection. RNA-Seq was used to identify transcriptional programmes expressed by hLEC that actviates pathways relating to cellular pro-survival and cyosolic surveillance of intracellular Mtb. Mtb cording was found to be a mechanism to evade xenophagy in the cytosol of endothelial cells.

overall_design: A total of 6 samples were sequenced: 3 samples of Mtb-infected hLEC and 3 samples of uninfected hLEC

first: Max

last: Gutierrez

first: Tom

last: Lerner


title: Adipose tissue network analyses reveal negative association between GDF15 and BMI during weight management intervention.

city: Lausanne

first: Gregory

last: Lefebvre

address: EPFL Scientific Parc

country: Switzerland

institute: Nestlé Institute of Health Science

summary:
CONTEXT: Adipose tissue (AT) transcriptome studies provide holistic pictures of adaptation to weight and related bioclinical settings changes.
OBJECTIVE: To implement AT gene expression profiling and investigate the link between changes in bioclinical parameters and AT gene expression during three steps of a two-phase dietary intervention (DI).
RESULTS: During LCD, 5 modules were found, 3 of which including at least 1 bio-clinical variable. Especially, the change in BMI was connected to changes in mRNA level of genes with Inflammatory response signature. In this module, BMI was negatively connected to several genes encoding secreted proteins including GDF15, CCL3 and SPP1. Whatever the phase of the DI, change in GDF15 was connected to changes in SPP1, CCL3, LIPA and CD68.
CONCLUSION: network analyses confirm GDF15 as upregulated with calorie restriction-induced weight loss, concomitantly to macrophages markers.

overall_design: AT transcriptome profiling was obtained from sequencing 1051 samples, corresponding to 556 distinct individuals enrolled in a weight loss intervention (8-week low calorie diet (LCD) at 800 kcal/d) followed with a 6-month ad libitum randomized dietary intervention (DI).


title: Characterization of estrogen and the endocrine-disrupting chemical-induced mouse mammary gland reorganization via single-cell RNA-sequencing

status: Public on Nov 12 2019

city: Duarte

first: Xiwei

last: Wu

state: CA

address: 1500 E. Duarte Rd.

department: Computational and Quantitative Medicine

institute: City of Hope National Medical Center

summary: Exposure to estrogen and its mimics during menopausal transition is thought to modify the structure of mammary gland. Using a surgical menopausal (ovariectomized) mouse model, we assessed how mammary gland tissue was affected by both 17β-estradiol (E2) and polybrominated diphenyl ethers (PBDEs). Three analyzed flame retardants, BDE-47, BDE-100 and BDE-153, are most frequently detected in human serum from all over the world. During physiologically-relevant exposure to E2, PBDEs enhanced E2-mediated regrowth of mammary glands with terminal end bud-like structures. According to sequencing analysis of mammary gland RNA, PBDEs both augmented E2-facilitated cell proliferationgene expression changes and modulated immune regulation. To better elucidate the effects of E2 and PBDEs, single-cell RNA sequencing (scRNAseq) analysis was performed. In a definitive manner, E2 was found to induce Pgr expression in both Esr1+ and Esr1- cells. Significantly, molecular evidence was available to show functional differences among cells with differential expression of Esr1 and Pgr. The PBDEs + E2 treatment increased the number of double positive (Esr1+Pgr+) and Pgr only (Esr1-Pgr+) cells in both mature luminal epithelial and progenitor cells of luminal epithelialcells.  Moreover, the combination increased the mammary gland population of M2 macrophages. The results help clarify how exposure to both E2 and endocrine disrupting chemicals like PBDEs during menopausal transition  impact mammary gland structure. Ultimately, the findings advance understanding of how exposure can increase the risk of developing breast cancer.

overall_design: Female BALB/cj mice were exposed to E2 and a mixture of PBDEs for 1 week.  Single-cell RNA sequencing was done to evaluate gene expression changes among treatment groups. Mice were subjected to 3 different treatments (Vehicle, E2, E2+PBDE).

first: Chen

last: Shiuan

first: Noriko

last: Kanaya


title: EGR1 genomic binding in t-HESC cell line

status: Public on Nov 26 2019

first: Maria

last: Szwarc

address: One Baylor Plaza

institute: Baylor College of MEdicine

summary: Whole genome binding profile of EGR1 in human endometrial stromal cells

overall_design: ChIP-sequencing of EGR1 in human endometrial stromal cells

first: John

last: Lydon


title: TGFβ superfamily signaling regulates the state of human stem cell pluripotency and competency to create telencephalic organoids

status: Public on Jan 01 2020

city: Los Angeles

first: Bennett

last: Novitch

state: California

address: 650 Charles E Young Dr. S, CHS 66-200

department: Department of Neurobiology

institute: University of California, Los Angeles

summary: Telencephalic organoids generated from human pluripotent stem cells (hPSC) are emerging as an effective system to study the distinct features of the developing human brain and the underlying causes of many neurological disorders. While progress in organoid technology has been steadily advancing, many challenges remain including rampant batch-to-batch and cell line-to-cell line variability and irreproducibility. Here, we demonstrate that a major contributor to successful cortical organoid production is the manner in which hPSC are maintained prior to differentiation. Optimal results were achieved using fibroblast-feeder-supported hPSC compared to feeder-independent cells, related to differences in their transcriptomic states. Feeder-supported hPSC display elevated activation of diverse TGFβ superfamily signaling pathways and increased expression of genes associated with naïve pluripotency. We further identify combinations of TGFβ-related growth factors that are necessary and together sufficient to impart broad telencephalic organoid competency to feeder-free hPSC and enable reproducible formation of brain structures suitable for disease modeling.

overall_design: Bulk RNA sequencing (RNA-seq) analysis on triplicate samples of both H9 and XFiPSC telencephalic organoids grown under both mouse embryonic fibroblast and feeder-free conditions (12 samples in total).

first: Jillian

middle: R

last: Haney

first: Bennet

middle: G

first: Momoko

last: Watanabe


title: ISR activation mediates the behavioral and neurophysiological abnormalities in Down syndrome

status: Public on Nov 02 2019

first: Ya

last: Cui

state: TEXAS

address: One Baylor Plaza, Baylor College of Medicine, 529DH

summary: Down syndrome (DS) is the most common genetic cause of intellectual disability and a major biomedical concern. Despite a general consensus that protein homeostasis is essential for normal brain function, little is known about its role in DS. Here, we show that the integrated stress response (ISR)—a conserved signaling network that maintains proteostasis by controlling protein synthesis—is activated in the brains of DS mice and individuals with DS, leading to reduced translation. Genetic and pharmacological suppression of the ISR, either by inhibiting the ISR-initiating double-stranded RNA-activated protein kinase (PKR) or boosting the function of the eIF2•eIF2B complex, which becomes limiting upon ISR activation, de-repressed translation, reversed enhanced inhibitory synaptic transmission, and rescued the deficits in synaptic plasticity and memory in DS mice. Our findings unveil a crucial role for the ISR in DS pathophysiology and suggest tuning of the ISR as a promising therapeutic intervention for DS.

overall_design: Ribosome associated mRNA-seq of Ts65Dn, Ts65Dn-Pkr-/- and WT Mus Musculus

first: Ping

last: Zhu


title: 10X skeletal muscle regeneration scRNA-seq

status: Public on Mar 25 2020

city: West Lafayette

first: Shihuan

last: Kuang

state: IN

address: 901 W State St

institute: Purdue University

summary: Single-cell RNA sequencing of cells dissociated from skeletal muscle at discrete regeneration timepoints to reveal transcriptional identities of contributors to skeletal muscle regeneration.

overall_design:
Tibialis anterior muscle from C5Bl6 WT male mice were injected with cardiotoxin to induce muscle injury and collected at  at 0.5, 2, 3.5, 5, 10, 21 days post injury and a non-injured sample. Muscle was digested with collagenase and dispase to dissociate single cells, stained with a cell viability marker and subjected to FACS to generate single cell suspensions. Single cell suspensions were loaded onto the 10X Genomics Platform and processed according to the manufacturer's instructions. Each sample is an individual time point, n=3 male mice combined per time point.

The regen_data.rds file is list of two annotated data frames, of which each item contains the merged dataset with all cells from all timepoints.
Item [1] which contains the RNA counts while item [2] contains the SCT normalized values. Columns are the samples, rows and the gene values and the metadata is structures accordingly.

first: Stephanie

last: Oprescu


title: Transcriptomic analysis of mRNA expression changes induced by RANKL expression in the murine salivary gland

status: Public on Oct 28 2019

summary: Sequencing of mRNA isolated from salivary glands of control mice and mice overexpressing RANKL in keratin 5 (K5)-positive cells for 2 weeks.

overall_design: Control (monogenic control) and K5-rtTA:TetO-RANKL mice were administered doxycycline (in food and water) at 9-weeks of age for 2 weeks to induce transgenic RANKL expression. Each RNA sample was isolated from pooled salivary gland tissues from 5 mice.


title: oxLDL exposed trained monocytes

status: Public on Nov 07 2019

first: Boris

last: Novakovic

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

overall_design: Refer to individual Series


title: Epigenomes of oxLDL exposed trained monocytes

summary: Innate immune memory is the phenomenon whereby innate immune cells such as monocytes or macrophages undergo functional reprogramming after exposure to microbial components.  In this study we apply epigenetic, H3K27ac and H3K4me3 ChIP-seq, analysis to healthy human monocytes exposed to oxidized Low Density Lipoprotein (oxLDL), in vitro.

overall_design: Monocytes from healthy donors were cultured in vitro for 24 hours with media or oxLDL

first: Samuel

last: Keating

first: Niels

last: Riksen


title: Transcriptomes of human monocytes from patients with familial hypercholesterolemia before and after statin treatment

status: Public on Sep 30 2019

summary: Innate immune memory, also refered to as trained immunity (TRIM) is the phenomenon whereby innate immune cells such as monocytes or macrophages undergo functional reprogramming after exposure to microbial components or metabolites. The cholesterol pathway is also involved in TRIM, and in this study we compared the transcriptomes of patients with hypercholesterolemia before and after statin treatment for 3 months.

overall_design: Monocytes from healthy controls and patients with hypercholestoremia before and after 3 months of Statin treatment

first: Siroon

last: Bekkering

first: Mihai

last: Netea


title: Gene expression profile in mouse skeletal muscle after Mettl21e deletion

status: Public on Oct 22 2019

summary: Analysis of tibialis anterior (TA) muscle isolated from wildtype (WT) and Mettl21e deletion mice (KO). Results provide unbiased gene expression profile of TA muscle after Mettl21e deletion.

overall_design: Two-condition experiment, WT TA vs. KO TA. Biological replicates: 3 WT replicates, 3 KO replicates. Total RNAs were isolated using Trizol reagent (Invitrogen). Total RNAs were quality monitored by Agilent 2100 analysis, and sequenced by Illumina Genome Analyzer in Novogene Corporation., Beijing, China.

first: Chao

last: Wang

first: Bing

last: Zhang


title: Robust RNA-Seq of aRNA-amplified single cell material collected by patch clamp

status: Public on Jan 25 2020

first: Hugo

middle: Jae Mun

last: Kim

address: 4760 Templeton St, Unit 3113

department: Neuroscience

institute: UNIVERSITY OF CALIFORNIA, SAN DIEGO

summary: Most single cell RNA sequencing protocols start with single cells dispersed from intact tissue. High-throughput processing of the separated cells is enabled using microfluidics platforms. However, dissociation of tissue results in loss of information about cell location and morphology and potentially alters the transcriptome. An alternative approach for collecting RNA from single cells is to re-purpose the electrophysiological technique of patch clamp recording. A hollow patch pipette is attached to individual cells, enabling the recording of electrical activity, after which the cytoplasm may be extracted for single cell RNA-Seq (“Patch-Seq”). Since the tissue is not disaggregated, the location of cells is readily determined, and the morphology of the cells is maintained, making possible the correlation of single cell transcriptomes with cell location, morphology and electrophysiology. Recent Patch-Seq studies utilizes PCR amplification to increase amount of nucleic acid material to the level required for current sequencing technologies. PCR is prone to create biased libraries – especially with the extremely high degrees of exponential amplification required for single cell amounts of RNA. We compared a PCR-based approach with linear amplifications and demonstrate that aRNA amplification (in vitro transcription, IVT) is more sensitive and robust for single cell RNA collected by a patch clamp pipette.

overall_design:
We used 5 metrics to assess the RNA-Seq data of amplified products. The first two metrics were total mapping rate defined as a fraction of raw reads mapped to the genome and transcriptome (GenCode v22, GRCh38.p2) and transcriptome mapping ratio calculated as a percentage of mapped reads which mapped to the transcriptome, excluding both rRNA (ribosomal RNA) and mtRNA (mitochondrial RNA). The third metric was gene model discovery rate, assessed as the number of genes with more than 5 mapped reads detected per 3 million mapped reads. A Recent single cell RNA-Seq (scRNA-Seq) study indicated that more than one million reads are required to analyze the variance in expression[2]. We used 3 million as our baseline for our analysis to increase the gene complexity because we are targeting highly complex neuronal cells. With single cell amount (10pg) and sub-single cell amount (5pg) of RNA processed with several different library construction methods, we compared these five metrics .

Please note that the 'SMARTER-UCSD-UHR'* and 'Upenn_UHR10pg*' data columns (in the 'Partek_Hugo_SCUHRDS_transcript_counts.xlsx') contain re-analyzed data from publicly available data (GSE56638).

first: JaeMun

middle: H

first: Adrian

last: Camarena

first: Christopher

last: Walker

first: Ming-Yi

last: Lin

first: Victoria

last: Wolseley

first: Tade

last: Souaiaia

first: Matthew

last: Thornton

first: Brendan

last: Grubbs

first: Robert

last: Chow

first: Oleg

last: Evgrafov

first: James

last: Knowles


title: Mutant p63 disrupts the key specification switch from the multipotent cell state to stratified epithelia during epithelial differentiation/in ectodermal dysplasia disorders

status: Public on Aug 19 2019

city: Nijmegen

first: Jo Huiqing

last: Zhou

address: Geert Grooteplein 26/28

country: Netherlands

institute: Radboud University

summary: Transcription factor p63 is a key regulator of stratified epithelia. In humans mutations in p63 are associated with developmental disorders that manifest defects in stratified epithelia including the epidermis. We established an epidermal commitment model using human pluripotent stem cells (PSCs) and characterized differentiation defects of PSCs carrying p63 mutations. Transcriptome analyses revealed distinct phases of epidermal commitment, multipotent simple epithelial, basal stratified epithelial and mature epidermal fates. Differentiation defects of p63 mutant PSCs occurred during the specification switch from the simple epithelium to the basal stratified epithelial fate. Single-cell transcriptome and pseudotime analyses identified enhanced mesodermal signatures associated with the deviated commitment route of p63 mutant PSCs. Repressing mesodermal differentiation improved epidermal commitment of PSCs. Our study demonstrate that p63 is required for specification of stratified epithelia but not sufficient for epidermal maturation. It provides insights into disease mechanisms underlying defects of stratified epithelia caused by p63 mutations.

overall_design: Examination of gene expression during during epithelial differentiation.

first: Eduardo

last: Soares

first: Quan

last: Xu

first: Qingqing

last: Li

first: Fuchou

last: Tang

first: Huiqing


title: Combinatorial small molecule treatment enhances the in vitro maturation of pluripotent stem cell-derived myotubes

status: Public on Oct 29 2019

city: Minneapolis

first: Alessandro

last: Magli

state: MN

address: 2231 6th St SE

institute: University of Minnesota

summary: Targeted differentiation of pluripotent stem (PS) cells into myotubes enables in vitro disease modeling of skeletal muscle diseases. Although various protocols achieve myogenic differentiation in vitro, resulting myotubes invariably display an embryonic identity. This is a major hurdle for accurately recapitulating disease phenotypes in vitro, as disease typically does not manifest in the embryonic muscle, but at more mature stages. To address this problem, we identified four factors from a small molecule screen whose combinatorial treatment resulted in myotubes with enhanced maturation, as shown by increased expression of fetal, neonatal and adult myosin heavy-chain isoforms. These molecular changes were confirmed by global chromatin accessibility and transcriptome studies. Importantly, we also observed this maturation in three-dimensional muscle bundles, which displayed improved in vitro contractile force generation in response to electrical stimulus. Thus, we established a model for in vitro muscle maturation from PS cells.

overall_design: Three replicates each of human iPSC myotubes derived with vehicle (DMSO) or small molecule treatment (SDDF) were analyzed by 1) RNA seq to identify the differentially expressed genes and 2) ATAC-seq to determine changes in chromatin accessibility.

first: Selvaraj

last: Sridhar

first: Mondragon-Gonzalez

last: Ricardo

first: Magli

last: Alessandro

first: Perlingeiro

last: Rita


title: Transcriptomes of oxLDL exposed trained monocytes

first: Boris

last: Novakovic

summary: Innate immune memory is the phenomenon whereby innate immune cells such as monocytes or macrophages undergo functional reprogramming after exposure to microbial components.  In this study we apply RNA-seq analysis to healthy human monocytes exposed to oxidized Low Density Lipoprotein (oxLDL), in vitro.

first: Samuel

last: Keating

first: Niels

last: Riksen


title: HuR controls apoptosis and activation response without effects on cytokine 3′ UTRs

status: Public on Jun 05 2019

city: Riverside

first: Fedor

last: Karginov

address: 900 University Ave

department: MCSB

institute: UC Riverside

summary: RNA binding proteins (RBPs) regulate gene expression through several post-transcriptional mechanisms. The broadly expressed HuR/ELAVL1 RBP is important for proper function of multiple cell types in the immune system, and has been proposed to positively control the expression of cytokine and other mRNAs upon activation by regulating their 3′ UTRs. However, this mechanism has not been previously dissected in stable cellular settings. In this study, HuR demonstrated strong anti-apoptotic and activation roles in the Jurkat T cell line. Detailed transcriptomic analysis of HuR knockout cells revealed a substantial negative impact on the activation program, coordinately preventing the expression of several gene categories related to the immune response, including all cytokines. Measurements of IL-2 production showed a significant defect in knockout cells, which was rescued upon reintroduction of HuR. Interestingly, the mechanism of HuR regulation did not involve control of the cytokine 3′ UTRs: HuR knockout did not affect the activity of several cytokine 3′ UTR reporters in 293 cells. Similarly, HuR had no effect on the regulation of IL-2 and TNF 3′ UTRs in resting or activated Jurkat cells. Instead, impaired cytokine production corresponded with defective induction of the IL-2 promoter upon activation. Accordingly, upregulation of the master transcription factor NFATC1 upon activation was also impaired in HuR KOs, without effects on its 3′ UTR. Together, these results indicate that HuR controls cytokine production through coordinated upstream pathways, and that additional mechanisms must be considered in investigating its function.

overall_design: RNA-seq of mRNA in WT and HuR KO Jurkat cells


title: Transcriptome analysis of circulating monocytes in QFS and CFS compared to various control groups

status: Public on May 06 2019

summary: Transcriptomes of circulating monocytes in Q fever fatigue syndrome (QFS) patients, chronic fatigue syndrome (CFS) patients, asymptomatic Q fever seropositive controls and healthy controls

overall_design: Circulating monocytes from QFS patinets, CFS patients, asymptomatic Q fever seropositive controls and healthy controls were isolated from PBMCs by menas of Percoll

first: Ruud

last: Raijmakers

first: Mihai

last: Netea

first: Chantal

last: Bleeker-Rovers


title: Tuberculous Meningitis in Pediatric Patients

status: Public on Aug 06 2019

first: Rachel

last: Lai

summary: Whole blood transcriptional profiles in children with or without tuberculous meningitis (TBM) were compared using RNA-Seq and a biomarker signature driven by inflammasome activation and signaling was identified

overall_design: A total of 43 whole blood samples were sequenced, which include 15 TBM cases, 4 non-TBM infection cases and 24 non-infection healthy controls

first: Ursula

last: Rohlwink


title: Functional rare and low frequency variants in BLK and BANK contribute to human lupus pathogenesis

status: Public on May 01 2019

first: Nicole

last: Baldwin

address: 3434 Live Oak St

summary: Systemic lupus erythematosus (SLE) is the prototypic systemic autoimmune disease. It is thought that many common variant gene loci of weak effect act additively to predispose to common autoimmune diseases, while the contribution of rare variants remains unclear. Here we describe that rare coding variants in lupus-risk genes are present in most SLE patients and healthy controls. We demonstrate the functional consequences of rare and low frequency missense variants in the interacting proteins BLK and BANK, which are present alone, or in combination, in a substantial proportion of lupus patients. The rare variant found in patients, but not those found exclusively in controls, impair suppression of IRF and type-I IFN in human B cell lines and increase pathogenic lymphocytes in lupus-prone mice. Thus, rare gene variants are common in SLE and likely contribute to genetic risk.

overall_design: 31 SLE Samples, 9 controls

first: Simon

last: Jiang

first: Vicki

last: Athanasopoulos

first: Julia

middle: I

last: Ellyard

first: Aaron

last: Chuah

first: Jean

last: Cappello

first: Amelia

last: Cook

first: Savit

last: Prabhu

first: Jacob

last: Cardenas

first: Jinghua

last: Gu

first: Maurice

last: Stanley

first: Jonathan

last: Roco

first: Ilenia

last: Papa

first: Mehmet

last: Yabas

first: Giles

last: Walters

first: Gaetan

last: Burgio

first: Kathryn

last: McKeon

first: James

last: Byers

first: Charlotte

last: Burrin

first: Anselm

last: Enders

first: Lisa

last: Miosge

first: Pablo

middle: F

last: Canete

first: Marija

last: Jelusic

first: Velibor

last: Tasic

first: Adrian

last: Lungu

first: Stephen

last: Alexander

first: Arthur

last: Kitching

first: David

last: Fulcher

last: Shen

first: Todor

last: Arsov

first: Paul

last: Gatenby

first: Jeff

last: Babon

first: Dominic

last: Mallon

first: Carmen

last: de Lucas Collantes

first: Eric

last: Stone

first: Philip

last: Wu

first: Matthew

last: Field

first: Thomas

last: Andrews

first: Eun

last: Cho

first: Virginia

last: Pascual

first: Carola


title: Blood immunomonitoring uncovers pregnancy-induced immune programs and their dysregulation in lupus pregnancy destined for complications

status: Public on Mar 22 2019

first: Nicole

last: Baldwin

summary: Systemic lupus erythematosus carries increased risk of pregnancy complications.  To understand the underlying molecular mechanisms, we characterized the blood transcriptome of lupus patients and healthy controls during pregnancy and postpartum and performed multicolor flow-cytometry in a subset. We also followed healthy women undergoing assisted reproductive technology, allowing for analysis around embryo implantation. A striking and sustained down modulation of two lupus-related signatures, interferon and plasma cells, takes place during healthy pregnancy. These changes appear immediately post embryo implantation and are mirrored in uncomplicated lupus pregnancies. Patients with preeclampsia displayed early upregulation of neutrophil signatures and expansion of immature neutrophils. Lupus pregnancies with fetal complications carried the highest interferon and plasma cell signatures as well as ICOS+ CD4+ T cell counts. Thus, healthy and uncomplicated lupus pregnancies exhibit early and sustained transcriptional modulation of lupus-related signatures, and failure to do so is associated with adverse outcomes.

overall_design:
512 total samples; 325 SLE samples; 187 without SLE. Samples were taken under the following conditions: not pregnant or NP, <16 wks pregnant, 16-23 weeks pregnant, 24-31 weeks pregnant, 32-40 weeks pregnant, post-partum or PP.

Description of sample characteristics:
GRP_P_TP; Disease group, Pregnancy/non-pregnancy, timepoint, a variable used for mixed model
GA_AT_COLLECTION; Gestational age at collection (week)
GA_at_end_of_Pregnancy; Gestational age at the end of pregnancy (week)
SLE; yes=1, no=0
aPL; Antiphospholipid antibodies yes=1, no=0
LAC; Lupus Anticoagulant yes=1, no=0
TP; time point [1: <16 weeks of gestation (WG); 2: 16-23 WG; 3: 24-31 WG; 4: 32-40 WG, and between 8 and 20 weeks postpartum (5)]
PE; Preeclampsia yes=1, no=0
If_PE_before_or_after_36_weeks; If the patient had preeclampsia, before or after 36 WG
FD; Fetal death
NND; Neonatal death
PL_Insuff; preterm delivery <36 weeks because of placental insufficiency
IUGR; Intrauterine growth restriction
SGA; small-for-gestational-age neonate

first: Seunghee

last: Hong

first: Romain

last: Banchereau

first: Bat-Sheva

last: Maslow

first: Marta

last: Guerra

first: Jeanine

last: Baisch

first: D

middle: W

last: Branch

first: Flint

last: Porter

first: Allen

last: Sawitzke

first: Carl

last: Laskin

first: Jill

last: Buyon

first: Michael

middle: D

last: Lockshin

first: Lisa

last: Sammaritano

first: Michelle

last: Petri

first: Alma-Martina

last: Cepika

first: Marina

last: Ohouo

first: Gerlinde

last: Obermoser

first: Esperanza

last: Anguiano

first: Jacob

last: Turner

first: Derek

last: Blankenship

first: Tae

last: Kim

first: John

last: Nulsen

first: Djamel

last: Nehar-Belaid

first: Jacques

first: Jane

last: Salmon


title: Molecular Distance to Health Transcriptional Score and Disease Severity in Children Hospitalized with Community-Acquired Pneumonia

status: Public on Dec 26 2018

first: Nicole

last: Baldwin

summary: Community-acquired pneumonia (CAP) is a leading cause of hospitalization and mortality in children.  Diagnosis remains challenging and there are no reliable tools to objectively classify patients according to disease severity or predict clinical outcomes. Molecular distance to health (MDTH) is a genomic score that measures the global perturbation of the transcriptional profile that may help classify patients by disease severity. We assessed the value of MDTH to assess disease severity in children hospitalized with CAP.

overall_design: 172 total samples (152 CAP samples, 20 matched healthy control samples)

first: Rebecca

last: Wallihan

first: Nicolas

last: Suarez

first: Daniel

last: Cohen

first: Mario

last: Marcon

first: Melissa

last: Moore-Clingenpeel

first: Asuncion

last: Mejias

first: Octavio

last: Ramilo


title: Steroid Receptor Coactivator-2 Regulated Transcriptome in Human Endometrial Stromal Cells

status: Public on Oct 23 2018

first: Maria

last: Szwarc

summary: RNA-sequencing of mRNA isolated from in vitro decidualizaing human endometrial stromal cells with or without siRNA-mediated knockdown of steroid receptor coactivator-2/nuclear receptor coactivator 2 (SRC-2/NCOA2)

overall_design: Primary human endometrial stromal cells isolated from 3 healthy volunteers. Transfected with nontargeting or SRC-2/NCOA2 siRNA. Treated with estradiol, medroxyprogesterone acetate, and cAMP (EPC) for 0 or 3 days

last: Lydon

middle: M


title: EGR1-controlled transcriptome of T HESCs

status: Public on Apr 22 2019

summary: Sequencing of mRNA isolated from a human endometrial stromal cell line (T HESCs) following control or EGR1 siRNA-mediated knockdown.

overall_design: RNA isolated from THESCs following 48h transfection with nontargeting or EGR1 siRNA. Triplicate samples for each siRNA are derived 3 from independent experiments.


title: SRC-2/NCOA genomic binding in decidualizing human endometrial stromal cells

status: Public on Dec 04 2018

summary: Whole genome binding profile of SRC-2/NCOA2 in human endometrial stromal cells

overall_design: ChIP-sequencing of SRC-2/NCOA2 in decidualizing human endometrial stromal cells

first: Francesco

middle: J

last: DeMayo


title: Parkinson-associated SNCA enhancer variants revealed by open chromatin in mouse dopamine neurons

status: Public on Nov 14 2018

city: Baltimore

first: Andy

last: McCallion

state: MD

address: 733 N Broadway, MRB 446

department: Institute of Genetic Medicine

institute: Johns Hopkins School of Medicine

summary: The progressive loss of midbrain (MB) dopaminergic (DA) neurons defines the motor features of Parkinson disease (PD) and modulation of risk by common variation in PD has been well established through GWAS. Anticipating that a fraction of PD-associated genetic variation mediates their effects within this neuronal population, we acquired open chromatin signatures of purified embryonic mouse MB DA neurons. Correlation with >2,300 putative enhancers assayed in mice reveals enrichment for MB cis-regulatory elements (CRE), data reinforced by transgenic analyses of six additional sequences in zebrafish and mice. One CRE, within intron 4 of the familial PD gene SNCA, directs reporter expression in catecholaminergic neurons of transgenic mice and zebrafish. Sequencing of this CRE in 986 PD patients and 992 controls reveals two common variants associated with elevated PD risk. To assess potential mechanisms of action, we identify proteins whose binding is impacted by these enhancer variants. Additional genotyping across the SNCA locus identifies a single PD-associated haplotype, containing the minor alleles of both of the aforementioned PD-risk variants. Our work posits a model for how common variation at SNCA may modulate PD risk and highlights the value of cell context-dependent guided searches for functional non-coding variation.

overall_design: 6 ATAC-seq libraries and 8 RNA-seq samples from sorted mouse Th-EGFP dopaminergic neurons collected at E15.5 from two distinct brain regions (midbrain and forebrain)

first: Sarah

last: McClymont

first: Paul

last: Hook

first: Alexandra

last: Soto

first: Xylena

last: Reed

first: William

last: Law

first: Samuel

last: Kerans

first: Eric

middle: L

last: Waite

last: Briceno

first: Joey

last: Thole

first: Michael

last: Heckman

first: Nancy

last: Diehl

first: Zbigniew

middle: K

last: Wszolek

first: Cedric

last: Moore

first: Heng

last: Zhu

first: Jennifer

last: Akiyama

first: Diane

middle: E

last: Dickel

first: Axel

last: Visel

first: Len

last: Pennacchio

first: Owen

last: Ross

last: Beer

first: Andrew

middle: S


title: Tuberculous Meningitis in Pediatric Patients [CSF]

first: Rachel

last: Lai

summary: Cerebrospinal fluid transcriptional profiles in children with tuberculous meningitis (TBM) or other infections were compared using RNA-Seq and a biomarker signature driven by NMDA-receptor activation was identified

overall_design: A total of 31 CSF samples were sequenced, which include 18 TBM cases (ventricular and lumbar puncture CSF) and 7 other infections (ventricular CSF only)

first: Ursula

last: Rohlwink


title: Cancer stem cells associated miRNAs serve as prognostic biomarkers in colorectal cancer

status: Public on Feb 22 2019

first: Nicole

last: Baldwin

institute: Baylor Research Institute

type: Non-coding RNA profiling by high throughput sequencing

summary: Chemoresistance in cancer is linked to a subset of cancer cells termed “cancer stem cells” (CSCs), and in particular, those expressing the CD44 variant appear to represent a more aggressive disease phenotype. Herein, we demonstrate that CD44v6 represents a CSC population with increased resistance to chemotherapeutic agents, and its high expression is frequently associated with poor overall survival (OS) (HR:3.04; CI:1.35–6.85; p<0.001) and disease-free survival (DFS) (HR:5.30, CI:1.64–17.12, p<0.01) in CRC patients. CD44v6+ cells showed elevated resistance to chemotherapeutic drugs and significantly high tumor initiation capacity. The inhibition of CD44v6 resulted in the attenuation of self-renewal capacity and re-sensitization to chemotherapeutic agents. Of note, miRNA profiling of CD44v6+ SDCSCs identified a unique panel of miRNAs indicative of high self-renewal capacity. In particular, miR-1246 was overexpressed in CD44v6+ cells, and associated with poor OS (HR:2.44, CI:1.15–5.18, p<0.05) and DFS (HR:2.37, CI:1.03–5.44, p<0.05) in CRC patients. We demonstrate that CD44v6+ CSCs induced chemoresistance and enhance tumorigenicity in CRC cells, which is in part orchestrated by dysregulated profiles of a distinct panel of miRNAs. These findings provide a rationale for the therapeutic targeting of specific miRNAs, as well as serving as promising prognostic biomarkers in patients with colorectal neoplasia.

overall_design: 8 total samples, 4 CD44v6− HCT116 replicates and 4 CD44v6+ HCT116 replicates

first: Shusuke

last: Toden

first: Shigeyasu

last: Kunitoshi

first: Jacob

last: Cardenas

first: Jinghua

last: Gu

first: Elizabeth

last: Hutchins

first: Kendall

last: Van Keuren-Jensen

first: Hiroyuki

last: Uetake

first: Yuji

last: Toiyama

first: Ajay

last: Goel


title: Combinatorial mRNA Regulation by miRNAs and Pumilio proteins

status: Public on Oct 17 2018

first: Fedor

last: Karginov

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

overall_design: Refer to individual Series


title: A novel CD4+ T cell population expanded in SLE blood provides B cell help through IL10 and succinate

status: Public on Sep 26 2018

summary: A better understanding of the mechanisms involved in human plasma cell differentiation will accelerate therapeutic target identification in autoantibody-mediated diseases such as Systemic Lupus Erythematosus (SLE). Here, we describe a novel CXCR5- CXCR3+ PD1hi CD4+ T cell ‘helper’ population distinct from follicular helper T cells (Tfh) and expanded in blood and inflamed kidneys of SLE patients. Upon activation, these cells express IFNand high levels of IL10. Additionally, they accumulate high amounts of mitochondrial ROS (mtROS) as the result of reverse electron transport (RET) fueled by succinate.  These cells provide potent help to B cells through the synergistic effect of IL10 and succinate. Cells with similar phenotype and function are generated in vitro upon priming naïve CD4+ T cells with oxidized mitochondrial DNA (Ox mtDNA)-activated plasmacytoid dendritic cells (pDCs) in a PD1-dependent manner. Our results provide a novel mechanism for the initiation and/or perpetuation of extrafollicular humoral responses in SLE.

overall_design: 9 total samples; 3 groups of 3 biological replicates: control group Th0, co-culture group CpGA-pDC, and co-culture group Ox mtDNA-pDC

first: Simone

last: Caielli

first: Diogo

middle: T

last: Veiga

first: Preetha

last: Balasubramanian

first: Shruti

last: Athale

first: Bojana

last: Domic

first: Elise

last: Murat

first: Romain

last: Banchereau

first: Zhaohui

last: Xu

first: Manjari

last: Chandra

first: Chang

last: Chung

first: Lynnette

last: Walters

first: Jeanine

last: Baisch

first: Tracey

last: Wright

first: Marilynn

last: Punaro

first: Lorien

last: Nassi

first: Katie

last: Stewart

first: Julie

last: Fuller

first: Duygu

last: Ucar

first: Hideki

last: Ueno

first: Joseph

last: Zhou

first: Jacques

first: Virginia

last: Pascual


title: A novel CD4+ T cell population expanded in Systemic Lupus Erythematosus (SLE) blood provides B cell help through IL10 and succinate

first: Nicole

last: Baldwin

summary: A better understanding of the molecular and cellular factors involved in human plasma cell differentiation will accelerate therapeutic target identification in autoantibody-mediated diseases such as Systemic Lupus Erythematosus (SLE). Here, we describe a novel CXCR3+ PD1hi CD4+ T cell ‘helper’ population expanded in blood and inflamed kidneys of SLE patients. Upon activation, these cells express IFNg and IL10 and display high levels of mitochondrial ROS (mtROS) as the result of reverse electron transfer (RET) fueled by succinate. Furthermore, T cell-derived succinate synergizes with IL10 to provide B cell help. Cells with similar phenotype and function are generated in vitro upon priming naive CD4+ T cells with oxidized mitochondrial DNA (Ox mtDNA)- activated plasmacytoid dendritic cells (pDCs) in a PD1-dependent manner. Our results provide a novel mechanism for the initiation and/or perpetuation of extrafollicular humoral responses in SLE.

overall_design: 2 independent datasets; dataset1: total 9 samples (3 subjects, 3 groups, 3 replicates); dataset2: total 20 samples, 2 samples(PD1POS, Tfh) from each of 10 SLE patients

first: Simone

last: Caielli

first: Diogo

last: Veiga

first: Bojana

last: Domic

first: Elise

first: Shruti

last: Athale


title: Interleukin 10 and succinate synergize to provide B cell help in human lupus

summary: Autoantibodies against nucleic acids are a hallmark of Systemic Lupus Erythematosus.  We recently uncovered that human oxidized DNA of mitochondrial origin released by activated lupus neutrophils represents a distinct class of interferogenic TLR9 ligand for plasmacytoid dendritic cells. We now show that oxidized mitochondrial DNA-activated plasmacytoid dendritic cells skew naïve CD4+ T cells towards IL2low, IFNγhigh, IL10high secreting B helper cells different from follicular helper and Type 1 regulatory CD4+ T cells. Furthermore, PD1-induced succinate and mitochondrial ROS accumulation revoke anergy, while IL10 and succinate synergize to deliver B cell help. We provide evidence that IL10-producing CD4+ T cells infiltrate the SLE kidney insterstitium, where they might play a role in extrafollicular B cell responses. Thus, we describe a novel B cell helper pathway that links innate and adaptive immunity alterations in human lupus.

overall_design: 9 total samples. 3 biological replicates of each of 3 conditions. Stimulation with media alone (MEDIA) as a control, stimulation with oxidated mitochondrial DNA (Ox mtDNA), stimulation with a Class-A CpG ODN that preferentially binds to human TLR9 (CpGA).

first: Elis

last: Murat

first: Manjari

last: Chandra

first: Lynnette

first: Romain

last: Banchereau

first: Zhaohui

last: Xu

first: Jeanine


title: Combinatorial mRNA Regulation by miRNAs and Pumilio proteins [PUM1CLIP]

first: Fedor

last: Karginov

summary: Approximately 1500 RNA-binding proteins (RBPs) profoundly impact mammalian cellular function by controlling distinct sets of transcripts, often using sequence-specific binding to 3′ untranslated regions (UTRs) to regulate mRNA stability and translation. Aside from their individual effects, higher-order combinatorial interactions between RBPs on specific mRNAs have been proposed to underpin the regulatory network. To assess the extent of such co-regulatory control, we took a global experimental approach followed by targeted validation to examine interactions between two well-characterized and highly conserved RBPs, Argonaute2 (AGO2) and Pumilio (PUM1 and PUM2). Transcriptome-wide changes in AGO2-mRNA binding upon PUM knockdown were quantified by CLIP-seq, and the presence of PUM binding on the same 3′ UTR corresponded with cooperative and antagonistic effects on AGO2 occupancy. In addition, PUM binding sites that overlap with AGO2 showed differential, weakened binding profiles upon abrogation of AGO2 association, indicative of cooperative interactions. In luciferase reporter validation of candidate 3′ UTR sites where AGO2 and PUM colocalized, three sites were identified to host antagonistic interactions, where PUM counteracts miRNA-guided repression. Interestingly, the binding sites for the two proteins are too far for potential antagonism due to steric hindrance, suggesting an alternate mechanism. Our data experimentally confirms the combinatorial regulatory model and indicates that the mostly repressive PUM proteins can change their behavior in a context-dependent manner. Overall, the approach underscores the importance of further elucidation of complex interactions between RBPs and their transcriptome-wide extent.

overall_design: Determination of PUM1 and PUM2 binding by CLIP-seq in 293T cells and DICER-null derivatives

first: Erin

last: Sternburg

first: Jason

middle: A

last: Estep

first: Yahui

last: Li

middle: V


title: Transcriptome Mapping of Internal N7-methylguanosine Methylome in Mammalian mRNA

status: Public on Apr 26 2019

first: Li-Sheng

last: Zhang

state: IL

address: 929 E 57th St Rm E313

department: Department of Chemistry

institute: The University of Chicago

type: Methylation profiling by high throughput sequencing

summary: N7-methylguanosine (m7G) is a positively charged, essential modification at the 5′ cap of eukaryotic mRNA, regulating mRNA export, translation, and splicing. m7G also occurs internally within tRNA and rRNA, but its existence and distribution within eukaryotic mRNA remain to be investigated. Here, we show the presence of internal m7G sites within mammalian mRNA. We then performed transcriptome-wide profiling of internal m7G methylome using m7G-MeRIP sequencing (MeRIP-seq). To map this modification at base resolution, we developed a chemical-assisted sequencing approach that selectively converts internal m7G sites into abasic sites, inducing misincorporation at these sites during reverse transcription. This base-resolution m7G-seq enabled transcriptome-wide mapping of m7G in human tRNA and mRNA, revealing distribution features of the internal m7G methylome in human cells. We also identified METTL1 as a methyltransferase that installs a subset of m7G within mRNA and showed that internal m7G methylation could affect mRNA translation.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design:
[dataset 1] 12 samples. Duplicates for PAR-CLIP of METTL1 and WDR4 in HeLa cell line. Duplicates for PAR-CLIP of eIF4E, RNMT, METTL1 and WDR4 in HEK293T cell line.
[dataset 2] 8 samples. Duplicates for Ribo-Seq samples, mRNA input and ribosome-bound RNA of siControl vs siMETTL1 in HeLa cell.
[dataset 3] 46 samples. Duplicates for m7G-seq of small RNA (<200 nt) and mRNA samples, including three sections of RNA input, before_pulldown, and pulldown in HeLa, HepG2, and HEK293T cell lines.
[dataset 4] 38 samples. Duplicates for m7G-MeRIP-seq (anti-m7G antibody, MBL) of cell mRNA samples, including two sections of RNA input and IP in HeLa, HepG2, HEK293T, MEF and mESC wild-type cell lines; also in HeLa cell siControl vs siMETTL1, and HepG2 cell shControl vs shMETTL1.

first: Chang

last: Liu

first: Chuan

last: He


title: Gene expression profiling of the whole mouse brain at 7 days post-natal age using transcriptome tomography

status: Public on Aug 07 2018

city: Wako-shi

first: Yuko

last: Okamura-Oho

state: Saitama

address: 2-1, Hirosawa

country: Japan

institute: RIKEN

summary: We developed a novel framework to assess comprehensive endogenous gene expression maps in a broad and isotropic 3D-anatomical context using a relatively small number of original data points: transcriptome tomography. Using this method, we produced a dataset of gene expression maps of the whole mouse brain at 7 days after birth, computationally reconstructed from fraction data.

overall_design: A total of 22 fraction samples obtained with body axes-based slicing preparation conjugated with anatomical image acquisition were analysed. There are a sets of body axes-based preparation series: the S series composed of 6 fractions/ C of 10 fractions/ H of 6 fractions.

first: Y

first: H

last: Yokota


title: Respiratory Syncytial Virus Genotypes and Disease Severity in Young Children Hospitalized with Bronchiolitis

status: Public on Mar 21 2018

first: Nicole

last: Baldwin

summary:
Background: There is limited data on how different RSV genotypes and associated viral loads influence disease phenotypes. We characterized the genetic variability of RSV strains during five non-consecutive respiratory seasons, and evaluated the role of RSV subtypes, genotypes and viral loads on clinical disease severity.
Methods: Healthy infants hospitalized with RSV bronchiolitis were prospectively enrolled and nasopharyngeal samples obtained within 24h of hospitalization for RSV load quantitation by PCR, typing and genotyping. Parameters of disease severity were assessed, and multivariate models constructed to identify virologic and clinical factors predictive of clinical outcomes.
Results: From March 2004 to April 2011, we enrolled 253 patients (56.5 % males; median age 2.1 (1.1-4.0) months). RSV A infections predominated over RSV B (69% vs. 31%; p<0.001) and showed greater genotype variability. The most common genotypes were RSV A/GA2, A/GA5 and RSV B/BA. Infants infected with RSV GA5 had higher viral loads compared with GA2 or BA infection (p<0.01), independent of duration of symptoms. After adjusting for other covariates, RSV A/GA5 infections were associated with longer hospital stay.
Conclusions: RSV A infections were more frequent than RSV B infections and displayed greater genetic variability. Infections with GA5 were independently associated with clinical disease severity.

overall_design: To define the host systemic immune response according to the different RSV types or genotypes identified, blood transcriptional profiles were analyzed in a subset of infants with RSV bronchiolitis enrolled during the last study season (2010-11; n=62) and 12 healthy asymptomatic age-matched controls.  Of the 62 infants with RSV LRTI enrolled, 49 (79%) were infected with RSV A (GA2 n=43, GA5 n=5, non-typed n=1) and 13 (21%) with RSV B (BA n=12, non-typed n=1).

first: Rosa

last: Rodriguez-Fernandez

first: Lorena

last: Tapia

first: Chin-Fen

last: Yang

first: Juan

middle: P

last: Torres

first: Susana

last: Chavez-Bueno

first: Carla

last: Garcia

first: Lisa

last: Jaramillo

first: Melissa

last: Moore-Clingenpeel

first: Hasan

last: Jafri

first: Pedro

last: Piedra

first: Mark

last: Peeples

first: Octavio

last: Ramilo

first: Asuncion

last: Mejias


title: Comprehensive gene expression analysis of mouse brains in postnatal maturation stages using Transcriptome Tomography

first: Yuko


title: RNA-Sequencing approach for the identification of novel long non-coding RNA biomarkers in colorectal cancer

status: Public on Feb 14 2018

summary: Long non-coding RNA (lncRNA) have been implicated in human pathology, however, their roles in colorectal carcinogenesis has not been fully elucidated. In the current study, whole-transcriptome was analyzed in 3 pairs of colorectal cancer (CRC) and matched normal mucosa (NM) by RNA sequencing (RNA-seq). Followed by confirmation using the Cancer Genome Atlas (TCGA) dataset, we identified 27 up-regulated and 22 down-regulated lncRNAs in CRC. Up-regulation of four lncRNAs, hereby named colorectal cancer associated lncRNA (CRCAL)-1 [AC021218.2], CRCAL-2 [LINC00858], CRCAL-3 [RP11-138J23.1] and CRCAL-4 [RP11-435O5.2], was further validated by real-time RT-PCR in 139 colorectal neoplasms and matched NM tissues. Knockdown of CRCAL-3 and CRCAL-4 in colon cancer cells reduced cell viability and colony formation ability, and induced cell cycle arrest. TCGA dataset supported the associations of CRCAL-3 and CRCAL-4 with cell cycle and revealed a co-expression network comprising dysregulated lncRNAs associated with protein-coding genes. In conclusion, RNA-seq identified numbers of novel lncRNAs dysregulated in CRC. In vitro experiments and GO term enrichment analysis indicated the functional relevance of CRCAL-3 and CRCAL-4 in association with cell cycle. Our data highlight the capability of RNA-seq to discover novel lncRNAs involved in human carcinogenesis, which may serve as alternative biomarkers and/or molecular treatment targets.

overall_design: 6 total samples consisting of 3 matched pairs of colorectal cancer (CRC) and matched normal mucosa (NM)

first: Atsushi

last: Yamada

first: Pingjian

last: Yu

first: Wei

last: Lin

first: Yoshinaga

last: Okugawa

first: C

middle: R

last: Boland

first: Ajay

last: Goel


title: Combinatorial mRNA Regulation by miRNAs and Pumilio proteins [CLIP-seq]

first: Fedor

last: Karginov

overall_design: Determination of AGO2 binding by CLIP-seq in T-REx-293 cells with and without PUM1 and PUM2 knockdown

first: Erin

last: Sternburg

first: Jason

last: Estep

first: Yahui

last: Li


title: The Promyelocytic Leukemia Zinc Finger Dependent Transcriptome during Human Endometrial Stromal Cell Decidualization

status: Public on Mar 29 2018

first: Maria

last: Szwarc

summary: RNA-sequencing of mRNA isolated from in vitro decidualizaing human endometrial stromal cells with or without siRNA-mediated knockdown of PLZF

overall_design: Primary human endometrial stromal cells isolated from 3 healthy volunteers. Transfected with nontargeting or PLZF siRNA. Treated with estradiol, medroxyprogesterone acetate, and cAMP (EPC) for 0 or 3 days

first: John

last: Lydon


title: Canakinumab treatment in hyperimmunoglobulinemia D with periodic fever syndrome

status: Public on Sep 17 2018

first: Nicole

last: Baldwin

summary:
Objective. To evaluate the efficacy and safety of canakinumab treatment in active hyperimmunoglobulinemia D with periodic fever syndrome (HIDS).
Methods. This is a 3-part open-label study with a 6-month treatment period (P1; 300 mg or 4 mg/kg q6w) a 6-month withdrawal period (P2) and a 24-month treatment period (P3). The primary endpoint was reduction in frequency of flares during treatment period compared with the historical period (HP; period in which patients did not receive drugs other than NSAIDs and/or steroids).

overall_design: 63 samples total. 7 at visit 1: anakinra treatment (V1_AK) 9 at visit 2: during active disease after anakinra withdrawal (V2_NT) 9 at visit 3: after anakinra withdrawal and on canakinumab (V3) 8 at visit 4: after anakinra withdrawal and on canakinumab (V4) 8 at visit 5: after anakinra withdrawal and on canakinumab (V5) 15 age- and sex-matched healthy controls (HC). Note: V2_NT is BL in the associated manuscript. 7 at visit 6: during active disease after canakinumab withdrawal (ATTACK)

first: Juan

last: Arostegui

first: Jordi

last: Anton

first: Inmaculada

last: Calvo

first: Angel

last: Robles

first: Estibaliz

last: Iglesias

first: Berta

last: López-Montesinos

first: Romain

last: Banchereau

first: Seunghee

last: Hong

first: Antonio

last: Speziale

first: Yolandi

last: Joubert

first: Guido

last: Junge

first: Virginia

last: Pascual

last: Yagüe


title: Mechanistic insights into anti-cancer properties of oligomeric proanthocyanidins from grape seeds in colorectal cancer

status: Public on Feb 28 2018

summary: Although the anti-cancer properties of Oligomeric Proanthocyanidins (OPCs) from grape seeds has been well recognized, the molecular mechanisms by which they exert anti-cancer effects are poorly understood. In this study, through comprehensive RNA-sequencing based gene-expression profiling in multiple colorectal cancer cell lines, we for the first time illuminate the genome-wide effects of OPCs from grape seeds in colorectal cancer. Our data revealed that OPCs affects several key cancer-associated genes. In particular, genes involved in cell cycle and DNA replication were most significantly and consistently altered by OPCs across multiple cell lines. Intriguingly, our in vivo experiments showed that OPCs was significantly more potent at decreasing xenograft tumor growth compared to the unfractionated grape seed extract (GSE) that includes the larger polymers of proanthocyanidins. These findings were further confirmed in colorectal cancer patient-derived organoids, wherein OPCs more potently inhibited the formation of organoids compared to GSE. Further, we validated alteration of cell cycle and DNA replication associated genes in cancer cell lines, mice xenografts as well as patient-derived organoids. Overall, this study provides an unbiased and comprehensive look at the mechanisms by which OPCs exerts anti-cancer properties in colorectal cancer.

overall_design: 30 total samples consisting of 6 samples for each of 5 cell lines; From each group of 6 samples 2 were treated with DMSO (control), 2 were treated with grape seed extract (GSE), and 2 were treated with oligomeric proanthocyanidins (OPCs)

first: Preethi

last: Ravindranathan

first: Divya

last: Pasham

first: Uthra

last: Balaji

first: Jacob

last: Cardenas

first: Jinghua

last: Gu

first: Shusuke

last: Toden

first: Ajay

last: Goel


title: Gene expression profiling of the whole mouse brain at 3 weeks of post-natal age using transcriptome tomography

first: Yuko

last: Okamura-Oho

summary: We developed a novel framework to assess comprehensive endogenous gene expression maps in a broad and isotropic 3D-anatomical context using a relatively small number of original data points: transcriptome tomography. Using this method, we produced a dataset of gene expression maps of the whole mouse brain at 3 weeks after birth, computationally reconstructed from fraction data.

overall_design: A total of 28 fraction samples obtained with body axes-based slicing preparation conjugated with anatomical image acquisition were analysed. There are a sets of body axes-based preparation series: the S series composed of10 fractions/ C of 12 fractions/ H of 6 fractions.

first: Y

first: H

last: Yokota


title: Transcriptional repression of IRF7 by MYC is critical for antiviral immune response in human pDC [pDC]

status: Public on Jun 01 2018

first: Nicole

last: Baldwin

summary: Type I interferons (IFN) are crucial mediators of human innate and adaptive immunity and are massively produced from plasmacytoid dendritic cells (pDC). IRF7 is a critical regulator of type I IFN production when pathogens are detected by TLR7/9 in pDC. However, hyperactivation of pDC can cause life-threatening autoimmune diseases. To avoid the deleterious effects of aberrant pDC activation, tight regulation of IRF7 is required. Nonetheless, the detailed mechanisms of how IRF7 transcription is regulated in pDC are still elusive. To this end, we identified the global gene expression changes after stimulation of human primary pDC with the TLR9 agonist CpGB. We identified that the transcription factor MYC is prominently upregulated upon CpGB engagement in pDC. Moreover, when we knocked down MYC in the pDC-like cell line GEN2.2, production of interferon-stimulated genes (ISGs) was dramatically increased and was further enhanced by CpGB.  Interestingly, MYC is shown to be recruited to the IRF7 promoter region through interaction with NCOR2/HDAC3 for its repression, and HDAC3 inhibition enhanced IRF7 expression and IFNβ production. Interestingly, activation of TLR9-mediated NF-kB and MAPK and nuclear translocation of IRF7 were greatly enhanced by MYC depletion. Pharmaceutical inhibition of MYC recovered IRF7 expression, further confirming the negative role of MYC in the antiviral response by pDC. Furthermore, the inverse correlation of MYC and IRF7 was validated in psoriasis skin sample datasets. Therefore, our results identify the novel immunomodulatory role of MYC in human pDC and may add to our understanding of aberrant pDC function in autoimmune diseases.

overall_design: 12 total pDC samples assayed at 4 time points post CpGB stimulation (3 replicates of 0hrs; 3 replicates of 4 hrs; 3 replicates of 8 hrs; 3 replicates of 12 hrs).

first: Tae

last: Kim

first: Yin

last: Lin

first: HyeMee

last: Joo

first: Taeil

first: Yong-Jun

last: Liu


title: RET Ligands Mediate Endocrine Sensitivity via a Bi-stable Feedback Loop with ERα

status: Public on Feb 22 2018

city: Ithaca

first: Charles

middle: G

last: Danko

state: NY

address: Hungerford Hill Rd

department: Baker Institute

institute: Cornell University

summary: The molecular mechanisms of endocrine resistance in breast cancer remain poorly understood. Here we used PRO-seq to map the location of hundreds of genes and thousands of distal enhancers whose activities differ between endocrine sensitive and resistant MCF-7 cells. Our genome-wide screen identified a 16-fold transcriptional increase in glial-cell line derived neurotrophic factor (GDNF), a RET tyrosine kinase receptor ligand, which is both necessary and sufficient for resistance in MCF-7 cells. GDNF causes endocrine resistance by switching the active state of a bi-stable feedback loop from ERα signaling to GDNF-RET signaling. To catalyze this switch, we found that GDNF directly downregulates ERα transcription and activates the transcription factor EGR1, which, in turn, induces GDNF expression. Remarkably, both MCF-7 cells and ER+ primary tumors appear poised for endocrine resistance via the RET signaling pathway, but lack robust RET ligand expression and only develop resistance upon expression of GDNF or other RET ligands.

overall_design: PRO-seq was used to analyze four different MCF-7 cell subclones in seven different biological conditions.


title: Gene expression profiling of the whole mouse brain at 3 days post-natal age using transcriptome tomography

summary: We developed a novel framework to assess comprehensive endogenous gene expression maps in a broad and isotropic 3D-anatomical context using a relatively small number of original data points: transcriptome tomography. Using this method, we produced a dataset of gene expression maps of the whole mouse brain at 3 days after birth, computationally reconstructed from fraction data.

overall_design: A total of 34 fraction samples obtained with body axes-based slicing preparation conjugated with anatomical image acquisition were analysed. There are a sets of body axes-based preparation series: the S series, composed of11 fractions/ C of 15 fractions/ H of 8 fractions.


title: Epigenomes and Transcriptomes of human monocytes from Hyper IgD Syndrome (HIDS) patients

status: Public on Jan 08 2018

first: Boris

last: Novakovic

address: Murdoch Children's Research Institute, Royal Children's Hospital, Flemington Road

institute: Murdoch Children's Research Institute

summary: Innate immune memory, also refered to as trained immunity (TRIM) is the phenomenon whereby innate immune cells such as monocytes or macrophages undergo functional reprogramming after exposure to microbial components.  In this study we performed epigenomic and transcriptomic analysis of HIDS (Hyper IgD Syndrome) patients at baseline, and after 24 hours culture in media (RPMI only) or media + 10ng/mL LPS. HIDS is associated with the accumulation of mevalonate, a monocyte trainer.

overall_design: HIDS patients were recruited and monocytes were isolated for ChIP-seq and RNA-seq.

first: Siroon

last: Bekkering

first: Rob

last: Arts

first: Mihai

last: Netea


title: Transcriptome-wide identification and validation of interactions between the miRNA machinery and HuR on mRNA targets

status: Public on Jan 19 2018

first: Fedor

last: Karginov

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

overall_design: Refer to individual Series


title: Transcriptome-wide identification and validation of interactions between the miRNA machinery and HuR on mRNA targets [HuR]

summary: The 3’ UTR of messenger RNAs serves as the regulatory region that mediates post-transcriptional control by microRNAs and RNA-binding proteins (RBPs). Aside from individual sequence-specific binding and regulation, examples of interaction between these factors at particular 3’ UTR sites have emerged in recent studies. However, the whole picture of such higher-order regulatory modules across the transcriptome is lacking. Here, we investigate the interactions between HuR, a ubiquitous RBP, and Ago2, a core effector of the miRNA pathway, at the transcriptome-wide level. Using HITS-CLIP, we map HuR and miRNA binding sites on human 3’UTRs and assess their co-occurrence. Additionally, we demonstrate global effects of HuR knockdown on Ago2 occupancy, suggesting a co-regulatory relationship. Focusing on sites of Ago2-HuR overlap, 13 candidates were screened in luciferase reporter assays, compared to miRNA site mutant controls. Eleven of the sites showed a repressive activity, which displayed significant de-repression upon subsequent testing of the reporters in Dicer-null cells, substantiating miRNA dependence. To experimentally test for HuR’s role in co-regulation, we tested the reporters in CRISPR-generated HuR KO cells. Three of the miRNA sites demonstrated altered activities, indicating that HuR has an effect on miRNA repression at those sites. Our study presents an efficient search and validation system for studying miRNA-HuR interactions, which expands our understanding of the combinatorial post-transcriptional control of gene expression at the 3’ UTR.

overall_design: Determination of HuR binding by CLIP-seq in 293T cells and 293T NoDice cells

first: Yahui

last: Li

first: Jason

last: Estep


title: Transcriptome-wide identification and validation of interactions between the miRNA machinery and HuR on mRNA targets [Ago2]

overall_design: Determination of Ago2 binding by CLIP-seq in T-REx-293 cells with and without HuR knockdown


title: Epigenomes and Transcriptomes of human macrophages after ex vivo exposure to mevalonate

summary: Innate immune memory, also refered to as trained immunity (TRIM) is the phenomenon whereby innate immune cells such as monocytes or macrophages undergo functional reprogramming after exposure to microbial components.  In this study  we exposed monocytes from healthy donors to mevalonate ex vivo, and analyzed the effect of this exposure on gene expression and histone modification changes.

overall_design: Monocytes from healthy donors were cultured in the presence of mevalonate and differentiated to macrophages to study mevalonate-induced trained innate immunity (TRIM). Cells were then collected for ChIP-seq and RNA-seq.


title: Transcriptomes of human monocytes after ex vivo exposure to uric acid

status: Public on Dec 08 2017

summary: Innate immune memory, also refered to as trained immunity (TRIM) is the phenomenon whereby innate immune cells such as monocytes or macrophages undergo functional reprogramming after exposure to microbial components.  In this study  we exposed monocytes from healthy donors to uric acid ex vivo, and analyzed the effect of this exposure on gene expression changes. After 'uric acid' or 'media only' treatments, monocytes were exposed to LPS for 4 hours to measure they response to this pyrogen.

overall_design: Monocytes from healthy donors were cultured in the presence of uric acid for 24 hours and then exposed to LPS for 4 hours. Cells were then collected for RNA-seq.

first: Tania

last: Crisan

first: Leo

last: Joosten


title: Self-organized cerebral organoids with human specific features predict effective drugs to combat Zika virus infection

status: Public on Oct 11 2017

first: Bennett

last: Novitch


title: Self-organized cerebral organoids with human specific features predict effective drugs to combat Zika virus infection [expression array]

summary: The human cerebral cortex possess distinct structural and functional features that are not found in the lower species traditionally used to model brain development and disease. Accordingly, considerable attention has been placed on the development of methods to direct pluripotency stem cells to form human brain-like structures termed organoids. However, many organoid differentiation protocols are inefficient and display marked variability in their ability to recapitulate the three-dimensional architecture and course of neurogenesis in the developing human brain. Here, we report optimized organoid culture methods that efficiently and reliably produce cortical and basal ganglia structures similar to those in the human fetal brain in vivo. Neurons within the organoids are functional and exhibit network-like activities. We further demonstrate the utility of the organoid system for modeling the teratogenic effects of Zika virus on the developing brain and identifying new candidate receptors and therapeutic compounds that can mitigate its desructive actions.

overall_design: Human cortical organoids were lysed in QIAzol reagents and RNA was extracted following manufacturer's instructions (miRNeasy Micro Kit, Qiagen). Cerebral organoids derived from H9 were collected with 3 replicates (3 independent experiments, 12 samples total) at W5, W8, W11, and W14 and RNA samples were sent to the UCLA Neuroscience Genomic Core. RNA integrity was cofirmed with the Agilent 2100 bioanalyzer (RIN > 8) and samples were hybridized to the HumanHT 12v4.0.

first: Momoko

last: Watanabe

first: Jessie

last: Buth

first: Neda

last: Vishlaghi

first: Luis

last: de la Torre-Ubeita

first: Jiannis

last: Taxidis

first: Baljit

last: Khakh

first: Giovanni

last: Coppola

first: Caroline

last: Pearson

first: Ken

last: Yamauchi

first: Danyang

last: Gong

first: Xinghong

last: Dai

first: Robert

last: Damoiseaux

first: Roghiyh

last: Aliyari

first: Simone

last: Liebscher

first: Katja

last: Schenke-Layland

first: Daniel

middle: H

last: Geschwind

first: Genhong

last: Cheng

first: Peyman

last: Golshani

first: Ren

last: Sun


title: Epigenomes and transcriptomes of human monocytes before and after in vivo exposure to Bacillus Calmette-Guérin vaccine

first: Boris

last: Novakovic

summary: Innate immune memory is the phenomenon whereby innate immune cells such as monocytes or macrophages undergo functional reprogramming after exposure to microbial components. In this study, we apply epigenomic and transcriptomic analysis to a clinical trial of BCG vaccination in healthy adults.

overall_design: Healthy volunteers were injected with the BCG vaccine, and monocytes were collected before vaccination, and 1 month after vaccination.

first: Rob

last: Arts

last: Moorlag

first: Mihai

last: Netea


title: Self-organized cerebral organoids with human specific features predict effective drugs to combat Zika virus infection [RNA-seq]

first: Bennett

last: Novitch

overall_design: Human cerebral organoids were differenitated from H9. For ZIKV analyses, ~ 8W-old cortical organoids with mock or ZIKV infection (~MOI 1.67) after 3 days and 5 days of post infection were collected with 3 biological replicates (6 samples total). RNA samples were sent to the UCLA Clinical Microarray Core (CMC) and RNA integrity was confirmed with the Agilent 2100 bioanalyzer. The cDNA libraries were generated using the KAPA stranded mRNA-Seq kits (KAPA Biosystems) and sequenced by Illumina HiSeq 3000, yielding between 27 and 101 million reads per sample.

first: Momoko

last: Watanabe

first: Jessie

last: Buth

first: Neda

last: Vishlaghi

first: Luis

last: de la Torre-Ubeita

first: Jiannis

last: Taxidis

first: Baljit

last: Khakh

last: Coppola


title: Blood transcriptional signature of HIV-associated tuberculous meningitis

status: Public on Jun 10 2017

first: Rachel

last: Lai

address: Du Cane Road

summary: Patients with HIV-associated TB meningitis (TBM) are known to experience systemic hyperinflammation, clinically known as immune reconstitution inflammatory syndrome (IRIS), following the commencement of antiretroviral therapy (ART). No prognostic markers or biomarkers have been identified to date and little is known about the mechanism mediating the hyperinflammation. We recruited a prospective cohort of 33 patients with HIV-associated TBM, 16 of whom developed TBM-IRIS, and performed transcriptomic profiling. Transcriptomic signatures that distinguish patients who would eventually develop IRIS were identified and indicated neutrophil and inflammasome activation as being the predominant mediator of TBM-IRIS pathogenesis.

overall_design: Patients are classified either as TBM-IRIS and non-IRIS and longitudinal samples (TBM-diagnosis, 2wpTBRx and 2wpART) were anlayzed. A control group of 17 patients HIV-1 infected patients without TBM and not on ART were also included. There are a total of 116 samples (33 TBM at 3 timepoints and 17 control at 1 timepoint). For noramlization, all samples were baseline transformed to the median of all samples.

first: Suzaan

last: Marais

first: Robert

last: Wilkinson


title: A multidimensional blood stimulation assay reveals immune alterations underlying systemic juvenile idiopathic arthritis [RNA-Seq]

status: Public on Sep 19 2017

city: Dallas

first: Nicole

last: Baldwin

summary: Despite great scientific and technological advances, immune alterations predisposing to sporadic human inflammatory diseases remain mostly unknown. To fill this gap, we developed a strategy to evaluate blood leukocyte responses to innate stimuli, simultaneously at the transcriptional, cellular and secreted protein levels. The data were integrated using weighted gene co-expression network analysis. When applied to systemic juvenile idiopathic arthritis (sJIA), an autoinflammatory disease of unknown etiology, this approach identified gene sets associated with specific cytokine environments and activated leukocyte subsets. During the remission phase of disease and off treatment, sJIA patients displayed dysregulated responses to TLR4, TLR8 and TLR7 stimulation in blood. Isolated sJIA monocytes accumulated higher levels of intracellular IL-1β after stimulation, and underexpressed the IL-1 inhibitor AHR at baseline. In line with the recent demonstration that AHR downregulation in monocytes is linked to macrophage differentiation, we show that the differentiation of sJIA monocytes in vitro was skewed towards macrophages, away from dendritic cell phenotype. This might contribute to the increased incidence of macrophage activation syndrome in these patients. The integrated analysis of these high-dimensional data can thus help unravel underlying immune alterations predisposing to complex inflammatory diseases.

overall_design: 32 total samples, 4 control healthy in vitro, 6 control systemic JIA in vitro, 4 LPS healthy in vitro, 6 LPS systemic JIA in vitro, 5 healthy ex vivo, 7 systemic JIA ex vivo

first: Alma-Martina

last: Cepika

first: Romain

last: Banchereau

first: Elodie

last: Segura

first: Marina

last: Ohouo

first: Brandi

last: Cantarel

first: Kristina

last: Goller

first: Victoria

last: Cantrell

first: Emily

last: Ruchaud

first: Elizabeth

last: Gatewood

first: Phuong

last: Nguyen

first: Jinghua

last: Gu

first: Esperanza

last: Anguiano

first: Sandra

last: Zurawski

first: Jeanine

last: Baisch

first: Marilynn

last: Punaro

first: Gerlinde

last: Obermoser

first: Karolina

last: Palucka

first: Jacques

first: Sebastian

last: Amigorena

first: Virginia

last: Pascual


title: Differential gene expression profile correlates with bortezomib responsiveness in steroid-resistant cGVHD

status: Public on Jan 18 2017

first: Nicole

last: Baldwin

summary: Nineteen patients with steroid-refractory cGVHD (chronic graft versus host disease) received weekly bortezomib from 2 – 12 months (1.6 mg/m2 q wk x 4 followed by one week of rest). Treatment was well tolerated with no > grade 3 adverse events. Six patients achieved CR or good PR (gPR). Average Rodnan scores decreased from 22.6 ± 12.8 to 5.9 ± 6.2 (p<0.005) for 8 patients with sclerosis who completed > 2 treatments, including one patient with marked healing of suppurating lesions and significantly reduced chronic diarrhea in another. Thus, weekly bortezomib was safe and produced early improvements in some patients with longstanding refractory disease. CR or gPR correlated with a younger age and lower number of involved organs with a severity score of > 2. Based on gene expression array and pathway analyses, High Responders displayed an immune response profile that resembled that of hematopoietic stem cell-transplanted patients without cGVHD. By comparison, Low Responders showed markedly altered gene expressions in both cell-mediated immune response and proinflammatory pathways. Our preliminary findings suggest that bortezomib responsiveness may be predicated by distinct immunopathological manifestations among cGVHD patients. This trial was registered at www.clinicaltrials.gov as NCT01158105.

overall_design:
54 total samples, 0 replicates, 8 healthy controls, 6 disease controls. Sample details follow.

Eligible patients were 18 years or older with a confirmed clinical diagnosis of steroid-refractory cGVHD, defined as either failure to improve after 2 months or progression after 1 month of standard steroid-based therapy, and had no prior bortezomib treatment for cGVHD. Enrolled patients were required to be in remission from his/her primary malignancy for which HSCT was indicated, ECOG performance status < 3, life expectancy > 3 months, have not had myocardial infarction within 6 months prior to enrollment and not having received radiation therapy within 3 weeks, platelet count ≥ 50 x 10^9/L, absolute neutrophil count ≥ 1.0 x 10^9/L, total bilirubin ≤ 1.5x upper normal limit, creatinine clearance ≥ 30 mL/min and no peripheral neuropathy episodes (≥ Grade 2) within 14 days before enrollment. Patients with prior hypersensitivity to bortezomib, boron or mannitol were excluded from trial.

Bortezomib was supplied in vials as open-label stock. Patients received Kytril premedication per institutional standards (1 mg intravenous/IV push) before start of VELCADE® (bortezomib; Millennium Pharmaceuticals) infusion (1.6 mg/m2). Bortezomib was administered on days 1, 8, 15, 22 of a 35-day cycle. Steroid dose that was in effect on Day 1 was maintained. All other agents given for GVHD treatment were withdrawn as tolerated. Patients with stable disease or better after two cycles continued for up to six cycles of bortezomib. Patients who continued to do well were eligible for up to six months of maintenance. Those patients continued on the same dose of bortezomib but on days 1 and 15 of a 28-day cycle. Followup after the final dose was for 6 months.

Subjects were assessed for signs and symptoms of cGVHD on day 1 of each cycle by standardized criteria for cGVHD diagnosis, new clinical scoring system, and new guidelines for global assessment of cGVHD severity (NIH Consensus Development Project on Criteria for Clinical Trials in cGVHD). Safety was assessed according to adverse event incidents for 30 days following the last dose for up to 6 months. Adverse events were recorded throughout the trial and followed by investigator until resolution. Enrolled patients also completed a neurotoxicity-directed questionnaire on presence and intensity of neuropathic pain and/or peripheral neuropathy from the patient's perspective. Responses were reviewed to assist with evaluation of onset and intensity of peripheral neuropathy and other neurotoxicities that may require intervention or dose modification.

No change from baseline was considered as stable disease (SD). >50% improvement in any specific organ score or overall performance was considered partial response (PR). Disappearance of symptoms of cGVHD was considered complete response (CR). Worsening of symptoms in specific organ or overall observed in two consecutive assessment time points was considered as progressive disease (PD). All human subject-related study protocols were reviewed and approved by the institutional Review Board for Human Protection, Baylor University Medical Center. All patients signed informed consent before being enrolled into study. This trial was registered at www.clinicaltrials.gov as NCT01158105.

first: Indira

last: Munagala

first: Allen

last: Miller

first: Reva

last: Schneider

first: Estil

last: Vance

first: Edward

last: Agura

first: Sheila

last: Powell

first: Aditi

last: Acharya

first: Mamta

last: Sharma

first: Huanyin

last: Qin

first: Derek

last: Blankenship

first: Alex

last: Tong


title: Healthy adult and sJIA blood stimulation with innate immune ligands

summary: The goal of this study was to characterize altered inducible immune networks in Systemic onset juvenile idiopathic arthritis (sJIA), an IL-1-driven autoinflammatory disease of unknown etiology. To this end, we developed a high-throughput assay that quantifies the transcriptional and protein-level responses of blood leukocytes to innate stimuli. Herein, we report transcriptional data from healthy adult blood stimulated with 16 different conditions, including TLR ligands, cytosolic receptor ligands and inflammatory cytokines. We further report blood transcriptional profiles from sJIA patients with various disease activity and treatment statuses, both ex vivo (baseline) and after in vitro stimulation with a subset of innate stimuli including heat-killed bacterial pathogens.

overall_design:
291 total samples, 31 donors and 17 stimuli (excluding media controls). All stimuli were not used with all donors. There is one technical replicate for donors BD1277, BD1350, BD1635 for all stimuli - the replicate was run in a later batch (larger barcode number).

Stimuli used were agonists for surface Toll-like receptors (CpG-C, R837,  R848, TLR8L, Flagellin, LPS), a protein kinase C activator (PMA/I: phorbol myristate acetate and ionomycin), cytokines (IFNalpha, IFNgamma, IL-17A/F,  IL-18, IL-1beta, TNFalpha), gram+ derived ligands (PGN, LTA), and  heat-killed Staph. aureus (HKSA) as well as heat-killed Staph enteric (HKSE).

Ligands were diluted in medium (RPMI 1640 with GlutaMax, Thermo Fisher  Scientific, Grand Island, NY, USA) and aliquoted into 2 mL polypropylene  96-well plates (Greiner Bio-One, Monroe, NC, USA). Blood was drawn in the  morning into the BD Vacutainers containing ACD anticoagulant (BD, Franklin  Lakes, NJ, USA). Time between blood draw and culture setup was less than  2h. Blood and appropriately diluted ligands were mixed in 1:1 ratio in  total volume of 1 mL/well and incubated for 6h at 37oC/5% CO2. After the culture, blood was mixed and  100 uL aliquot set aside for activation marker phenotyping and  intracellular cytokine staining (ICS) for IL-1b. The plate was then  centrifuged at 12,000 x g/15  and 400 mL of plasma aliquoted and stored at  -80oC until the Luminex analysis. The pellet was the  lysed with Tempus  solution (Tempus blood RNA tubes, Thermo Fisher Scientific) in 2:1 ratio  and stored at -20oC until the RNA extraction.

Please note that data were analyzed using either V1 manifest (pre-Oct 2010) or the V2 manifest (post-Oct 2010) as indicated in the sample title.

Sample Group abbreviation definitions:
sIU - SoJIA, Inactive, Untreated
sAU - SoJIA, Active, Untreated
sIU - SoJIA, Inactive, Anakinra

first: Alma-Martina

last: Cepika

first: Romain

last: Banchereau

first: Brandi

last: Cantarel

first: Marina

last: Ohouo

first: Kristina

last: Goller

first: Victoria

last: Cantrell

first: Emily

last: Ruchaud

first: Elizabeth

last: Gatewood

first: Phuong

last: Nguyen

first: Esperanza

last: Anguiano

first: Sandra

last: Zurawski

first: Jeanine

last: Baisch

first: Marilynn

last: Punaro

first: Gerlinde


title: A multidimensional blood stimulation assay reveals immune alterations underlying systemic juvenile idiopathic arthritis

status: Public on Sep 18 2017

first: Nicole

last: Baldwin

last: Obermoser

first: Jacques

first: Karolina

last: Palucka

first: Virginia

last: Pascual


title: Influenza Vaccines Differentially Regulate the Interferon Response in Human Dendritic Cell Subsets [HumanHT-12 V4.0 Beadchip Experiments]

status: Public on Dec 25 2016

summary: Human dendritic cells play a fundamental role in the initiation of long-term adaptive immunity during vaccination against influenza. Understanding the early response of human DCs to vaccine challenge is thus essential to determine the nature and magnitude of maturation signals that strongly correlate with vaccine effectiveness. Here, we show that the non-adjuvanted trivalent Fluzone®09-10 (TIV-09) induces monocyte-derived DCs (moDCs), purified blood conventional DCs (cDCs) and plasmacytoid DCs (pDCs) to express CD80, CD83, and CD86, and secrete cytokines. TIV-09 stimulated the secretion of type I interferons (IFN) IFN-α and IFN-β and type III IFN IL-29 by moDCs and cDCs subsets. The vaccine also induced the production of IL-6, TNF and the chemokines IP-10 and MIP-1β. Conversely, the non-adjuvanted monovalent H1N1 California vaccine (MIV-09), which was commercialized in 2009 during the H1N1 epidemics and displayed lower effectiveness in retrospective studies, did not induce the production of type I IFN in moDCs and blood cDCs. Furthermore, it inhibited the TIV-09-induced secretion of type I IFN by these DCs. Yet, both vaccines induced pDCs to secrete type I IFN indicating that different influenza vaccines activate different molecular signaling pathways in different DC subsets. In vivo, the baseline IFN signature was decreased upon administration of the H1N1 vaccine in 3/3 pediatric lupus patients. These results suggest that subtypes of non-adjuvanted influenza vaccines elicit long-term protection through different mechanisms and that a vaccine’s ability to induce an IFN response in DCs may palliate the absence of adjuvant and increase vaccine efficacy.

overall_design: 9 total samples. No replicates. 9 in vitro IL4 DC samples with 3 medium controls; 3 stimulated by non-adjuvanted monovalent H1N1 California vaccine (MIV-09); 3 stimulated by non-adjuvanted trivalent Fluzone®09-10 (TIV-09).

first: Athale

last: Shruti

first: Romain

first: LuAnn

last: Thompson-Snipes

first: Yuanyuan

last: Wang


title: Influenza Vaccines Differentially Regulate the Interferon Response in Human Dendritic Cell Subsets [HumanWG-6 v3.0 Beadchip Experiments]

overall_design: 108 total samples. No replicates. 9 in vitro pDC samples with 3 medium controls; 3 stimulated by non-adjuvanted monovalent H1N1 California vaccine (MIV-09); 3 stimulated by non-adjuvanted trivalent Fluzone®09-10 (TIV-09). 8 in vitro mDC samples with 3 medium controls; 3 stimulated by MIV-09; 2 stimulated by TIV-09. 91 ex vivo whole blood samples with 26 total saline controls at 13 time points with 2 donors; 26 total stimulated by MIV-09 at 13 time points with 2 donors; 39 total stimulated by TIV-09 at 13 time points with 3 donors

last: Banchereau