title: CNS lymphatic drainage and neuroinflammation are regulated by meningeal lymphatic vasculature [2D2 T cells]

status: Public on Sep 18 2018

city: Charlottesville

first: Christopher

last: Overall

state: Virginia

address: 409 Lane Road

department: Neuroscience

institute: University of Virginia

summary: Neuroinflammatory diseases, such as multiple sclerosis, are characterized by invasion of the brain by autoreactive T cells. The mechanism for how T cells acquire their encephalitogenic phenotype and trigger disease remains, however, unclear. The existence of lymphatic vessels in the meninges indicates a relevant link between the CNS and peripheral immune system, perhaps affecting autoimmunity. Here we demonstrate that meningeal lymphatics fulfill two critical criteria: they assist in the drainage of cerebrospinal fluid components and enable immune cells to enter draining lymph nodes in a CCR7-dependent manner. Unlike other tissues, meningeal lymphatic endothelial cells do not undergo expansion during inflammation, and they express a unique transcriptional signature. Notably, the ablation of meningeal lymphatics diminishes pathology and reduces the inflammatory response of brain-reactive T cells during an animal model of multiple sclerosis. Our findings demonstrate that meningeal lymphatics govern inflammatory processes and immune surveillance of the CNS and pose a valuable target for therapeutic intervention.

overall_design: Adult C57Bl6 mice were injected with 2D2 tdTOMATO T cells prior to meningeal lymphatic ablation (VIS) or control (PBS). EAE was induced with MOG(35-55). Activated 2D2 T cells (TCRb+CD4+CD44+tdTOMATO+) were isolated from dCLN and spleen at day8 post EAE induction

first: Antoine

last: Louveau

first: Jasmin

last: Herz

first: Maria

middle: N

last: Alme

first: Andrea

middle: F

last: Salvador

first: Michael

middle: Q

last: Dong

first: Kenneth

middle: E

last: Viar

first: Grace

last: Herod

first: James

last: Knopp

first: Joshua

last: Setliff

first: Alexander

last: Lupi

first: Sandro

last: Da Mesquita

first: Elizabeth

last: Frost

first: Alban

last: Gaultier

first: Tajie

middle: H

last: Harris

first: Rui

last: Cao

first: Song

last: Hu

first: John

last: Lukens

first: Igor

last: Smirnov

first: Guillermo

last: Oliver

first: Jonathan

last: Kipnis


title: Mrp1 is involved in lipid presentation and iNKT cell activation by Streptococcus pneumoniae

status: Public on Sep 14 2018

city: La Jolla

first: Shilpi

last: Chandra

state: California

address: 9420 Athena Circle

department: Division of Developmental Immunology

country: USA

institute: La Jolla Institute for Allergy and Immunology

summary: Invariant natural killer T cells (iNKT cells) are activated by lipid antigens presented by CD1d, but the pathway leading to lipid antigen presentation remains incompletely characterized. Here we show a whole genome siRNA screen to elucidate the CD1d presentation pathway. A majority of gene knockdowns that diminish antigen presentation reduced formation of glycolipid-CD1d complexes on the cell surface, including members of the HOPS and ESCRT complexes, genes affecting cytoskeletal rearrangement, and ABC family transporters. We validated the role in vivo for the multidrug resistance protein 1 (Mrp1) in CD1d antigen presentation. Mrp1 deficiency reduces surface clustering of CD1d, which decreased iNKT cell activation. Infected Mrp1 knock out mice show decreased iNKT cell responses to antigens from Streptococcus pneumoniae and were associated with increased mortality. Our results highlight the unique cellular events involved in lipid antigen presentation and show how modification of this pathway can lead to lethal infection.

overall_design: Expression data from macrophage cell line J774 overexpressing CD1d and F4/80+ peritoneal macrophages at steady state

last: Gray

first: William

middle: B

last: Kiosses

first: Archana

last: Khurana

first: Kaori

last: Hitomi

first: Catherine

middle: M

last: Crosby

first: Ashu

last: Chawla

first: Zheng

last: Fu

first: Meng

last: Zhao

first: Natacha

last: Veerapen

first: Stewart

middle: K

last: Richardson

first: Steven

middle: A

last: Porcelli

first: Gurdyal

last: Besra

first: Amy

middle: R

last: Howell

first: Sonia

last: Sharma

first: Bjoern

last: Peters

first: Mitchell

last: Kronenberg


title: Differentially expressed genes after miRNA or siRNA transfection in human cancer cell lines (T24, BOY, A549 and SK-MES)

status: Public on Dec 05 2018

city: Chiba

first: Naohiko

last: Seki

state: Chiba

address: 1-8-1, Inohana

department: Functional Genomics

country: Japan

institute: Chiba University

summary: To identify differentially expressed genes by anti cancer treatments (microRNAs or siRNAs) in human cancer, several cell lines (pancreatic cancer, esophageal cancer and lung squamous cell carcinoma) were subjected to Agilent whole genome microarrays.

overall_design: Human cell lines (T24, BOY, A549 and SK-MES) were treated with miRNAs (miR-143-5p and 143-3p), siRNA (si-KIF2A) and PLOD inhibitor.

first: Yasutaka

last: Yamada


title: Genetic and epigenetic evolution as a contributor to WT1-mutant leukemogenesis

status: Public on Sep 01 2018

city: New York

first: Richard

last: Koche

state: New York

institute: Memorial Sloan Kettering Cancer Center

type: Expression profiling by high throughput sequencing

summary: Genetic studies have identified recurrent somatic mutations in Acute Myeloid Leukemia (AML) patients, including in WT1 (Wilms’ tumor gene 1).  The molecular mechanisms by which WT1 mutations contribute to leukemogenesis have not yet been fully elucidated.  We investigated the role of Wt1 gene dosage in steady state and pathologic hematopoiesis.  Wt1 heterozygous loss enhanced stem cell self-renewal in an age-dependent manner, with increased stem cell function over time and age-dependent leukemic transformation. Wt1-haploinsufficient leukemias were characterized by progressive genetic and epigenetic alterations, including in known leukemia disease alleles, demonstrating a requirement for additional events to promote hematopoietic transformation.  Consistent with this observation, we found that Wt1 haploinsufficiency cooperates with Flt3-ITD mutation to induce fully penetrant AML.  Our studies provide insight into mechanisms of Wt1-loss in leukemogenesis and into the evolutionary events required to induce transformation of Wt1-haploinsufficient stem/progenitor cells.

overall_design: RNA-seq of sorted myeloid progenitors from Mx+ Wt1fl/+ mice of different ages compared to age-matched control counterparts

first: Elodie

last: Pronier

middle: P

first: Ross

middle: L

last: Levine


title: Chemical inhibition of the Mycobacterium tuberculosis DosRST two-component regulatory system by targeting sensor kinase heme and response regulator DNA binding

status: Public on Sep 10 2018

city: East Lansing

first: Robert

middle: B.

last: Abramovitch

state: MI

address: 567 Wilson Road

department: Microbiology and Molecular Genetics

institute: Michigan State University

summary: The purpose of this study was to characterize the mechanism of action of the small molecules HC104 and HC106

overall_design: Mtb strain CDC1551 or the DosR mutant was grown at 37ºC in T-25 vented, standing tissue culture flasks in 8 mL of buffered 7H9 medium to an OD600 of 0.1. Cultures were subsequently treated with 40 µM HC104A or HC106A. For the control, cultures treated with an equal amount of DMSO.  Following a 6 day incubation period, total bacterial RNA was extracted, quality controlled and sequenced. Each treatment was repeated with two independent (biological) replicates.

first: Huiqing

last: Zheng


title: Transcriptional and Epigenomic Landscapes of CNS and non-CNS Vascular Endothelial Cells

status: Public on Sep 06 2018

city: Baltimore

first: Mark

last: Sabbagh

state: MD

address: 725 North Wolfe Street

department: Molecular Biology and Genetics

institute: Johns Hopkins University School of Medicine

type: Methylation profiling by high throughput sequencing

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Vascular endothelial cell (EC) function depends on appropriate organ-specific molecular and cellular specializations. To explore genomic mechanisms that control this specialization, we have analyzed and compared the transcriptome, accessible chromatin, and DNA methylome landscapes from mouse brain, liver, lung, and kidney ECs. Analysis of transcription factor (TF) motifs at candidate cis-regulatory elements together with TF gene expression reveals both shared and organ-specific EC TFs. In the embryo, only those ECs that are adjacent to or within the developing CNS exhibit canonical Wnt signaling, which correlates precisely with blood-brain barrier (BBB) differentiation and Zic3 expression. In acutely cultured brain ECs, the rapid loss of BBB markers is closely correlated with down-regulation of canonical Wnt signaling. In the early postnatal brain, single cell RNA-seq of purified ECs reveals close relationships between veins and mitotic cells and between arteries and tip cells, and a division of capillary ECs into vein-like and artery-like classes.

overall_design: RNA-seq and MethylC-seq of postnatal day 7 (P7) vascular endothelial cells (ECs) from brain, liver, lung, and kidney of Tie2-GFP mice; RNA-seq and ATAC-seq of primary brain EC culture; single cell RNA-seq of P7 mouse brain ECs

first: Jacob

last: Heng

first: Chongyuan

last: Luo

first: Rosa

middle: G

last: Castanon

first: Joseph

last: Nery

first: Amir

last: Rattner

first: Loyal

last: Goff

last: Ecker

first: Jeremy

last: Nathans


title: Single cell RNA-sequencing of visceral adipose tissue Pdgfrβ+ cells

city: Durham

first: Rana

last: Gupta

state: North Carolina

address: Carmichael Building, 300 North Duke St.

department: Duke Molecular Physiology Institute

institute: Duke University

summary: We previously derived a doxycycline-inducible (Tet-On) lineage-tracing model that allows for the indelible labeling of Pdgfrb-expressing perivascular cells in adipose tissue of adult mice (PdgfrbrtTA; TRE-Cre; Rosa26RmT/mG; herein, “MuralChaser mice”). Prior to exposing animals to doxycyline, all cells within the stromal-vascular fraction (SVF) of adult gonadal WAT (gWAT) express membrane tdTomato from the Rosa26 locus. Following 9 days of exposure to doxycycline-containing chow diet, Cre-mediated excision of the loxP-flanked tdTomato cassette occurs in Pdgfrb-expressing cells, and membrane-bound GFP (mGFP) expression is constitutively activated. We set out to test the hypothesis that Pdgfrb-expressing perivascular cells in gonadal visceral WAT of adult mice are heterogeneous, with subpopulations harboring functionally distinct phenotypes. To this end, we performed single cell RNA-sequencing  of mGFP+ cells isolated from gWAT of lean (chow fed) 8 weeks-old male MuralChaser mice following 9 days of doxycycline exposure.

overall_design: We pooled tdTomato- GFP+ cells from gonadal white adipose tissue (WAT) from 5 MuralChaser mice. Single cell library preparation was performed using the 10X Genomics Single Cell 3’ v2 according to the manufacturer’s instructions.

first: R

first: C

last: Hepler


title: RNA-seq data

status: Public on Sep 05 2018

city: Seattle

first: Diego

last: Calderon

address: 3720 15th Ave NE

institute: University of Washington

summary: The immune system depends on a balanced interplay among a variety of specialized cell types transitioning between resting and stimulated states. While epigenome mapping efforts have focused on resting ex vivo immune cells, efforts to characterize stimulation-induced chromatin changes of diverse cell types has been limited. Thus, we collected ATAC-seq and RNA-seq data from immune cell types under resting and stimulated conditions from blood of healthy individuals, and cell types from fetal thymus samples.

overall_design:
166 RNA-seq samples of 25 blood cell types from 8 healthy donors.

* NOTE: The raw data of some Samples were updated on Jan 30, 2020. *

first: Michelle

last: Nguyen

first: Anja

last: Mezger


title: UGDH knockdown in MDA-MB-231

status: Public on Aug 21 2018

city: Houston

first: James

middle: Michael

last: Arnold

state: Texas

address: 1 Baylor Plaza

department: Biochemistry and Molecular Biology

institute: Baylor College of Medicine

summary:
Transcriptome profiling (RNA-Seq) to evaluate the connection the metabolic enzyme UGDH on gene expression.
The RNA libraries were prepared and sequenced at University of Houston Seq-N-Edit Core per protocols. Total libraries were prepared with Ovation® Universal Plus mRNA-Seq (NuGen) using 200 ng input RNA. The size selection for libraries were performed using SPRIselect beads (Beckman Coulter) and purity of the libraries were analyzed using the High Sensitivity DNA chip on Bioanalyzer 2100 (Agilent). The prepared libraries pooled and sequenced using NextSeq 500 (Illumina), generating ~15 million 2×76 bp paired-end reads per samples.
The RNA-seq raw fastq data were processed with RNA-Seq Express app within the Illumina BaseSpace app suite (www.basespace.illumina.com), which performed alignment with STAR aligner, assignment of aligned reads to genes, and differential gene expression with DESeq2
Analysis was performed comparing controls (shNT) against individual knockdowns (shU1 or shU2). Then, results from both analyses were compared for the intersection to identify commonly altered genes.

overall_design: mRNA profiles of MDA-MB-231 cells carrying either a control shRNA (shNT) or one of two independent shRNAs targeting UGDH (shUGDH-1, shUGDH-2) were generated by deep sequencing, in triplicate, using Illumina NextSeq 500.

first: Arun

last: Sreekumar


title: Specific Oxylipins Enhance Vertebrate Hematopoiesis via the Receptor GPR132

status: Public on Aug 10 2018

city: Boston

first: Leonard

last: Zon

state: MA

address: 1 Blackfan Circle

department: Oncology/Hematology

institute: Boston Children's Hospital

summary: Epoxyeicosatrienoic acids (EETs) are endogenous lipid signaling molecules with cardioprotective and vasodilatory actions. We recently showed that exogenous addition of 11,12-EET enhances hematopoietic induction and engraftment in mice and zebrafish. EETs are known to signal via a G-protein coupled receptor(s), but no specific EET receptor has been identified. Identification of an EET receptor would enable genetic interrogation of the EET signaling pathway and perhaps clinical use of this molecule. We developed a bioinformatic approach to identify the EET receptor based on the expression of GPCRs in cell lines with differential responses to EETs. We found 10 candidate EET receptors that are commonly expressed in three EET-responsive human cell lines, but not expressed in an EET-unresponsive line. Of these candidates, only GPR132 showed EET-responsiveness in vitro using a luminescence-based assay for β-arrestin recruitment. Knockdown of zebrafish gpr132b prevented EET-induced hematopoiesis, and marrow from GPR132 knockout mice showed decreased long-term engraftment capability. GPR132 has affinity for certain fatty acids in vitro, and we found that these same fatty acids enhance hematopoietic stem cell specification in the zebrafish. We conducted structure-activity relationship analyses using both in vitro and in vivo assays on diverse medium chain fatty acids. Certain oxygenated, unsaturated free fatty acids showed high activation of GPR132, while unoxygenated or saturated fatty acids had lower activity. Absence of the carboxylic acid moiety prevented activity, suggesting that this moiety is required for receptor activation. GPR132 responds to a select panel of polyunsaturated, oxygenated fatty acids to enhance both embryonic and adult hematopoiesis.

overall_design: To identify potential EET receptors, we performed RNAseq on 3 EET-binding cell lines and one non-binding cell lines, then identified GPCRs expressed in common in the EET-binding lines but missing from the non-binding line.

first: Jamie

last: Lahvic

first: Song

last: Yang


title: A dynamic co-expression map of early inflorescence development in Setaria viridis provides a resource for gene discovery and comparative genomics

status: Public on Aug 17 2018

city: St. Louis

first: Andrea

last: Eveland

state: MO

address: 975 N. Warson Road

institute: Donald Danforth Plant Science Center

summary: Setaria viridis is a small, rapidly growing grass species in the subfamily Panicoideae, a group that includes economically important cereal crops such as maize and sorghum. The S. viridis inflorescence displays complex branching patterns, but its early development is similar to that of other panicoid grasses, and thus is an ideal model for studying inflorescence architecture. Here we report detailed transcriptional resource that captures dynamic transitions across six sequential stages of S. viridis inflorescence development, from reproductive onset to floral organ differentiation. Co-expression analyses identified stage-specific signatures of development, which include homologs of previously known developmental genes from maize and rice, suites of transcription factors and gene family members, and genes of unknown function. This spatiotemporal co-expression map and associated analyses provide a foundation for gene discovery in S. viridis inflorescence development, and a comparative model for exploring related architectural features in agronomically important cereals.

overall_design: We profiled the transcriptome of S.viridis inflorescence primordia across six sequential developmental stages that captured key events from the reproductive transition to floral organ differentiation. Three to four biological replicates were used for each stage.

first: Jiani

first: Chuanmei

last: Zhu

first: Mathew

middle: S

last: Box

first: Elizabeth

last: Kellogg


title: Transforming growth factor beta-signaling in dendritic cells is required for immunotolerance to sperm in the epididymis

status: Public on Aug 08 2018

city: Tucson

first: Pawel

middle: R.

last: Kiela

state: AZ

address: 1501 N. Campbell Ave.

department: Pediatrics

institute: University of Arizona

summary: The epididymis exhibits a less restrictive physical blood-tissue barrier than the testis and, while numerous immunosuppressive factors have been identified in the latter, no mechanisms for epididymal immunotolerance have been identified to date. Therefore, data are currently insufficient to explain how the immune system tolerates the extremely large load of novel antigens expressed on sperm, which become present in the male body after puberty, i.e. long after central tolerance was established. This study tested the hypothesis that TGFβ-signaling in dendritic cells (DCs) is required for immunotolerance to sperm located in the epididymis, and that male mice lacking TGFβ-signaling in DCs would develop severe epididymal inflammation. To test this, we employed adult Tgfbr2ΔDC males, which exhibit a significant reduction of Tgfbr2 expression and TGFβ-signaling in DCs, as reported previously. Results show that Tgfbr2ΔDC males exhibit sperm-specific immune response and severe epididymal leukocytosis. This phenotype is consistent with epididymal loss of immunotolerance to sperm, and suggests that TGFβ-signaling in DCs is a factor required for a non-inflammatory steady state in the epididymis, and therefore for male tract homeostasis and function.

overall_design: We used microarrays to compare the global patterns of gene expression in the epididymis and testis as a function of TGFbR2 deletion specifically in dendritic cells

first: Fernando

last: Pierucci-Alves


title: SDR9C7 Promotes Lymph Node Metastases in Patients with Esophageal Squamous Cell Carcinoma

status: Public on Aug 14 2018

city: Chengdu

first: sen

last: qin

address: Tianhui Ave

country: China

institute: Chengdu military general hospital

summary: To  identify the lymph node (LN) metastasis-associated genes in primary ESCC tumors, gene expression proﬁling assay (GEP) was performed to identify the differences in gene expression profiles between primary ESCC tumors that were with LN metastases (N+) and those without LN metastases (N-).

overall_design: The primary surgical tissues from 3 esophageal squamous cell carcinoma (ESCC) tumors that were N-, and 3 patients with strong N+ tumors, were collected for GEP assays from December 2009 to May 2010

first: Shanhong

last: Tang


title: Expression analysis of A549 cells following treatment with conditioned medium from polarized macrophages

status: Public on Jul 26 2018

city: Taipei

first: Chih-Chan

last: Lee

address: No.155, Sec.2, Linong Street

country: Taiwan

institute: Molecular medicine program

summary: Tumor-associated macrophages (TAMs) are one of the most abundant host immune cells in tumor microenvironments. In this study, we investigated the functional role of M1 and M2 polarized macrophages on cancer cells. Expression microarray and bioinformatics analysis were performed in A549 cells treated with conditioned media from different subtype macrophages.

overall_design: Lung cancer cells, A549, were cultured in M0, M1, M2 macrophages conditioned media for two days. The different subtype macrophages were differentiated by IFN-gamma, LPS, IL-4 and dexamethasone.

first: Muh-Hwa


title: Expression analysis of tumor-associated macrophages from primary tumor and metastatic lung

summary: In tumor microenvironment, tumor-associated macrophages (TAMs) have been characterized as M1-like or M2-like phenotype. In this study, we investigated the characteristics and functional roles of different TAMs on cancer metastasis. We isolated TAMs from primary tumor and metastatic lung and performed microarrays to identify the gene expression in distinct TAMs populations.

overall_design: TAMs were sorted by CD11B+ and F4/80+ from tumor-bearing mice. Bone marrow-derived macrophages were differentiated from the bone marrow of health mice and sorted by CD11b+ and F4/80+.


title: Exposure to polybrominated biphenyl (PBB) associates with DNA methylation differences across the genome

status: Public on Aug 01 2018

city: Atlanta

first: Sarah

middle: Whelan

last: Curtis

state: GA

address: 101 Woodruff Circle NE, Ste 42

institute: Emory University

type: Methylation profiling by genome tiling array

summary: Polybrominated biphenyl (PBB) is an endocrine-disrupting compound which was accidently added to the food supply in Michigan in the 1970’s. Highly exposed individuals and their children have numerous health problems, though the underlying mechanism behind these health problems remains unknown. Other endocrine-disrupting compounds have been linked to epigenetic differences, but no epigenetic studies have been done for PBB. In this study, DNA from the blood of individuals with PBB exposure from the Michigan PBB Registry was interrogated with the MethylationEPIC BeadChip (N = 659). Associations between each of the ~850,000 CpG sites and current, serum PBB levels were tested with a linear regression that controlled for age, sex, and cell type proportions. We show that exposure to PBB is associated with differences in epigenetic marks that suggest that it is acting similarly to estrogen and is associated with dysregulated immune system pathways.

overall_design: DNA methylation was assessed in peripheral blood from people exposed to polybrominated biphenyl (PBB). Peripheral blood DNA was extracted, bisulfite converted and genome-wide DNA methylation was interrogated using the HumanMethylationEPIC BeadChip (Illumina).

middle: W

first: Varun

last: Kilaru

first: Dawayland

middle: O

last: Cobb

first: Michele

last: Marcus

first: Karen

middle: N

last: Conneely

first: Alicia

last: Smith


title: Tdrd6a recruits Ziwi-bound piRNAs and coordinates deposition of germ plasm mRNAs into primordial germ cells [small RNA]

status: Public on Jul 24 2018

city: mainz

first: lucas

last: kaaij

address: ackermannweg 4

department: RNA silencing

country: Germany

institute: IMB

type: Non-coding RNA profiling by high throughput sequencing

summary: Germ plasm, the Balbiani body and nuage are evolutionary conserved structures essential for germ cell specification and maintenance. We describe Tdrd6a as a component of these structures with two distinct molecular functions. First, Tdrd6a facilitates the accumulation of the typical antisense-bias of piRNAs, without having effects on piRNA biogenesis signatures. Second, we show that Tdrd6a is required for Balbiani body and germ plasm integrity, and associates with RNA-binding proteins and germ plasm mRNAs. On the cell-biological level, maternally contributed Tdrd6a strongly impacts germ cell formation, but is dispensable for fertility. Using single-cell RNA-sequencing we demonstrate that Tdrd6a promotes early germ cell development and regulates the stoichiometry of germ plasm mRNAs. We propose that Tdrd6a functions as a scaffold to recruit correct ratios of germ plasm transcripts and to accumulate antisense piRNA complexes in order to ensure both specification and maintenance of germ cells.

overall_design: Zebrafish were grown under standard conditions. Total ovary or size selected Oocytes were used to extact RNA from het of tdrd6a mutant fish.

first: Lucas

last: Kaaij


title: Global mapping of polyadenylation site use in proliferating and quiescent (contact-inhibited and serum-starved) cells

status: Public on Jul 19 2018

city: Los Angeles

first: Hilary

last: Coller

state: CA

address: 610 Charles E Young Drive E

department: Molecular, Cell and Developmental Biology

institute: University of California, Los Angeles

summary:
Purpose: To determine genes that undergo alternative polyadenylation in proliferating versus quiescent fibroblasts.
Methods: Three different biological replicates (fibroblast strains, 10-1, 12-1, and 12-2) were used for generating proliferating and quiescent (7-day contact inhibited and 7-day serum-starved) cells. RNA extracted from these cells were used for library preparation. cDNA fragments were enriched for the junction between the poly(A) site and the end of the 3' UTR using a modified high throughput sequencing protocol (GnomeGen). cDNA libraries were created according to Gnome-Gen RNA-seq library preparation kit for RNA profiling except Amgen Ampure XP beads were used instead of the Gnome-gen size selector product to remove ligation reaction products before proceeding to the reverse transcription step.  The libraries were sequenced on an Illumina HiSeq 2000 instrument. The sequencing reaction wasa run for 147 cycles. Reads from pol(A) enriched cDNA libraries were aligned to the genome using the STAR alignment algorithm after the computational removal of untemplated adenosines.
Results: Polyadenylation site selection was significantly altered in approximately 10% of genes in quiescent compared with proliferating fibroblasts.  Contact inhibited and serum starved fibroblasts had similar polyadenylation site selection profiles.
Conclusions: Quiescence is associated with changes in polyadenylation site selection.

overall_design: Three samples of proliferating fibroblasts, three samples of contact inhibited fibroblasts and three samples of 7-day serum starved fibroblasts were analyzed with polyadenylation site-enriched RNA-Seq.

last: Johnson

first: Wei

last: Wang

first: Jessica

last: Buckles

first: Mithun

last: Mitra


title: Tdrd6a recruits Ziwi-bound piRNAs and coordinates deposition of germ plasm mRNAs into primordial germ cells [scRNA-Seq]

overall_design: Single cell were sorted directly in Trizolfrom embryos spawned by mz tdrd6a-/- mother and wt mother carrying a kop::egfp-f-nos1-3’UTR  transgene. Thereafter single cell trizol extractio was performed followed by RT, IVT and RNA-seq library prep.


title: Tdrd6a recruits Ziwi-bound piRNAs and coordinates deposition of germ plasm mRNAs into primordial germ cells [RIP-Seq]

overall_design: RNA was extracted from ovary tissue or from RIP experiments by Trizol extraction.


title: Tdrd6a recruits Ziwi-bound piRNAs and coordinates deposition of germ plasm mRNAs into primordial germ cells

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

overall_design: Refer to individual Series


title: The effect of productive HPV16 infection on the cervical epithelium transcriptome

city: Cambridge

first: Anna

last: Salzberg

address: 399 Binney St UNIT 300

institute: Rubius Therapeutics

summary: Microarray analysis of the effect of early stage, productive HPV16 infection on the transcriptome of human cervical keratinocytes. Our results provide for the first time, global gene expression changes in cervical epithelium that produces high levels of viral titers.

overall_design: RNA was isolated from 10-day organotypic raft cultures of human cervical keratinicoytes persistently infected with HPV16.  RNA isolated from raft cultures of primary cervical keratinocytes were used as control.

first: Craig

last: Meyers

first: Sa-Do

last: Kang


title: RNA-Sequencing identifies differentially expressed genes in adult male tilapia following low Benzo (a) Pyrene (BaP) exposure

status: Public on Jul 06 2018

city: Mérida

first: Reyna

middle: Cristina

last: Colli-Dula

state: Yucatán

address: Km. 6 Ant Carretera a Progreso

department: Recursos del Mar

country: Mexico

institute: CINVESTAV

summary:
Purpose: The objective of this study was to identifiy if a low dose of BaP would alter gene expression profiles and key pathways in the liver and testis of adult male tilapia

Methods: The liver and testis tissue from the BaP-treated or control groups fish were collected and then total RNA was extracted, RNA integrity (RIN) was evaluated (RIN >7) for Illumina RNA-Seq library construction and Sequencing. Gene read mapping was performed with TopHat and differential expression analysis (DE) was conducted using exact test with R package EdgeR (p < 0.05; fold change > ± 1.5 were considered as significant). Pathway analysis was performed using the gene set enrichment analysis (GSEA) and subnetwork enrichment analysis (SNEA) applying the Mann-Whitney test with an alpha level of p < 0.05 to identify the biological mechanism that underlie the BaP effects in male tilapia.

Results: RNA-Seq analysis using EdgeR showed a larger number of altered genes in liver than in testis as a result of BaP exposure. In liver, 1444 genes were significantly differentially expressed, while in the testis only 309 showed any changes using a p < 0.05 and fold change > ± 1.5

Conclusions: Low dose of BaP can alter genes and molecular pathways in the liver and testis in male adult tilapia, and these changes could be associated with biological endpoints disruption.

overall_design: Illumina NextSeq500 Sequencing was used to generate transcriptomic profiles for BaP-exposure liver (n=4 or control, n=4 for BaP) and testis (n=3 for control, n=3 for BaP) from adult male tilapia

first: R

middle: C

first: D

last: Moreno

first: X

last: Fang


title: Transcriptional profiling of drug-induced changes in mice nucleus accumbens and prefrontal cortex

status: Public on Jul 17 2018

city: Krakow

first: Marcin

last: Piechota

address: Smetna 12

department: Molecular Neuropharmacology

country: Poland

institute: Institute of Pharmacology PAS

summary: We applied next-generation sequencing to analyse changes in the  expression of Dclk1 gene isoforms in the brain in response to several psychoactive  drugs with diverse pharmacological mechanisms of action. We used bioinformatics  tools to define the range and levels of Dclk1 transcriptional regulation in the mouse  nucleus accumbens and prefrontal cortex

overall_design: Sequencing was performed on ribo-depleted RNA samples derived from the mouse nucleus accumbens septi (Nac) and prefrontal cortex (PFCx) 2 hours after treatment with antidepressants (venlafaxine and mianserin), antipsychotics (haloperidoland risperidone), a psychostimulant (methamphetamine) or a psychotomimetic (ketamine)

first: Magdalena

last: Zygmunt

first: Michal

last: Korostynski


title: Inter-tumoral heterogeneity in SCLC is influenced by the cell-type of origin

status: Public on Jul 13 2018

city: Stanford

first: Dian

last: Yang

address: B261 Beckman Center, 1291 Welch Rd.

institute: Stanford University

summary: We describe two different routes of SCLC metastatic progression

overall_design: We performed RNA-seq on primary tumors and metastasis from SCLC mouse model (Rb/p53/p130/mTmG) transduced by Ad-CMV-Cre or Ad-CGRP-Cre

first: Peyton

last: Greenside

first: Julien

last: Sage

first: Monte

last: Winslow


title: DNA topoisomerase and supercoil accumulation across yeast genome

status: Public on Jul 02 2018

city: Milano

first: Mohamood Adhil

last: Mohammed Iqbal

state: Lombardia

address: Via Adamello 16

country: Italy

institute: IFOM - The FIRC Institute of Molecular Oncology

type: Genome binding/occupancy profiling by genome tiling array

summary:
DNA topoisomerases assist DNA replication & transcription events by controlling supercoiling alterations. We investigated supercoil distribution across the yeast genome and compared with the accumulation of RNA pol2 and DNA topoisomerases particularly in S-phase. Our data indicate that Top2 along with Hmo1 maintain negative supercoil at gene boundaries by stabilizing alternative DNA structures.
To understand how DNA superhelical tension accumulates across the genome we have adopted previously described method [Naughton C et al., 2013] to budding yeast where a biotin molecule was attached to TMP via a linker (bTMP). The Chip on chip analysis for proteins was carried out as described (Bermejo R et al., 2009).  For RNA-DNA hybrids DRIP-chip is carried out as described previously (Chan YA et al., 2014). Supercoiled regions are then compared with RNA pol2 (RPB3-chip), DNA Topoisomerase (Top1-chip) & RNA-DNA hybrid (DRIP-chip).

overall_design: Protein-chip, bTMP-chip and DRIP-chip were carried out in following Saccharomyces Cerevisiae strains:  wildtype (W303),  top2-1△, hmo1△, top1△, top2-1;hmo1△, top2-1;top1△, WT-Ctrlplasmid, Wt-TopAplasmid, top2-1;top1△-Ctrlplasmid, top2-1;top1△-TopAplasmid, sen1-cl, rrm3△, rnh1△

first: Yathish

middle: J

last: Achar

first: Mohamood

last: Adhil

first: Marco

last: Foiani


title: Environment-induced same-sex mating suggests homothallism is a primary mode of sexual reproduction in the yeast Candida albicans

status: Public on Aug 29 2018

city: Shanghai

first: Jian

last: Bing

address: Songhu Road, Yangpu District

department: School of Life Sciences

institute: Fudan University

summary: In this study, we report that glucose starvation and oxidative stress, common environmental stresses encountered by the pathogen, induce the development of mating projections and efficiently permit same-sex mating in C. albicans with an “a” mating type (MTLa/a).

overall_design: total RNA profiles of cells grown in YPD-K or YP-K

first: Guanghua

last: Huang

first: Guobo

last: Guan


title: Effect of top2 and hmo1 on chromatin architecture and transcription in yeast

summary: DNA topoisomerase-2 and high mobility group protein Hmo1 are known to regulate chromatin architecture by regulating gene boundaries. Here we report how these proteins affect global RNA level after inactivation of Top2 and Hmo1. Our data indicate that inactivating Hmo1 has a drastic effect on transcription levels of 20% yeast genes, however, this phenomenon can slightly be rescued by inactivating Top2 functions. Also, we study the Top2 and Top1 role in nucleosome architecture with and without expressing E.coli TopA. In top2-1;top1∆ condition with TopA expressed, it affects the nucleosome occupancy at the global level compared with top2-1;top1∆-Control plasmid. The ChIA-PET (Chromatin interaction analysis by paired-end tag sequencing) method is used to address whether a specific protein is engaged in the chromosomal interactions. Epitope tagged Top2 protein is used as probe in ChIA-PET experiments to map Top2 mediated chromatin-chromatin interactions.

overall_design: RNA-seq for yeast strains: wildtype, top2-1, hmo1∆, and hmo1∆;top2-1. ChIP-seq for Hmo1 Protein: Hmo1tag-ControlPlasmid, Hmo1tag-top2-1;top1∆-TopAPlasmid. Histone H3 ChIP-seq: wildtype-ControlPlasmid, wildtype-TopAPlasmid, top2-1;top1∆-ControlPlasmid, top2-1;top1∆-TopAPlasmid.


title: Large-scale differences in gene expression, including genes encoding type IV pili-associated proteins, are seen when Clostridium perfringens is grown in liquid or on surfaces

status: Public on Jun 30 2018

city: Blacksburg

first: Stephen

last: Melville

address: 217 LS1, Washington St.

department: Biological Sciences

institute: Virginia Tech

summary: Clostridium perfringens is a Gram-positive anaerobic pathogen that causes multiple diseases in humans and animals.  C. perfringens lack flagella but have type IV pili (TFP) and can glide on agar surfaces by forming filaments of cells aligned end to end.  When cells are placed on agar surfaces, they become elongated, flexible and have TFP on their surface.  To understand the basis of these phenotypes, cells were grown in three types of liquid media and on agar plates with the same medium to compare gene expression using RNA-seq.   Hundreds of genes were differentially expressed, including genes associated with TFP functions, which were higher on plates than in liquid.  The gusA gene, encoding the enzyme β-glucuronidase, was inserted into the chromosome downstream of TFP promoters and expression measured in liquid and on plates.  β-glucuronidase activity was proportional to the amount of transcripts seen with RNA-seq n liquid-grown cells, but not plate-grown cells, suggesting post-transcriptional regulation of these genes on surfaces.  An sRNA adjacent to pilA1 (SR79) was expressed at higher levels on plates in all media; overexpression of this sRNA resulted in longer cells on plates and shorter lag times in liquid suggesting SR79 plays a role in adapting to surface growth.

overall_design: Comparison of gene expression in bacteria grown in liquid or on plates

first: Samantha

last: Murray

first: Andrea

middle: H

last: Hartman

first: Tara

middle: M

last: Gallagher

first: Roderick

middle: V

last: Jensen

middle: B

first: Gary

last: Camper


title: Transcriptome analysis of mouse Cnot6l-/- preovulatory oocytes (GV), ovulated eggs (MII), and 1-cell zygotes

status: Public on Jul 10 2018

city: Zagreb

first: Vedran

last: Franke

address: Horvatovac 102a

department: Department of biology

country: Croatia

institute: University of Zagreb

summary: We profiled transcriptomes from Cnot6l deadenylase knock-out mouse GV oocytes, MII eggs and 1-cell zygotes in order to analyse its function during the oocyte-to-embryo (OET) transition.  Transcriptome of wild-type golden hamster GV oocytes was also profiled.

overall_design:
bulk RNA-seq of 20 samples:
- 3 replicates of each WT and Cnot6l knock-outs in 3 mouse developmental stages (GV, MII, 1-cell)
- 2 replicates of WT golden hamster GV oocytes

first: Filip

last: Horvat

first: Petr

last: Svoboda


title: Asf1a resolves bivalent chromatin domains for the induction of lineage-specific genes during mouse embryonic stem cell differentiation

status: Public on Jun 22 2018

city: New York

first: zhiguo

last: zhang

state: NY

address: 1130 St. Nicholas Avenue

department: Pediatric and Genetics and Development

institute: Columbia University

summary: Bivalent chromatin domains containing both repressive H3K27me3 and active H3K4me3 modifications are barriers for the expression of lineage-specific genes in embryonic stem (ES) cells and must be resolved for the transcription induction of these genes during differentiation, a process that remains largely unknown. Here, we show that Asf1a, a histone chaperone involved in both nucleosome assembly and disassembly, regulates the resolution of bivalent domains and activation of lineage-specific genes during mouse ES cell differentiation. Deletion of Asf1a does not affect the silencing of pluripotent genes, but compromises the expression of lineage-specific genes during ES cell differentiation. Mechanistically, the Asf1a-histone interaction, but not the role of Asf1a in nucleosome assembly is required for gene transcription. Asf1a is recruited to several bivalent promoters, partially through association with transcription factors, and mediates nucleosome disassembly during differentiation. We suggest that Asf1a-mediated nucleosome disassembly provides a means for resolution of bivalent domain barriers for induction of lineage-specific genes during differentiation.

overall_design: To analyze how Asf1a regulates bivalent gene expression during ES differentiation, we performed RNA-seq, H3K4me3 and H3K27me3 ChIP-seq in WT and Asf1a KO mouse ES cells and differentiated neural precursors

first: Yuan

last: Gao

first: Haiyun

last: Gan

first: Zhenkun

last: Lou

first: Zhiguo

last: Zhang


title: BORIS knockdown in colorectal cancer cells HCT116 and Caco-2

status: Public on Jul 01 2018

city: Hangzhou

first: Yanmei

state: Zhejiang

address: Tianmushan Road No.182

department: Center for Molecular Medicine

institute: Zhejiang Academy of Medical Science

summary:
BORIS expresses abnormally in colorectal cancer cells. Both the expression and the copy number of BORIS are remarkably higher in colorectal cancer cells than in normal colon or rectal cells. BORIS is potential diagnosis, prognosis or therapeutic target for colorectal cancer. To expand our view of the signaling pathway related with BORIS, altered gene expression by BORIS knockdown was assessed by microarray assay.
To study the gene regulation by BORIS knockdown, microarray assay was applied to screen the gene expression regulated by BORIS siRNA in colorectal cancer cells HCT116 and Caco-2

overall_design: HCT116 and Caco-2 cells were silenced by siRNA of BORIS for 3 days and extracted for the RNA. Silenced samples and the controls were compared to screen for gene expression difference by Affymetrix PrimeView human gene expression array.

first: Mengdie

last: Fang

first: Juan

last: Ren

first: Yongfei

last: Song

first: Haining

last: Yuan

first: Jianfei

first: Dongmei

last: Yan

first: Xiaoju

last: Wang


title: LincU Preserves Naive Pluripotency by Restricting ERK Activity in Embryonic Stem Cells

status: Public on Jun 20 2018

first: Zeyidan

last: Jiapaer

address: No.1239, Siping Road

institute: Tongji University


title: LincU Preserves Naive Pluripotency by Restricting ERK Activity in Embryonic Stem Cells [knockdown]

summary: While the functional roles of long non-coding RNAs (lncRNAs) have been increasingly identified, few lncRNAs that control the naive state of embryonic stem cells (ESCs) are known. Here, we report a naive-state-associated lncRNA, lincU, which is intrinsically activated by NANOG in naive mESCs. LincU-deficient mESCs exhibit a primed-like pluripotent state and potentiate the transition from naive state to primed state, whereas ectopic lincU expression maintains mESCs in the naive state. Mechanistically, we demonstrate that lincU directly binds and stabilizes the DUSP9 protein, an ERK-specific phosphatase, and then constitutively inhibits the ERK1/2 signaling pathway, which critically contributes to maintenance of the naive state. Importantly, we reveal the functional role of lincU to be evolutionarily conserved in human. Our findings thus unveil lincU as a conserved lncRNA that intrinsically restricts MAPK/ERK activity and maintains the naive state of ESCs.

overall_design: LincU knockdown RNA-Microarray in 46C mouse embryonic stem cells. LincU sequence is available in a supplementary file at the foot of this record.

first: Guoping

last: Li


title: Gene expression analysis of mouse ovarian cancer treated with anti-VEGF antibody

status: Public on Jun 19 2018

city: Kyoto

first: Naoki

last: Horikawa

address: 54, Kawahara-cho, Shogoin, Sakyo-ku

department: Gynecology and Obstetrics

institute: Kyoto University

summary:
Ovarian cancer is a highly aggressive female tract cancer. Anti-VEGF therapy is widely used for the treatment of ovarian cancer, however it has shown moderate impact on patient prognosis.
The elucidation of mechanism of drug resistance is highly beneficial for the advances in ovarian cancer treatment. Here, we investigated gene expression profile of HM-1 tumor which were treated with or without anti-VEGF antibody (B20-4.1.1) using transcriptome array to identify special molecular features of ovarian tumor after anti-VEGF treatment.

overall_design: To reveal molecular mechanisms underlying ovarian tumor after anti-VEGF treatment, we established mouse ovarian cancer model by subcutaneously injected HM-1 mouse ovarian cancer cells into B6C3F1 mice. We confirmed that the efficacy of anti-VEGF treatment against HM-1 tumor is slight. HM-1 tumor samples were collected after anti-VEGF therapy or Rat IgG control, and we extracted RNA from tumors and analyzed gene expression profiling.

first: Kaoru

last: Abiko

first: Tsukasa

last: Baba

first: Junzo

last: Hamanishi

first: Ryusuke

last: Murakami

first: Mana

last: Taki

first: Masayo

last: Ukita

first: Yuko

last: Hosoe

first: Ikuo

last: Konishi

first: Noriomi

last: Matsumura

first: Masaki

last: Mandai


title: LincU Preserves Naive Pluripotency by Restricting ERK Activity in Embryonic Stem Cells [overexpression]

overall_design: To determine the responsible mechanism underlying lincU-induced key phenotypes in mESCs，we performed global transcriptome analyses of  lincU overexpression mESCs by microarray. LincU sequence is available in a supplementary file at the foot of this record.


title: Analysis of the translational landscape of Arabidopsis mitochondria

status: Public on May 31 2018

city: ORSAY

first: Pierre

last: Bertin

address: Batiment 400, Université Paris Sud

country: France

institute: Institut de Biologie Intégrative de la Cellule

summary: Messenger RNA translation is a complex process that is still poorly understood in eukaryotic organelles like mitochondria. Growing evidence indicate though that mitochondrial translation differs from its bacterial counterpart in many key aspects. In this analysis, we used the ribosome profiling technology to generate a genome-wide snapshot view of mitochondrial translation in Arabidopsis. We show that, unlike in humans, most Arabidopsis mitochondrial ribosomes footprints measure 27 and 28 bases. We also reveal that respiratory subunits encoding mRNAs show much higher ribosome association than other mitochondrial mRNAs, implying that they are translated to higher levels. Homogenous ribosome densities were generally detected within each respiratory complex except for complex V where higher ribosome coverage corroborate with higher needs in specific subunits. In complex I respiratory mutants, a slight reorganization of mitochondrial mRNAs ribosome association was detected involving an increase in ribosome densities on certain ribosomal protein encoding transcripts and a reduction in the translation of a few complex V mRNAs. Altogether, our observations reveal that plant mitochondrial translation is a dynamic process and that translational control is important for gene expression in plant mitochondria. This study paves the way for future advances in the understanding of translation in higher plant mitochondria.

overall_design: In this study, 2 Arabidopsis mutants are compared to the wild type using duplicates for the Ribosome Profiling experiment. RNASEQ: was conducted to observe the editing rates of Col0 transcripts. RIBOSEQ: was conducted to observe the ribosome occupancy on each transcript of wild type and mutants.

first: Noelya

last: Planchard

first: Martine

last: Quadrado

first: Céline

last: Dargel-Graffin

first: Isabelle

last: Hatin

first: Olivier

last: Namy

first: Hakim

last: Mireau


title: Cooperation of GRSF1 and the mitochondrial degradosome (hSuv3-PNPase complex) in degradation of mitochondrial RNA

status: Public on Jun 17 2018

city: Warszawa

first: Tomasz

middle: M.

last: Kuliński

state: Mazowieckie

address: ul. Ks. Trojdena 4

institute: International Institute of Molecular and Cell Biology in Warsaw

summary: Vertebrate mitochondrial genomes show extraordinary GC skew which results in the synthesis of RNAs prone to form G-quadruplexes (G4). Such RNAs, although mostly non-coding, are transcribed at high rate and swiftly degraded by an unknown mechanism. Comprehensive proteomic and transcriptomic approaches revealed that quasi-RRM protein GRSF1 together with the mitochondrial degradosome composed of RNA helicase hSuv3 and PNPase control the levels of G4 containing RNAs in mitochondria. When this machinery is inactivated non-coding mtRNAs are upregulated and novel, previously undescribed transcript, which we named tRNA-like, strongly accumulates. In vitro reconstitution experiments showed that GRSF is responsible for G4 RNA melting essential for degradosome-mediated decay.

overall_design: There are 2 sets of data which correspond to two RNAseq studies (degradosome set and GRSF1 set). Each set includes experiments which were performed in triplicates. The aim of the first studies was to analyze the effect of inactivation of the mitochondrial degradosome components (hSuv3, PNPase) on mitochondrial RNA (mtRNA). In the second studies we examined influence of GRSF1 silencing of mtRNA. In both cases we analyzed long transcripts and small RNAs. Strand-specific library preparation procedures were applied. Ligation-based approach was used to analyze short RNAs (from ~20 to ~200 nucleotides) whereas longer RNAs (> ~100 nucleotides) were analyzed after their random fragmentation followed by reverse transcription primed with random oligomers (dUTP-based protocol). RNA was isolated from unfractionated cells using TRI-Reagent. Before preparation of long transcripts libraries total RNA was subjected to rRNA depletion. This step was omitted in preparation of libraries dedicated for analysis of short RNAs. Total RNA was used for preparation of small RNA libraries. Libraries from different studies (degradosome and GRSF1) were sequenced with the help of different Illumina sequencing platforms. Dysfunction of hSuv3 was achieved by inducible expression of its dominant-negative mutant form (hSuv3-G207V), which we shown to be equivalent to hSuv3 silencing (PMID: 19864255), whereas PNPase or GRSF1 were depleted by siRNA transient transfections. In the case of GRSF1 studies as controls we analyzed untreated cells and cells transfected with non-targeting siRNA. In the case of degradosome studies as controls we analyzed untreated cells and cells expressing siRNA-insensitive version of PNPase (rescue experiment).

first: Zbigniew

last: Pietras

last: Kulinski

first: Andrzej

last: Dziembowski

first: Roman

last: Szczesny


title: Genome Wide Distribution of Linker Histone H1.0 is Independent of MeCP2 II

status: Public on May 28 2018

city: Houston,

first: Hari Krishna

last: Yalamanchili

state: TX

address: 1250 Moursund St #1125,

institute: Baylor College of Medicine

summary: Previous studies suggested that MeCP2 binds to linker DNA and competes with linker histone H1 to regulate chromatin structure, but this hypothesis has never been tested in vivo. Here, we expressed Flag-tagged H1.0 in forebrain excitatory neurons in mice and performed ChIP-Seq to reveal H1.0 distribution and its relationship with MeCP2. Unexpectedly, we detected no major change in H1.0 upon MeCP2 depletion, revealing that MeCP2 functions independent of linker H1.0.

overall_design:
ChIP-seq of MeCP2 from the mouse frontal cortices, which were obtained from the wild-type (WT) and Cam-tTA:Fl-H1.0 mice expressing Flag-tagged H1.0 in the Camk2a+ excitatory cells (three biological replicates).

MeCP2 ChIP fh-Mecp2+/y peaks are included in the file Mecp2.FH.MACS_peaks.bed available on the SuperSeries record.

first: Aya

last: Ito-Ishida

first: Hari

middle: K

first: Yingyao

last: Shao

first: Zhandong

last: Liu

first: Huda

middle: Y

last: Zoghbi


title: Genome Wide Distribution of Linker Histone H1.0 is Independent of MeCP2

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

overall_design: Refer to individual Series


title: Genome Wide Distribution of Linker Histone H1.0 is Independent of MeCP2 I

overall_design:
ChIP-seq of Flag-tagged H1.0 in the Camk2a+ excitatory cells of the frontal cortices, which were obtained from the Mecp2-positive and Mecp2-null male mice (three biological replicates per genotype).

MeCP2 ChIP fh-Mecp2+/y peaks are included in the file Mecp2.FH.MACS_peaks.bed available on the SuperSeries record.


title: SWI/SNF remains localized to chromatin in the presence of SCHLAP1

status: Public on May 14 2018

city: Chapel Hill

first: Jesse

last: Raab

address: 120 Mason Farm Rd

department: Genetics

institute: University of North Carolina Chapel Hill


title: Disruption of GRIN2B impairs differentiation in human neurons

status: Public on May 21 2018

city: Montreal

first: Carl

last: Ernst

state: Quebec

address: 6875 Boulevard LaSalle

country: Canada

institute: Douglas Mental Health University Institute

summary: Mutations in GRIN2B are associated with intellectual disability in humans. We generated iPSC derived mature cortical neurons with mutations in GRIN2B and compared them to isogenic control cells. We found that both loss of function (LOF) and reduced dosage (RD) mutations in GRIN2B lead to reduced expression of NMDAR genes and increased expression of marker of immaturity, including KI67 and MET.

overall_design: Examination of transcriptome in iPSC-derved mature neurons with and without the presence of mutations in GRIN2B

first: Scott

last: Bell

first: Gilles

last: Maussion


title: SWI/SNF remains localized to chromatin in the presence of SCHLAP1 (ATAC-seq)

summary: Changes to chromatin openness were measured by ATAC seq in RWPE1 cells expression SCHLAP1 or a control cell line infected with a LACZ lentivirus.

overall_design: 3 indpendent replicates of RWPE1 cells infected with either LACZ or SCHLAP1 lentivirus were used to prepare ATAC-seq libraries as described in Buenrostro et al 2013.

middle: R

first: Terry

last: Magnuson


title: The distribution of Brahma in the genome of Drosophila melanogaster S2 cells and the roles of Brahma in transcription and pre-mRNA processing

status: Public on May 11 2018

city: Stockholm

first: Neus

last: Visa

address: Svante Arrhenius väg 20c

department: Molecular Biosciences, The Wenner-Gren Institute (MBW)

country: Sweden

institute: Stockholm University

summary: We have carried out ChIP-seq to study the distribution of Brahma in the genome of S2 cells and to identify the Brahma-bound genes. We have also knocked down Brahma by RNA interference and we have carried out RNA-seq to determine the changes induced by Brahma depletion in the transcriptome of S2 cells. Furthermore, we have carried out RNA-seq experiments to profile S2 cells in which we first deplete Brahma by RNA interference and then express recombinant Brahma (either wild-type or ATPase mutant). These datasets are being used in several projects aimed at gaining understanding on the roles of Brahma in transcription regulation and pre-mRNA processing.

overall_design: 12 samples total; 8 RNA-Seq samples (4 conditions, 2 biological replicates each); and 4 ChIP-seq samples (Drosophila BRM IP and Input; 2 biological replicates).

first: Antonio

last: Jordán-Pla

first: Simei

last: Yu

first: Johan

last: Waldholm

first: Thomas

last: Källman

first: Ann-Kristin

last: Östlund Farrants


title: SWI/SNF remains localized to chromatin in the presence of SCHLAP1 (ChIP-seq)

summary: ChIP-seq of SWI/SNF subunits and histone modifications in RWPE1 cells

overall_design: For histone modifications ChIP was performed on RWPE1; LACZ and RWPE:1SCHLAP1 cells using sonicated chromatin as input. For SWI/SNF subunits ChIP was performed in RWPE1;SCHLAP1 cells fragmented by Mnase


title: SWI/SNF remains localized to chromatin in the presence of SCHLAP1 (RIP-seq)

summary: RNA immunoprecipitation of SWI/SNF in prostate cacner

overall_design: Crosslinked cells were immunoprecipitated with SMARCA4 or SFPQ antibodies from 22Rv1 or LNCaP prostate cancer cell lines


title: Discovery and characterization of variance QTLs in human induced pluripotent stem cells

status: Public on Sep 21 2018

city: Chicago

first: John

middle: D

last: Blischak

state: IL

address: 920 E. 58th Street, CLSC 317

department: Human Genetics

institute: University of Chicago

summary: Quantification of gene expression levels at the single cell level has revealed that gene expression can vary substantially even across a population of homogeneous cells. However, it is currently unclear what genomic features control variation in gene expression levels, and whether common genetic variants may impact gene expression variation. Here, we take a genome-wide approach to identify expression variance quantitative trait loci (vQTLs). To this end, we generated single cell RNA-seq (scRNA-seq) data from induced pluripotent stem cells (iPSCs) derived from 53 Yoruba individuals. We collected data for a median of 95 cells per individual and a total of 5,447 single cells, and identified 241 mean expression QTLs (eQTLs) at 10% FDR, of which 82% replicate in bulk RNA-seq data from the same individuals. We further identified 14 vQTLs at 10% FDR, but demonstrate that these can also be explained as effects on mean expression. Our study suggests that dispersion QTLs (dQTLs), which could alter the variance of expression independently of the mean, have systematically smaller effect sizes than eQTLs. We estimate that at least 300 cells per individual and 400 individuals would be required to have modest power to detect the strongest dQTLs in iPSCs. These results will guide the design of future studies on understanding the genetic control of gene expression variance.

overall_design: The goal of our study was to identify quantitative trait loci associated with gene expression variance across cells (vQTLs). Using the Fluidigm C1 platform, we isolated and collected scRNA-seq from 7,585 single cells from induced pluripotent stem cell (iPSC) lines of 54 Yoruba in Ibadan, Nigeria (YRI) individuals. We used unique molecular identifiers (UMIs) to tag RNA molecules and account for amplification bias in the single cell data (Islam et al., 2014). To estimate technical confounding effects without requiring separate technical replicates, we used a mixed-individual plate study design. The key idea of this approach is that having observations from the same individual under different confounding effects and observations from different individuals under the same confounding effect allows us to distinguish the two sources of variation (Tung et al., 2017).

first: Abhishek

last: Sarkar

first: Po-Yuan

last: Tung

first: Yang

middle: I

last: Li

first: Matthew

last: Stephens

first: Yoav

last: Gilad


title: Gene expression of the bacterium Glutamicibacter arilaitensis when grown with Penicillium in cheese

status: Public on Apr 01 2018

city: Medford

first: Benjamin

last: Wolfe

address: Tufts University - Biology, 200 Boston Ave, Suite 4700

department: Biology

institute: Tufts University - Biology

summary: RNA-seq was used in combination with various analytical chemistry approaches to identify the chemical and genetic basis of pigment production of the bacterium Glutamicibacter arilaitensis when growing on cheese. This bacterium commonly found in cheese rinds where it co-occurs with Penicillium species and other molds. Pinkish-red pigments are produced by the bacterium in response to growth with Penicillium. Both chemical analyses and RNA-seq point to coproporphyrin III as the major metabolite leading to pigment formation.

overall_design: A total of 8 samples were analyzed. 4 biological replicates were prepared for each of the two treatments (Glutamicibacter arilaitensis growing alone and Glutamicibacter arilaitensis growing with Penicillium sp. #12)

middle: E


title: Centromeric chromatin in Koala

status: Public on Apr 26 2018

city: Storrs

last: Heider

address: 181 Auditorium Road, Unit 3197

institute: University of Connecticut

summary: We report high throughput sequencing of DNA associated with centromeric chromatin in the koala genome.

overall_design: Examination of 2 centromeric chromatin antibodies: CENP-A and CREST sera

first: Rachel

last: O'Neill


title: The Epstein Barr virus circleRNAome

status: Public on Jul 12 2018

city: New Orleans

first: Erik

last: Flemington

state: LA

address: 1430 Tulane Ave., SL79

department: Pathology

institute: Tulane Health Sciences Center

summary: Our appreciation for the extent of Epstein Barr virus (EBV) transcriptome complexity continues to grow through findings of EBV encoded microRNAs, new long non-coding RNAs, and hundreds of new polyadenylated lytic transcripts. Here we report an additional layer to the EBV transcriptome through the identification of a repertoire of latent and lytic viral circRNAs. Utilizing RNase R-sequencing with cell models representing latency types I, II, and III, we identified circRNAs expressed from the latency Cp promoter involving backsplicing from the W1 and W2 exons to the C1 exon, from the EBNA BamHI U exon, and from the latency long-non-coding RPMS1 locus. We also identified circRNAs expressed during reactivation including an exon 8-to-2 backspliced LMP2 transcript and a highly expressed circRNA derived from the BHLF1 gene. Altogether we identified over 30 EBV circRNA candidates and validated and determined the structural features, expression profiles and nuclear-cytoplasmic distributions of several prominent viral circRNAs. Further, we show that two RPMS1 circRNAs are expressed in stomach cancer specimens. This study increases the known EBV latency and lytic transcriptome repertoires to include viral circRNAs and provides an essential foundation for investigations into the functions of this new class of EBV transcripts in EBV biology and disease.

overall_design: RNase R treated, Ribodepletion, and Poly-A selection libraries of EBV+ B-cell lymphoma and gastric carcinoma cell lines were generated and RNA-sequencing was done.


title: Identification of LINC00473-regulated transcriptional program in human mucoepidermoid carcinoma

status: Public on Apr 18 2018

city: Gainesville

first: Lizi

last: Wu

state: FL

address: 2033 Mowry Road

institute: University of Florida

summary: Gene expression profiling was performed on control and long intergenic non-protein coding RNA 473 (LINC00473)-depleted human mucoepidermoid carcinoma H3118 cells, and differentially expressed genes after LINC00473 depletion were identified.

overall_design: There were two groups of H3118 cells: one transduced with pLKO.1 lentiviruses expressing scrambled shRNA control and the other transduced with pLKO.1 expressing LINC00473 shRNA. Two biological replicates for each group were included. Cells were independently cultured and RNAs were then harvested for microarray analysis.

first: Jian

middle: L


title: Postnatal DNA demethylation and its role in tissue maturation

status: Public on May 09 2018

city: Philadelphia

first: Klaus

last: Kaestner

state: PA

address: University of Pennsylvania, 12-126 Smilow Center for Translational Research, 3400 Civic Center Boulevard

department: Department of Genetics and Institute for Diabetes Obesity and Metabolism, Perelman School of Medicine

institute: University of Pennsylvania

summary: Development in mammals is accompanied by specific de novo and demethylation events that are thought to stabilize differentiated cell phenotypes. We demonstrate that large percentage of the tissue-specific methylation pattern is generated postnatally. Demethylation in the liver is observed in thousands of enhancer-like sequences associated with genes that undergo activation during the first few weeks of life. Using conditional gene ablation strategy we show that the removal of these methyl groups is stable and necessary for assuring proper hepatocyte gene expression and function through its effect on chromatin accessibility. These postnatal changes in methylation come about through exposure to hormone signaling. These results define the molecular rules of 5-methyl-cytosine regulation as an epigenetic mechanism underlying cellular responses to changing environment.

first: Yitzhak

last: Reizel

first: Ofra

last: Sabag

first: Adam

last: Spiro

first: Howard

last: Cedar


title: Investigation of the gamma-tubulin binding site in the nuclear and mitochondrial DNA of cells in S-phase

status: Public on Apr 04 2018

city: Malmö

first: Maria

middle: Alvarado

last: Kristensson

address: 59 Jan Waldenströms street

institute: Lund University

summary:
In a cell, g-tubulin establishes a cellular network of threads named g-string meshwork. However, the functions of this meshwork remain to be determined. We investigated the traits of the meshwork and show that g-strings have the ability to connect the cytoplasm and the mitochondrial DNA together. We also show that g-tubulin has a role in the maintenance of the mitochondrial meshwork and homeostasis as reduced levels of g-tubulin or impairment of its GTPase domain disrupts the mitochondria network and alters both their respiratory capacity and the expression of mitochondria-related genes. By contrast, reduced mitochondria number or increased protein levels of g-tubulin DNA-binding domain favour the association of g-tubulin with mitochondria.  Our results demonstrate that g-tubulin is an important mitochondrial structural component that maintains the mitochondria meshwork, providing mitochondria with a cellular infrastructure. We propose that g-tubulin provides a cytoskeletal element that maintains mitochondria homeostasis and gives form to the mitochondria meshwork.

Keywords: Genome binding/occupancy profiling by high throughput sequencing

overall_design:
The experiment was performed in an MCF10A cell line and a total number of 16 samples were included. There are two samples per timepoint (one and two hours) and a whole cell lysate (WCE) is included as a control for each sample.
Please note that processed data was generated from two samples (replicates *.1 and *.2) and is linked to the first replicate (i.e. *1.1, *2.1) of each pair.

last: Alvarado-Kristensson

first: Lisa

last: Lindström


title: microRNA expression patterns from humans with dialated cardiomyopathy

status: Public on Mar 31 2018

city: Changzhou

first: Lichan

last: Tao

address: Juqian Road 185, Changzhou, Jiangsu province, China

department: cardiology

institute: The third affilated hospital of Soochow University

summary: This study was to identify the expression patterns of miRNA in patients with dilated cardiomyopathy

overall_design: To compare expression patterns of miRNA in dilated cardiomyopathy patients and normal

first: Fei

last: Hua


title: Essential Nucleoid Associated Protein mIHF (Rv1388) controls virulence and housekeeping genes in Mycobacterium tuberculosis

status: Public on Feb 28 2018

city: Bern

first: Andrej

last: Benjak

address: Murtenstrasse 24-28

department: Department for BioMedical Research

country: Switzerland

institute: University of Bern


title: The effects of testosterone and lipopolysaccharide on gene expression in immortalized ocular surface and adnexal cells

status: Public on Mar 07 2018

city: Boston

first: David

middle: A.

last: Sullivan

address: 20 Staniford St.

institute: Schepens Eye Research Institute

summary: Immortalized human corneal, conjunctival, and meibomian gland epithelial cells were pre-treated with dihydrotesteosterone (DHT) or vehicle, then exposed to lipopolysaccharide (LPS) and ligand binding protein (LBP) or vehicle, lysed, and subjected to microarray analysis of RNA expression.

overall_design:
Cells were cultured in keratinocyte serum-free medium (KSFM) supplemented with bovine pituitary extract (BPE; 25 ug/mL HCEC and HCjE, 50 ug/mL HMGEC), 5 ng/mL epidermal growth factor (EGF), penicillin and streptomycin. On reaching confluence, cells were rinsed twice with a phosphate-buffered saline solution and exposed to a stratification medium consisting of DMEM and F12 (Mediatech, Inc) with 10% fetal bovine serum, epidermal growth factor (10 ng/mL), penicillin, and streptomycin for 2 days. After this time period, cells were incubated in serum-free DMEM/F12 and exposed to vehicle (1% bovine serum albumin [BSA]), or LPS (15 μg/ml) and LBP (150 ng/ml), for six hours. The LPS and LBP were dissolved in DMEM and the BSA in PBS.
For androgen-related studies, the stratification medium contained 10% charcoal and dextran-treated fetal bovine serum with ethanol or DHT (10nM); the poststratification medium also contained ethanol or DHT.

middle: A

first: Afsun

last: Sahin

first: Raheleh

last: Darabad


title: Impact of mIHF on gene expression

summary: mIHF is a possible nucleoid associated protein. Its effect on gene regulation remains unknown. We performed an RNA-seq analysis of the mihF conditional knock-down mutant in Mycobacterium tuberculosis. We show that mihF is essential for in vitro growth and that depeltion of mIHF results in deregulation of a large number of genes. Notably, the most downregulated gene was the espACD operon, which is necessary for secretion of M. tuberculosis main virulence factor EsxA.

overall_design: The mihF conditional knock-down (mihF-cKD) has an anhydrotetracycline (ATc)-inducible promoter. The comparison was done between the presence and the absence of ATc in the growing medium of mihF-cKD, i.e. exprssed vs. non-expressed mihF. To check for the possible influence of ATc alone on gene expression, presence and absence of ATc in the growing medium was compared in a wild-type strain. All conditions were repeated twice.

first: Nina

middle: T

last: Odermatt

first: Claudia

last: Sala

first: Stewart

last: Cole


title: Chromosomal DNA binding of mIHF

summary: mIHF is a possible nucleoid associated protein, but its DNA binding characteristics are unknown. To address this, we performed a ChIP-seq analysis of mIHF in Mycobacterium tuberculosis.

overall_design: Monoclonal anti-mIHF (Alere) was used for the mIHF ChIP-Seq experiment. To check that the technique works, and for comparative analysis, we reproduced the experiment from Blasco et al. 2012 (PMID: 22479184), using polyclonal rat anti-EspR antibodies. Input control was not incubated with any antibody. For mIHF, two different concentrations of antibodies were tested. Spearman correlation of the resulting binding sites was 0.96, so the two datasets were pooled for the final peak calling.


title: Xrn2 accelerates termination by RNA polymerase II, which is underpinned by CPSF73 activity

status: Public on Feb 13 2018

city: Exeter

first: Steven

last: West

state: Devon

address: Stocker Rd

department: Living Systems Institute

country: United Kingdom

institute: University of Exeter

summary: Termination is a ubiquitous phase in every transcription cycle but is incompletely understood and a subject of debate.  We have used gene editing as a new approach to address its mechanism through engineered conditional depletion of the 5’-3’ exonuclease, Xrn2, or the polyadenylation signal (PAS) endonuclease, CPSF73.  The ability to rapidly control Xrn2 reveals a clear and general role for it in co-transcriptional degradation of 3’ flanking region RNA and transcriptional termination.  This defect is characterised genome-wide, at high resolution, using native elongating transcript sequencing (mNET-seq).  An Xrn2 effect on termination requires prior RNA cleavage and we provide evidence for this by showing that catalytically inactive CPSF73 cannot restore termination to cells lacking functional CPSF73.  Notably, Xrn2 plays no significant role in either Histone or snRNA gene termination even though both RNA classes undergo 3’ end cleavage.  In sum, efficient termination on most protein-coding genes involves CPSF73 mediated RNA cleavage and co-transcriptional degradation of polymerase-associated RNA by Xrn2.   However, as CPSF73 loss caused more extensive read-through transcription than Xrn2 elimination, it likely plays a more underpinning role in termination.

overall_design: We performed total CTD mNET-seq and nuclear RNA-seq with auxin-inducible degron(AID) tagged cell lines treated with or without auxin to induce rapid depletion of XRN2. Cell lines ectopically express TIR1 from a sleeping beauty transposon inserted cassette. AID tagged proteins contain the inframe insertion of AID tag, P2A cleavage site, antibiotic resistance marker and SV40 polyA site. Two repair templates containing different antibiotic resistance markers (hygromycin and neomycin) were used to ensure biallelic insertion of the cassette by CRISPR/Cas9.

first: Joshua

last: Eaton

first: Lee

last: Davidson

last: Bauer

first: Toyoaki

last: Natsume

first: Masato

last: Kanemaki

first: Steve


title: Systemic post-translational control of bacterial metabolism regulates adaptation in dynamic environments

status: Public on Feb 02 2018

city: La Jolla

first: Nathan

last: Lewis

state: California

address: 9500 Gilman Drive

department: Pediatrics

institute: University of California, San Diego

summary: E. coli K-12 MG1655 was grown on M9 minimal media supplemented with 0.2% w/v glucose, fructose, or acetate. The goal of this study was to determine differentially expressed genes in the different media conditions.

overall_design: Gene expression profiling data were generated from cultures of exponentially-growing E. coli under aerobic conditions on glucose, fructose, or acetate M9 minimal media. E. coli K12 MG1655 was grown and expression profiled on 0.2% (w/v) glucose M9 minimal media or 0.2% (w/v) fructose M9 minimal media or 0.2% (w/v) acetate M9 minimal media at 37°C. Expression profiling was done using Affymetrix E. coli Genome 2 Arrays. Each experimental condition was tested in triplicate in the respective carbon sources using independent cultures and processed following the manufacturer-recommended protocols. Cultures were grown to mid-exponential growth phase aerobically (OD600 = 0.3) in minimal media, supplemented with the appropriate carbon source. Three ml of culture was added to 2 volumes of RNAprotect Bacteria Reagent (Qiagen) and total RNA was then isolated using RNeasy columns (Qiagen) with DNase I treatment. cDNA synthesis, fragmentation, end-terminus biotin labeling, and array hybridization were performed as recommended by the Affymetrix standard protocol.

first: Elizabeth

last: Brunk

first: Roger

last: Chang

first: Jing

last: Xia

first: Hooman

last: Hefzi

first: James

last: Yurkovich

first: Donghyuk

last: Kim

first: Evan

last: Buckmiller

first: Harris

last: Wang

first: Byung-Kwan

last: Cho

first: Chen

last: Yang

first: Bernhard

middle: O

last: Palsson

first: George

last: Church


title: In-vitro stimulation of healthy donor blood with IL-3 cytokine

city: Parkville

first: CSL

last: Research

state: VIC

address: 30 Flemington Rd

country: Australia

institute: CSL Limited / bio21 Institute

summary: This experiment was designed to look for in vitro IL-3 gene signature in donor blood at two different time points (6 and 24 hours). RNA from lysed whole blood cells was used for the sequencing.

overall_design: Lysed whole blood from seven healthy donors was stimulated with recombinant human IL-3 for 6 hours, or 24 hours, prior to RNA extraction for next-generation sequencing on the Illumina HiSeq platform. Unstimulated samples were included as controls.

first: Shereen

last: Oon

first: Katherine

last: Monaghan

first: Milica

last: Ng

first: Alberta

last: Hoi

first: Eric

last: Morand

first: Gino

last: Vairo

first: Eugene

last: Maraskovsky

first: Andrew

last: Nash

first: Ian

middle: P

last: Wicks

first: Nicholas

middle: J

last: Wilson


title: Molecular Mechanisms of the Progression of Cisplatin-Induced Acute Kidney Injury to Chronic Kidney Disease

status: Public on Dec 13 2017

city: New Haven

first: Robert

last: Safirstein

state: Connecticut

address: 333 Cedar Street

department: Internal Medicine (Nephrology)

institute: Yale University School of Medicine

summary: Though there has been extensive investigation of the kidney's acute cellular and molecular responses following cisplatin treatment, the mechanisms of progression from acute to chronic disease have not been explored. In this study, we use functional and morphological metrics to establish a time point when the transition from acute repairable kidney injury to chronic irreparable disease is clearly established. We then used microarray and western blot analysis to investigate the molecular changes (i.e., gene expression, pathway activity, signaling) associated with the transition from acute to chronic cisplatin-induced kidney disease.

overall_design: In the study presented here we compared the molecular profiles of kidneys obtained from C57BL/6 mice belonging to one of three treatment groups: control (no cisplatin), one dose of cisplatin, and two doses of cisplatin. Tissue from control and single dose mice was harvested two weeks following cisplatin treatment. Two dose animals received two doses of cisplatin administered two weeks apart. Tissue from these mice was harvested two weeks after a second dose of cisplatin (four weeks after the first cisplatin dose). Each time point was comprised of a group of three mice. Comparison of gene expression of single and dose dose mice compared with control mice was used to generate a hypothesis for potential pathways driving the progression from acute to chronic kidney disease. Microarray analysis was performed by Phalanx Biotech Group Inc. using Phalanx Mouse OneArray® v2 technology (MOA 2.1).

first: Sarah

last: Landau

first: Xiaojia

last: Guo

first: Heino

last: Velazquez

first: Richard

last: Torres

first: Eben

last: Olson

first: Rolando

last: Garcia-Milian

first: Gilbert

last: Moeckel

first: Gary

middle: V

last: Desir


title: AR Expression in Breast Cancer CTCs Associates with Bone Metastases

status: Public on Feb 19 2018

first: Ben

middle: S.

last: Wittner

state: MA

address: 149 13th Street

department: Center for Cancer Research

institute: Massachusetts General Hospital

summary: Molecular drivers underlying bone metastases in human cancer are not well understood, in part due to constraints in bone tissue sampling. Here, we undertook RNA sequencing of circulating tumor cells (CTCs) obtained from blood samples of women with metastatic estrogen receptor (ER)+ breast cancer, comparing cases with progression in bone versus visceral organs. Among the activated cellular pathways in CTCs from bone-predominant breast cancer is Androgen Receptor (AR) signaling. AR gene expression is evident, as is its constitutively active splicing variant AR-v7. AR expression within CTCs is correlated with the duration of treatment with aromatase inhibitors, suggesting that it may contribute to acquired resistance to anti-estrogen therapy. In an established breast cancer xenograft model, a bone-tropic derivative displays increased AR expression, whose genetic or pharmacologic suppression reduces metastases to bone but not to lungs. Together, these observations identify AR signaling in CTCs from women with bone-predominant ER+ breast cancer, and provide a rationale for testing androgen inhibitors in this subset of patients.

overall_design: Thirty-two women with metastatic ER-positive breast cancer were enrolled in an IRB-approved observational study at Massachusetts General Hospital (MGH). All patients had clinical evidence of disease progression, and had discontinued their therapy at the time of CTC collection, in anticipation of starting an experimental early phase drug trial (“drug washout period”). Patients with detectable CTCs (n=22) were classified dynamically based on which metastatic sites were progressing at the time of blood sampling, identifying 13 (59%) who had the appearance of new bone metastases at the time of CTC isolation (termed “Bone (+)”), versus 9 (41%) who had either no bone involvement at all or had no new bone metastases, despite having disease progression at other soft tissue or visceral sites (termed “Visceral (+)”).   For each patient, approximately 10ml of blood was processed through the CTC-iChip, which first uses microfluidic size-based fractionation to separate nucleated cells from platelets and red blood cells (RBCs), and then effectively depletes magnetically-labeled white blood cells (WBCs) from untagged CTCs. From the initial sample of blood (10 x 106 WBC/ml), the final microfluidic product contains approximately 500 residual WBCs per ml of whole blood processed, along with candidate CTCs. Unfixed CTCs are identified by live staining with Alexa 488-conjugated antibodies against the cell surface markers EpCAM (epithelial-specific), HER2 (lineage-specific) and CDH11 (mesenchymal-specific), while contaminating WBCs are counterstained using TexasRed-conjugated antibodies against the hematopoietic lineage-specific markers CD45, CD16 and CD14. Marker-positive single and clustered CTCs were identified in 13 patients with Bone (+) and 9 patients with Visceral (+) disease, with a median of 3.5 CTCs per patient, leading to a total of 77 samples representing either single CTC, CTC-clusters, or in rare patients with very high numbers of CTCs, pools of up to three CTCs. RNA from these CTCs was then extracted, reverse transcribed, amplified and sequenced as described in PubMed ID 25171411.

first: Nicola

last: Aceto

first: Aditya

last: Bardia

middle: S

first: Maria

middle: C

last: Donaldson

first: Ryan

last: O’Keefe

first: Amanda

last: Engstrom

first: Francesca

last: Bersani

first: Yu

last: Zheng

first: Valentine

last: Comaills

first: Huili

last: Zhu

first: Olivia

last: Mackenzie

first: Toshi

last: Shioda

first: Dennis

last: Sgroi

first: Ravi

last: Kapur

first: David

last: Ting

first: Sridhar

last: Ramaswamy

first: Mehmet

last: Toner

first: Daniel

last: Haber

first: Shyamala

last: Maheswaran


title: Transcriptome-wide response to synthetic chromatin protein PcTF: Breast Cancer

status: Public on Jan 31 2018

city: Tempe

first: Karmella

last: Haynes

state: Arizona

address: 427 E Tyler Mall

department: Biological and Health Systems Engineering

institute: Arizona State University

summary: The goal of this study was to identify genes that become upregulated in response to a synthetic transcription factor, PcTF, in a panel of breast cancer model cell lines. PcTF contains a conserved H3K27me3-binding chromodomain expressed in-frame with an mCherry fluorescent tag and a VP64 activation domain. Therefore, we expected genes with H3K27me3-associated promoters to become upregulated upon PcTF expression. We observed that hundreds of genes became up or downregulated.

overall_design: MCF7, BT474, or BT549 cells were transfected with a plasmid that constitutively expressed the PcTF protein. 24, 48, and 72 hrs post-transfection, total mRNA was extracted from treated and untreated cells, as well as from untransfected MCF10A cells.

first: Melissa

last: Wilson Sayres

first: Kimberly

last: Olney

middle: B

last: Nyer


title: Allele-specific mRNA expression experiment in lactis-wickerhamii and lactis-marxianus interespecies hybrids

status: Public on Dec 22 2017

city: San Francisco

first: Trevor

last: Sorrells

state: CA

department: Biochemistry and Biophysics

institute: University of California, San Francisco

summary: This experiment was performed to measure differences in pathway-level evolution of cis-regulation between closely related Kluyveromyces species. Sequencing of mRNA as well as DNA was performed in hybrids of Kl. lactis with two other species, Kl. wickerhamii and Kl. marxianus. Cells were grown overnight for DNA sequencing and for 4-8h to an OD600=0.7-1.0 for mRNA sequencing. DNA and mRNA reads were mapped to hybrid genomes, multimapping reads discarded, and allele-specific expression ratio for each gene was calculated after first normalizing mRNA reads to the number of DNA reads for each gene in each species. We found that the ribosomal protein genes showed pathway-level allele-specific expression differences between species in both interspecies hybrids.

sample_organism: Kluyveromyces lactis x Kluyveromyces wickerhamii

sample_organism: Kluyveromyces lactis x Kluyveromyces marxianus

overall_design: Examination of allele-specific expression in two interspecies hybrids

middle: R


title: CD8 T cell phenotype following antigen education by lymphatic endothelial cells versus mature dendritic cells

status: Public on Dec 31 2017

first: Shann

last: Yu

address: 5640 S Ellis Ave, Eckhardt Research Center

department: PME-Immunoengineering Center

institute: University of Chicago

summary:
The goal of this study is to analyze differences in gene expression of CD8 T cells that are antigen-educated by lymphatic stromal cells versus traditional antigen-presenting cells. Naive antigen-specific CD8 T cells reactive against the model antigen ovalbumin (OVA) were isolated from spleens of OT-I transgenic mice. They were then co-cultured with OVA-pulsed mature dendritic cells (mDCs) versus lymphatic endothelial cells (LECs) for up to 3d. Each day, OT-I cells were isolated and prepared for RNAseq analysis to determine differences in gene expression as a function of antigen-presenting cell. Naive, LEC-educated, or DC-educated OT-I transcriptomes are provided here.

Grant ID - AdG-323053
Agency - European Research Council (ERC)
Grant Title - LYMPHIMMUNE - Flow in the tumor microenvironment: Linking mechanobiology with immunology

Grant ID - R01CA219304
Agency - US National Institutes of Health (NIH)
Grant Title - Paradoxical roles of tumor lymphangiogenesis on tumor immunity and implications for immunotherapy

first: Efthymia

last: Vokali

first: Melody

last: Swartz


title: Altered molecular signatures during kidney development after intrauterine growth restriction of different origins

status: Public on Dec 09 2017

city: Cologne

first: Kai

middle: Dietrich

last: Nüsken

state: NRW

address: Kerpener Str. 62

department: Department of Pediatrics

institute: University of Cologne, Medical Faculty

summary: Intrauterine growth restriction (IUGR) has been linked to predisposition towards an unfavorable course of glomerulopathies and early loss of kidney function. This study was performed to identify transcriptional alterations in male IUGR rats during and at the end of nephrogenesis in order to generate hypotheses which molecular mechanisms contribute to adverse kidney programming. IUGR was induced by low protein (LP) diet throughout pregnancy, bilateral uterine vessel ligation (LIG), or intrauterine stress (IUS) by sham operation. Offspring of unimpaired dams served as controls. Significant acute kidney damage was ruled out by negative results for proteins indicative of ER-stress, autophagy, apoptosis or infiltration with macrophages. Renal gene expression was examined by transcriptome microarrays, demonstrating 53 (LP, n=12; LIG, n=32; IUS, n=9) and 134 (LP, n=10; LIG, n=41; IUS, n=83) differentially expressed transcripts on postnatal days (PND) 1 and 7, respectively. Reduced Pilra (all IUGR groups, PND 7), Nupr1 (LP and LIG, PND 7) and Kap (LIG, PND 1) as well as increased Ccl20, S100a8/a9 (LIG, PND 1), Ifna4 and Ltb4r2 (IUS, PND 7) indicated that inflammation-related molecular dysregulation could be a “common” feature after IUGR of different origins. Network analyses of transcripts and predicted upstream regulators hinted at proinflammatory adaptions mainly in LIG (arachidonic acid-binding, neutrophil aggregation, toll-like-receptor, NF-kappa B and TNF signaling) and dysregulation of AMPK and PPAR signaling in LP pups. The latter may increase susceptibility towards obesity-associated kidney damage. Western blots of the most prominent predicted upstream regulators confirmed significant dysregulation of RICTOR in LP (PND 7) and LIG pups (PND 1), suggesting that mTOR-related processes could further modulate kidney programming in these groups of IUGR pups.

overall_design:
Direct comparison of kidneys during early postnatal life from IUGR offspring after (1) low protein (LP) diet throughout pregnancy, (2) bilateral uterine vessel ligation (LIG) during terminal pregnancy and (3) intrauterine stress (IUS) by sham operation in late pregnancy with a common control (C) group offers the unique opportunity to differentiate the details of early kidney programming after reduced uterine blood flow from alterations induced by intrauterine stress or nutritional deficiency in a critical window of kidney development. The models have been thoroughly validated in our laboratory with a special focus on translational relevance. In detail, Wistar rat dams (Janvier Labs, France) were time-mated [presence of sperm plug = gestational day (GD) 0] and either fed a defined diet (no. C1000, Altromin, Germany) containing 17.6% protein throughout pregnancy in groups LIG, IUS and C, or a diet containing 8.1% protein (no. C1003, Altromin) in group LP as described before [16,19]. In LIG dams (n=19), fetuses were counted intraoperatively after midline laparotomy and careful extra-abdominal positioning of both uterine horns on GD 18 under isoflurane anesthesia and metamizol analgesia. Consecutively, bilateral uterine vessel ligation was performed caudally of the most caudal vessel branch running to the most caudal feto-placental unit. The duration of the complete intervention was 15 – 20 minutes. The size of LIG-litters was 9 – 15 living fetuses, all uterine horns (n=38) contained 4 – 9 living fetuses. After ligation, blood flow to the LIG fetuses is reduced, as blood now is supplied from cranially via the ovarian arteries only. In IUS dams (n=13), the suture material was not fixed but removed after identical procedures. The size of IUS-litters was 10 – 16 living fetuses, all uterine horns (n=26) contained 3 – 10 living fetuses. After surgery, all dams started to drink water (containing tramadol for 24h after surgery) after 5 – 10 minutes, and their behavior recovered fully within 1 hour. Dams of groups C (n=19) and LP (n=12) did not undergo surgery. All dams delivered spontaneously after approximately 22.5 gestational days within a time frame of 12h. Two C litters (litter size was <9, but had to be in the range of 9 – 16 to reduce bias due to varying litter size) and one LIG litter (11 out of 13 fetuses were resorbed after LIG) were excluded on PND 1. Thus, we finally included 17 C litters, 18 LIG litters, 13 IUS litters and 12 LP litters (n=60 litters in total). All offspring [C, n=232; LP, n=152; LIG, n=167 born alive out of n=209 counted intraoperatively, i.e. n=42 (20%) resorbed; IUS, n=174 born alive out of 174 counted intraoperatively) were weighed and measured on PND 1, and sexed visually. The error of this sexing method is ~1% in our laboratory on PND 1. In LIG, only the six smallest pups per litter (n=108 out of 167) were selected since we know from thorough intrauterine evaluation that especially the fetuses near to the ligation suffer from insufficient blood and nutrient supply and display intrauterine growth restriction.  As there is sex-specific transcriptional regulation and male rodents seem to be more susceptible to renal programming than females, we focused on male pups in all further analyses (PND 1 male pups: C, n=105; LP, n=70; LIG, n=52 small; IUS, n=99; PND 1 kidneys: C, n=27; LP, n=17; LIG, n=21; IUS, n=24; PND 7 pups and kidneys: C, n=24; LP, n=16; LIG, n=14; IUS, n=17).
For PND 1 studies, newborn male rats were narcotized with pentobarbital and decapitated. Then, whole kidneys as well as further organs for studies on brain and metabolism were obtained and weighed. Right kidneys were shock frozen in liquid nitrogen and stored at  80°C, left kidneys dissected longitudinally in the middle, directly fixed in 4% PFA for 24h, transferred in 70% isopropyl alcohol for 24h, and processed for paraffin embedding.
For PND 7 studies, postnatal environmental conditions were standardized in all groups on the first day of life, because litter size and postnatal nutrition are known to affect the final stages of renal development. In detail, pups from C, LIG and IUS dams were transferred to non-operated dams from group C receiving control diet (C1000, Altromin) and the size of all foster litters was adjusted to four male and four female pups to ensure similar postnatal nutrition. LP pups (four male, four female) were transferred to LP foster dams as postnatal renal programming effects are in part mediated by altered milk quality in this model. On PND 7, kidneys and further organs were harvested and stored as described above.

first: Eva

first: Gregor

last: Fink

first: Felix

last: Lechner

first: Jenny

last: Voggel

first: Maria

last: Wohlfarth

first: Lisa

last: Sprenger

first: Nava

last: Mehdiani

first: Lutz

last: Weber

first: Max

last: Liebau

first: Bent

last: Brachvogel

first: Jörg

last: Dötsch

middle: D


title: NFIA regulates pancreatic cell fate and adult physiology through vesicle trafficking

status: Public on Sep 14 2017

city: Houston

first: Malgorzata

last: Borowiak

address: 1 Baylor Plaza

institute: Baylor College of Medicine

summary: Intracellular trafficking is essential for proper cell signaling. In the pancreas, secretory cells rely on trafficking to regulate blood glucose and digestion. Pancreatic disorders reflect defects in function or development, evoking considerable interest in understanding the molecular genetics governing pancreatic organogenesis. Here, we show the transcription factor NFIA regulates trafficking in both the embryonic and adult pancreas, affecting both developmental cell fate decisions and adult physiology. NFIA deletion from pancreatic progenitors led to the development of more acinar cells and ducts and fewer endocrine cells, whereas ectopic NFIA promoted endocrine formation. We found that NFIA’s effects on trafficking influence endocrine/exocrine cell fate decisions through regulation of Notch. Adult NFIA-deficient mice develop diabetic phenotypes due to impaired insulin granule trafficking and defects in acinar zymogen secretion. This study shows how a single transcription factor, NFIA, thus exerts profound effects on both embryonic cell fate and adult physiology by regulating vesicle trafficking.

overall_design: 2 control and 2 NFIA fl/fl; Pdx1-cre samples, from pooled embryonic litters at E17.5

first: Marissa

last: Scavuzzo

first: Jolanta

last: Chmieloweic

first: Diane

last: Yang


title: Neisseria gonorrhoeae with interrupted VSP pathway genes (ngoAXIII, ngoAXIV, mutS, mutL) versus wild type

status: Public on Sep 25 2017

city: Warsaw

first: Monika

middle: Julita

last: Adamczyk-Poplawska

address: Miecznikowa 1

institute: University of Warsaw

summary: Transcriptional profiling of N. gonorrhoeae comparing wild type cells to cells with inactivated by kanamycin cassette (km) genes involved in VSP pathway (ngoAXIII, ngoAXIV, mutS, mutL). The Goal was to study the role of these genes in overall expression profile.

overall_design: Two-condition experiment, wild type cells vs. mutant cells. Biological replicates: 4 control replicates, 4 mutant replicates.

first: Agnieszka

last: Kwiatek

last: Adamczyk-Popławska


title: Transcriptome analysis of single cells from the mouse dentate gyrus [C1]

city: Stockholm

first: Sten

last: Linnarsson

address: Scheeles väg 1

department: Medical Biochemistry and Biophysics

institute: Karolinska Institutet

summary: RNA sequencing was performed on 2303 single cells from dentate gyrus of CD-1 and hGFAP-GFP reporter mice, sampled at 17 time points from P8 to P68, with the aim of delineating the distinct cellular states along the granule cell lineage.

overall_design: 2140 single cells from the dentate gyrus of CD-1 mice and 163 from hGFAP-GFP expressing mice.

first: Hannah

last: Hochgerner

first: Amit

last: Zeisel

first: Patrik

last: Ernfors


title: Genome wide DNA methylation study of Parkinson's disease in whole blood samples

status: Public on Mar 10 2018

city: Los Angeles

first: Ritz

last: Beate

address: 650 Charles E Young Dr. South

department: Epidemiology

institute: UCLA Fielding School of Public Health

summary: Genome wide DNA methylation profiling of whole blood and saliva samples in Parkinson's disease (PD) patients and PD-free controls. The Illumina Infinium 450k Human DNA methylation Beadchip was used to obtain DNA methylation profiles. Samples included 335 PD patients and 237 controls with blood DNA.

overall_design: Bisulphite converted DNA from the 572 samples were hybridised to the Illumina Infinium 450k Human Methylation Beadchip

first: Beate

last: Ritz

first: Steve

last: Horvath


title: Culturing cells in human serum: differentiation and metabolic reprogramming in human hepatoma cells

status: Public on Oct 01 2017

city: Edmonton

first: Rineke

last: Steenbergen

state: Alberta

address: 6-0096 Katz Building

department: Medical Microbiology and Immunology

institute: University of Alberta

summary:
Huh7.5 cells, which are cultured in human serum containing media instead of FBS containing media, undergo growth arrest and become differentiated. In this study we further characterized human serum cultured Huh7.5 cells, particularly with regards to morphology, metabolic profile and hepatocyte specific functions.
Similar to primary hepatocytes, HS-cultured Huh7.5 cells become columnar, are polarized, and develop bile canalicular structures on the apical side of the cells. Further analysis by microarray showed that significant transcriptional changes occur in 22-32% of the genes. HS cultured Huh7.5 cells undergo extensive metabolic re-programming. We showed a reversal of a Warburg-like metabolic profile that is typical for proliferating cells. In HS cultured cells large lipid droplets can be found in addition to large glycogen stores. Metabolic modeling shows an increased reliance on beta-oxidation, which was confirmed by an increase in ketone body production, the end products of β-oxidation. Additionally we showed increased conversion of glucose to glycogen. Bile secretion is increased, as are the enzymes involved in the degradation of xenobiotics.
Overall these data show that only changing the serum in cell culture media can result in the establishment of a cell type that morphologically, metabolically and functionally resemble primary hepatocytes.

overall_design: Samples were analyzed for Huh7.5 cells cultured in fetal bovine serum as well as at different time points – 8 days, 15 days and 23 days – after being transferred to human serum. Biological triplicates were used in each case.

first: D

middle: L

last: Tyrrell