title: Ribosome Profiling of Mouse Embryonic Stem Cells Reveals the Complexity of Mammalian Proteomes

status: Public on Nov 03 2011

first: Nicholas

last: Ingolia

email: ingolia@berkeley.edu, nick@ingolia.org

state: CA

address: 1 Barker Hall # 3202

department: Molecular and Cell Biology

institute: University of California, Berkeley

summary: The ability to sequence genomes has far outstripped approaches for deciphering the  information they encode. Here we present a suite of techniques, based on ribosome profiling (the deep-sequencing of ribosome-protected mRNA fragments), to provide genome-wide maps of protein  synthesis as well as a pulse-chase strategy for determining rates of translation elongation. We exploit  the propensity of harringtonine to cause ribosomes to accumulate at sites of translation initiation  together with a machine learning algorithm to define protein products systematically. Analysis of  translation in mouse embryonic stem cells reveals thousands of strong pause sites and novel  translation products. These include amino-terminal extensions and truncations and upstream open  reading frames with regulatory potential, initiated at both AUG and non-AUG codons, whose  translation changes after differentiation. We also define a new class of short, polycistronic ribosome-associated coding RNAs (sprcRNAs) that encode small proteins. Our studies reveal an unanticipated  complexity to mammalian proteomes.

overall_design: Examination of translation in mouse embryonic stem cells and during differentiation into embryoid bodies

first: Liana

middle: F

last: Lareau

first: Jonathan

last: Weissman


title: Host influence on in vivo gene expression profiles of Plasmodium yoelii blood-stage malaria parasites

status: Public on Nov 01 2011

city: Philadelphia

first: James

middle: M.

last: Burns

state: PA

address: 2900 Queen Lane

department: Microbiology and Immunology

country: USA

institute: Drexel University College of Medicine

summary: Microarray studies using synchronized Plasmodium falciparum parasites have revealed a ‘continuous cascade’ of gene expression.  Reports vary regarding the stability in these transcriptional patterns in the presence of external stressors.  Using Plasmodium yoelii 17X parasites replicating in vivo, we have examined differential gene expression in parasites isolated from individual mice, from independent infections, during ascending and peak parasitemia and in the presence and absence of host antibody responses.  Across experimental conditions, transcription was surprisingly stable.  Differential gene expression was greatest when comparing differences due to parasite load and/or host cell availability; however, even these changes were modest.  Of genes that were differentially expressed, many are of unknown function.  There was little to no differential expression of members of the yir and pyst-a multigene families, although a relatively large number of these were expressed during blood-stage infection regardless of experimental condition. Taken together, these results indicate that 1) P. yoelii gene expression remains stable in the presence of a changing host environment and 2) concurrent expression of a large number of the yir and pyst-a genes may function to divert host immune responses away from invariant protective antigens.

overall_design:
1.  P. yoelii 17X gene expression profiles were determined in blood-stage parasites isolated from infected Balb/c mice  (male, 8-12 weeks old) as follows:
a.  Mouse 1 (M1), Mouse 2 (M2), Mouse 3 (M3), Donor (D) mouse – three mice simultaneously infected with P. yoelii 17X iRBCs from a single donor mouse.  Parasite RNA was isolated early during infection when parasitemia was ascending as follows: M1 - day 11 at 13.0% parasitemia; M2 - day 10 at 11.1% parasitemia; M3 - day 10 at 18.6% parasitemia; D – day 11 at 18.7% parasitemia.
b.  Infection #1 (I1), infection #2 (I2), infection #3 (I3), infection #4 (I4) – four groups of mice infected on four separate occasions using P. yoelii 17X iRBCs obtained from four separate donor mice.  In each case, P. yoelii 17X RNA was isolated from iRBCs obtained on days 10-11 of infection when parasitemia was ascending and was ~15%.
c.  Day 10 (D10), Day 14 (D14) – P. yoelii 17X RNA isolated from iRBCs during infection #2 on day 10 (14% parasitemia, 14.4% reticulocytes) and on day 14 of infection #2 (43.5% parasitemia, 57.9% reticulocytes).
d.  WT and JHD - P. yoelii 17X RNA isolated from iRBCs during infection #4 from immunologically intact, wild-type Balb/c mice and B cell deficient, JHD mice on a Balb/c background (ascending infection, 15% parasitemia).
e.  QA and RMP - P. yoelii 17X RNA isolated from iRBCs during infection #1 from mice previously immunized with a preparation of membrane proteins isolated from P. yoelii 17X infected reticulocytes (PyRMP) (day 12, 13.3% parasitemia and Quil A immunized (QA) control mice (day 10, 15.6% parasitemia)
2.  On each array, gene expression in P. yoelii 17X blood-stage parasites was evaluated relative to a standard comparator of purified P. yoelii 17XL total RNA (pooled RNA from 45 mice, mean parasitemia ~35%).  Each sample was analyzed on three replicate arrays, one of which was a standard dye-flip.  One exception was the infection #3 sample which was analyzed on only on two arrays.  On each array, oligonucleotides were spotted in duplicate.
3.  Through the use of the standard reference RNA, the following 15 pairwise comparisons across samples were made:
(M1 vs M2); (M2 vs M3); (M1 vs M3); (D vs M1); (D vs M2); (D vs M3); (I1 vs I2); (I1 vs I3); (I1 vs I4); (I2 vs I3); (I2 vs I4); (I3 vs I4); (D10 vs D14); (WT vs JHD); (QA vs RMP)

first: Amy

last: Cernetich-Ott

first: Thomas

middle: M

last: Daly

first: Akhil

middle: B

last: Vaidya

first: Lawrence

middle: W

last: Bergman


title: A profibrotic gene expression program induced by Egr-1 in skin fibroblasts

status: Public on Oct 26 2011

city: Chicago

first: Swati

last: Bhattacharyya

state: IL

address: 240 E Huron st, McGAW, M230

department: Medicine

institute: Northwestern University

summary: Systemic sclerosis (SSc) is characterized by vascular damage, autoimmunity and fibrosis and is associated with highly variable clinical presentation and disease course. Aberrant transforming growth factor-ß (TGF-ß) signaling via the early immediate transcription factor Egr-1 is implicated in the pathogenesis of SSc. To shed light on the role of Egr-1 in fibrosis, regulation of  gene expression in human skin fibroblasts overexpressing Egr-1 was examined by genome-wide expression analysis. Over 600 genes were found to be regulated by Egr-1. The Egr-1-responsive gene signature is largely comprised of genes involved in cell proliferation, TGF-ß signaling, wound healing, extracellular matrix synthesis and vascular development. Expression of the Egr-1 responsive genes was evaluated in a microarray dataset comprising skin biopsies from 17 patients with scleroderma and six healthy controls (GEO GSE9285; PMID 18648520). The “Egr-1 responsive gene signature” was enriched in the ‘diffuse-proliferation’ subset of skin biopsies in the patients with diffuse cutaneous SSc (dcSSc), but was not associated with other forms of scleroderma, or with healthy controls. Skin biopsies from patients with scleroderma can provide more insights into the relevant pathological processes in the subset of disease and could be developed into a diagnostic tool for identifying a subset of diffuse scleroderma patients who may be responsive to Egr-1 therapy.

overall_design: Cultures of primary fibroblasts from neonatal foreskin treated by Egr1 and Tgfb1 were measured at 24 and 48 hours. Control samples without any treatment were also measured at the same time points. Two biological replicates per condition/time point were measured using the Illumina HumanRef-8 V2 Expression BeadChip.


title: Gene expression from mouse white, brown, and perivascular adipose tissue

status: Public on Aug 23 2011

city: Worcester

first: Timothy

middle: P

last: Fitzgibbons

address: 55 Lake Avenue North

department: Molecular Medicine

institute: University of Massachusetts Medical School

summary: Thoracic perivascular adipose tissue (PVAT) is a unique adipose depot that likely influences vascular function and susceptibility to pathogenesis in obesity and metabolic syndrome. Surprisingly, PVAT has been reported to share characteristics of both brown and white adipose, but a detailed direct comparison to interscapular brown adipose tissue (BAT) has not been performed. Here we show by full genome DNA microarray analysis that global gene expression profiles of PVAT are virtually identical to BAT, with equally high expression of Ucp-1, Cidea and other genes known to be uniquely or very highly expressed in BAT. PVAT and BAT also displayed nearly identical phenotypes upon immunohistochemical analysis, and electron microscopy confirmed that PVAT contained multilocular lipid droplets and abundant mitochondria. Compared to white adipose tissue (WAT), PVAT and BAT from C57BL/6 mice fed a high fat diet for 13 weeks had markedly lower expression of immune cell-enriched mRNAs, suggesting resistance to obesity-induced inflammation. Indeed, staining of BAT and PVAT for macrophage markers (F4/80, CD68) in obese mice showed virtually no macrophage infiltration, and FACS analysis of BAT confirmed the presence of very few CD11b+/CD11c+ macrophages in BAT (1.0%) in comparison to WAT (31%). In summary, murine PVAT from the thoracic aorta is virtually identical to interscapular BAT, is resistant to diet-induced macrophage infiltration, and thus may play an important role in protecting the vascular bed from thermal and inflammatory stress.

overall_design: 8-week-old male C57BL6/J mice were fed a normal (ND) or high fat diet (HFD) (Research Diets 12451, 45 kcal% fat) for 13 weeks. Mice were then euthanized and four different adipose depots were harvested for RNA analysis: perivascular fat from the lesser curvature of the aortic arch (PVAT), interscapular brown adipose (BAT), inguinal adipose tissue (SAT), and epididymal adipose tissue (VAT). 250 ng total RNA pooled from two mice was used for cDNA synthesis; 3 biological replicates per tissue and diet were performed for a total of 24 hybridizations.

first: Michael

last: Czech


title: CBP is required for environmental enrichment-induced neurogenesis and cognitive enhancement.

status: Public on Aug 30 2011

city: Sant Joan d'Alacant

first: Angel

last: Barco

state: Alicante

address: Av. Santiago Ramón y Cajal

country: Spain

institute: Instituto de Neurociencias (UMH-CSIC)

summary:
The epigenetic changes of the chromatin represent an attractive molecular substrate for adaptation to the environment. We examined here the role of CBP, a histone acetyltransferase involved in mental retardation, in the genesis and maintenance of long-lasting systemic and behavioral adaptations to environmental enrichment (EE). Morphological and behavioral analyses demonstrated that EE ameliorates deficits associated to CBP-deficiency. However, CBP-deficient mice also showed a strong defect in environment-induced neurogenesis and impaired EE-enhanced spatial navigation and patter separation ability. These defects correlated with an attenuation of the transcriptional program induced in response to EE and with deficits in histone acetylation at the promoters of EE-regulated, neurogenesis-related genes. Additional experiments in CBP restricted and inducible knockout mice indicated that environment-induced adult neurogenesis is extrinsically regulated by CBP function in mature granule cells. Overall, our experiments demonstrate that the environment alters gene expression by impinging on activities involved in modifying the epigenome and identify CBP-dependent transcriptional neuroadaptation as an important mediator of EE-induced benefits, a finding with important implications for mental retardation therapeutics.
To identify those genes whose expression was altered in the hippocampus of cbp heterozygous mice both under basal conditions and after 2 weeks of exposure to environmental enrichment, we performed a gene profiling analysis of hippocampal tissue using high-density oligonucleotide microarrays.

overall_design: We obtained quintuplicate (standard housing) and triplicate (environmental enrichment) samples containing total RNA from the hippocampi of four 3-month old females of either genotype (in total 32 cbp+/- mice and 32 wild type littermates were used in the experiment).

first: Jose

last: Lopez-Atalaya


title: Gene expression comparison of colorectal cancer samples from patients diagnosed at a young and elderly age

status: Public on Sep 07 2011

city: Oslo

first: Trude

middle: Holmeide

last: Ågesen

address: Ullernch. 70

department: Department of Cancer Prevention

country: Norway

institute: Oslo University Hospital

summary: Colorectal cancer (CRC) incidence increases with age and early onset of the disease is an indication of genetic predisposition, estimated to cause up to 30 % of all CRC cases. To identify genes associated with an increased risk for CRC, genome-wide gene expression levels of CRCs from patients diagnosed at an early age  and CRCs from patients diagnosed at higher age were investigated. Patients with known hereditary predisposition syndromes were excluded from the study.

overall_design: Twenty four patients were diagnosed at a young age (mean, 43 years; range, 28-53 years), referred to as the early onset group. They were excluded from the HNPCC and FAP syndromes by clinical criteria and no other cancer syndromes were recorded for these patients. The second group consisted of 17 patients with primary diagnosis at old age (mean, 79 years; range, 69-87 years), referred to as the late onset group. The samples from the late onset group were selected to reflect the composition of the early onset group with respect to gender, tumor localization and tumor stage according to The International Union Against Cancer (UICC)/American Joint Committee on Cancer (AJCC). Microsatellite instability (MSI) status was previously analyzed in all samples to eliminate the potential risk of including patients with inherited DNA repair deficiencies.

middle: H

first: Marianne

last: Berg

first: Espen

last: Thiis-Evensen

first: Lina

last: Cekaite

first: Guro

middle: E

last: Lind

first: Jahn

last: Nesland

first: Arne

last: Bakka

first: Tom

last: Mala

first: Hans

middle: J

last: Hauss

first: Torunn

last: Fetveit

first: Arild

last: Nesbakken

first: Ragnhild

middle: A

last: Lothe

first: Rolf

middle: I

last: Skotheim


title: ChIP-Seq data from Arabidopsis thaliana under dark and far-red light

status: Public on Aug 02 2011

city: Shenzhen

first: Bosheng

last: Li

state: Guangdong

address: SUStech Huiyuan #1 406

department: MCDB

country: China

institute: SUSTC (current, may different with previous institution )

type: Genome binding/occupancy profiling by high throughput sequencing

summary: FAR-RED ELONGATED HYPOCOTYL 3 (FHY3) and its homolog FAR-RED IMPAIRED RESPONSE 1 (FAR1) are two transposase-derived transcription factors initially identified as the key components in phytochrome A signaling and recently shown to function in the circadian clock. However, whether FHY3 and FAR1 are involved in other processes of plant development remains largely unknown. Here, we explored chromatin immunoprecipitation-based sequencing (ChIP-seq) analysis to identify 1745 and 1171 FHY3 direct binding target genes in darkness and far-red light conditions, respectively in the Arabidopsis thaliana genome. This analysis revealed that FHY3 preferentially binds to the gene promoters through the previously identified typical FHY3/FAR1 binding motif. Interestingly, FHY3 also binds to two novel motifs in the 178-bp repeats of the Arabidopsis centromere regions in vivo. Comparison between the ChIP-seq and microarray data indicates that FHY3 regulates the expression of 196 and 85 genes in dark and far-red respectively by directly binding to their promoters. FHY3 also co-regulates a number of common target genes with PHYTOCHROME INTERACTING FACTOR 3-LIKE 5 (PIL5) and ELONGATED HYPOCOTYL 5 (HY5). Moreover, our genome-wide identification of FHY3 direct target genes ultimately led to the discovery and validation of a new role of FHY3 in controlling chloroplast development, by directly activating the expression of ACCUMULATION AND REPLICATION OF CHLOROPLASTS5 (ARC5), a key gene regulating chloroplast constriction and division. Taken together, our data suggest that FHY3 is involved in regulating multiple facets of plant development, thus providing new insights into the functions of this type of transposase-derived transcription factors.

overall_design: To determine the global in vivo binding sites of FHY3, we performed ChIP-seq analysis using 35S:3FLAG-FHY3-3HA fhy3-4 transgenic lines in which the 3FLAG-FHY3-3HA fusion proteins could largely rescue the long hypocotyl phenotype of the fhy3-4 mutants  The seedlings were grown in D or continuous FR light conditions for 4 days, and then chromatin fragments were prepared using the commercially monoclonal anti-FLAG antibodies. We then generated two DNA libraries, one for D and one for FR -grown samples, which were then subjected to ultra-high throughput Solexa (Illumina) sequencing

first: Xinhao

last: Ouyang

first: Jigang

first: Gang

first: Huaishun

last: Shen

first: Xi

last: Huang

first: Xiaorong

last: Mo

first: Xiangyuan

last: Wan

first: Rongcheng

last: Lin

first: Shigui

first: Haiyang

last: Wang

first: Xingwang

last: Deng


title: MMS toxicogenomics collaboration Affymetrix data

status: Public on Oct 27 2011

city: Setagaya-ku

first: Takayoshi

last: Suzuki

state: Tokyo

address: 1-18-1 Kamiyoga

department: Div. Cellular and Gene Therapy Products

country: Japan

institute: National Institute of Health Sciences, Japan

summary: In order to screen candidate genes that discriminate genotoxic hepatocarcinogens from nongenotoxic ones, we compared gene expression in the liver of mice treated with 6 genotoxic hepatocarcinogens or 5 nongenotoxic hepatocarcinogens.

overall_design: 6 genotoxic and 2 non-genotoxic hepatocarcinogens with expression time of 4h, 20h, 14 days, and 28 days

first: Luan

last: Yang


title: Transcriptional changes of the human pathogenic fungus Trichophyton rubrum related to dermatophyte-skin interaction

status: Public on Oct 03 2011

city: Beijing

first: Qi

last: Jin

address: East Street 6, Yizhuang

institute: Institute of Pathogen Biology

summary: Trichophyton rubrum, an anthropophilic and cosmopolitan fungus, is the most common agent of superficial mycoses, causing rarely deep dermatophytosis in immunocompromised hosts.

overall_design: In this study, an infection condition of T. rubrum was modeled by adding human skin sections into a limited medium containing glucose to monitor T. rubrum gene expression patterns using cDNA microarrays on a global level. We found that exposure to human skin resulted in up-regulation of the expression levels of T. rubrum genes related to many cellular and biological processes, including transcription and translation, metabolism and secondary transport, stress response, and signaling pathways. These results provide a reference set of T. rubrum genes whose expression patterns change upon infection and reveal previously unknown genes that probably corresponding to proteins that should be considered as virulence factor candidates and potential new drug targets for T. rubrum infections.

first: Tao

last: Liu

first: Wenchuan

last: Leng

first: Ruoyu


title: Galanthamine hydrobromide 72h

status: Public on Jul 23 2011

city: Eggenstein-Leopoldshafen

first: Jessica

last: Legradi

state: Baden-Wuerttemberg

address: Hermann-von-Helmholtz-Platz 1

department: Institute of Toxicology and Genomics ITG

country: Germany

institute: Karlsruhe Institute of Technology

summary: Microarray analysis of 72h 1mM Galanthamine hydrobromide treated embryos. At this concentration GAL causes myopathology without causing morbidity.

overall_design: Microarray analysis were performed, with 72h 1mM Galanthamine hydrobromide treated embryos compared with untreated control embryos of the same clutch.

first: Uwe

last: Straehle

first: Sandeep

last: Paul


title: Gene expression profiling of HUVEC after stimulation with an inflammatory mediator cocktail

status: Public on Jul 20 2011

city: Cambridge

first: Muna

last: Affara

address: Tennis Court Road

department: Pathology

country: United Kingdom

institute: University of Cambridge

summary: To understand the transcripts regulated by inflammation and how this regulation may differ between different individuals, the transcriptome of HUVECs stimulated with inflammatory mediators was examined in 9 different individuals.

overall_design: In the study present here, HUVEC isolates from 9 different individuals were cultured until passage 4 in fully supplemented growth conditions. Passage 4 HUVEC isolates were then treated with a cocktail of inflammatory mediators (10ng/ml TNF-α, Il-1β, Il-8) for a period of 24 hours prior to RNA extraction. RNA was extracted and hybridised onto CodeLink microarrays. The expression profiles in the inflammatory mediator treated condition were examined along side a superset of the same biological replicates that were untreated (also submitted), to facilitate assessment of the resting transcriptome.

first: D

middle: S

last: Charnock-Jones

first: Cristin

last: Print


title: Gene expression profiling of HUVEC under basal (resting) conditions

summary: To understand the biological variability that exists between different individuals, the resting (untreated) transcriptome from 15 different individuals was examined. Transcripts expressed in a bimodal manor across the 15 biological replicates were identified for further examination.

overall_design: In the study presented here, HUVEC isolates from 15 different individuals were cultured until passage 4 in fully supplemented growth conditions. RNA was extracted and hybridised onto CodeLink microarrays. The expression profiles in the untreated condition were examined along side a subset of the same biological replicates treated with a cocktail of inflammatory mediators (10ng/ml TNF-α, Il-1β, Il-8) for a period of 24 hours prior to RNA extraction (also submitted).


title: Cytoplasmic MTDH associates with mRNAs

status: Public on Jul 13 2011

city: Iowa City

first: xiangbing

last: meng

state: IA

address: 375 Newton Road

institute: University of Iowa

summary: To examine whether MTDH is a novel RNA binding protein and regulates either metabolism or translation of various mRNAs, we performed a RNA-binding protein immunoprecipitation (RIP) with MTDH and IgG antibodies, and the resulting immunoprecipitated RNA was subjected to a microarray to identify transcripts associating with MTDH. In addition we tested the effect of PI3K inhibition using BEZ235 (a dual PI3K/mTOR inhibitor) on the association of MTDH with target mRNAs.

overall_design: The microarray was performed on three biological triplicates as well as three experimental triplicates of immunoprecipitated MTDH and IgG in Hec50co endometrial cancer cells. Magna RIP (RNA-binding protein immunoprecipitation, Millipore) Kit and microarray were used to immunoprecipitate MTDH associated RNAs and to identify mRNAs that associate with MTDH in the absence or presence of 100nM BEZ235 (LC Labs). Fold changes represent immunoprecipitated MTDH RNA compared to IgG immunoprecipitated RNA. Representative mRNAs associated with MTDH and the effect of BEZ235 on the association of MTDH with mRNAs are shown.

first: Xiangbing

last: Meng

first: Pavla

last: Brachova

first: Kimberly

last: Leslie


title: Post-transcriptional generation of miRNA variants by multiple nucleotidyl transferases contributes to miRNA transcriptome complexity (Validation 2)

status: Public on Jun 15 2011

city: Seattle

first: Muneesh

last: Tewari

state: WA

address: 1100 Fairview Ave, N

department: Human Biology

institute: Fred Hutchinson Cancer Research Center

summary:
Modification of microRNA sequences by the 3' addition of nucleotides to generate so-called “isomiRs” adds to the complexity of miRNA function, with recent reports showing that 3' modifications can influence miRNA stability and efficiency of target repression. Here we show that the 3' modification of miRNAs is a physiological and common post-transcriptional event that shows selectivity for specific miRNAs and is observed across species ranging from C. elegans to human. The modifications result predominantly from adenylation and uridylation, and are seen across tissue types, disease states, and developmental stages. To quantitatively profile 3' nucleotide additions, we developed and validated a novel assay based on NanoString Technologies' nCounter platform. For certain miRNAs, the frequency of modification was altered by processes such as cell differentiation, indicating that 3' modification is a biologically regulated process.  To investigate the mechanism of 3' nucleotide additions, we used RNA interference to screen a panel of eight candidate miRNA nucleotidyl transferases for 3' miRNA modification activity in human cells. Multiple enzymes, including PAPD1, PAPD4, PAPD5, ZCCHC6, ZCCHC11, and TUT1, were found to govern 3' nucleotide addition to miRNAs in a miRNA-specific manner. Three of these enzymes–PAPD1, ZCCHC6 and TUT1–have not previously been known to modify miRNAs.  Collectively, our results indicate that 3' modification observed in next generation small RNA sequencing data is a biologically relevant process, and identify enzymatic mechanisms that may lead to new approaches for modulating miRNA activity in vivo.
The matrix presents the counts from when the two pools of synthetic oligos were profiled in the each of the three bridge pools.

overall_design: We validated the specificity of the platform for the miRNAs of interest by assaying two pools of synthetic RNA oligonucleotides representing canonical or 3' variant versions of 14 miRNAs. Each bridge pool (i.e.,  canonical, variant 1 or variant 2 sequences) was used to assay the two mixtures of synthetic miRNA oligonucleotides.  A single replicate of each pool was profiled in each assay.

first: Stacia

middle: K

last: Wyman


title: Genome-scale DNA methylation pattern profiling of human bone marrow mesenchymal stem cells in long-term culture

status: Public on Jun 17 2011

city: Ansan

first: Kim

last: Seung Jun

state: Kyunggido

address: Sangrokgu, sadong, Hanyang Univ., HBI 628

department: Genome Business

country: South Korea

institute: GenoCheck

type: Methylation profiling by genome tiling array

summary:
Background: Human bone marrow mesenchymal stem cells (MSCs) expanded in vitro exhibit not only a tendency to lose their proliferative potential, homing ability and telomere length but also genetic or epigenetic modifications such as DNA methylation and the noncoding RNA-mediated mechanism. This results in the senescence of MSCs. We compared differential methylation patterns of genes and miRNAs between early-passage and late-passage cells and estimated the relationship between senescence and DNA methylation patterns. Genomic DNA of MSCs cultured at passage 5 (P5) and passage 15 (P15) was enriched using methylated DNA immunoprecipitation assays, and DNA methylation changes due to long-term culture of MSCs were analyzed using Human 2.1M Deluxe Promoter Arrays (NimbleGen).

Results: When we analyzed the methylation differences between P5 and P15 more than twice, 3,338 genes showed more than two-fold higher methylation at P5 than P15, whereas 4,670 genes showed more than two-fold higher methylation at P15 than P5. When we examined hypermethylated genes (methylation peak ≥ 2) at P5 or P15, 2,739 genes, including those related to fructose and mannose metabolism and calcium signaling pathways, and 2,587 genes, including those related to DNA replication, cell cycle and the PPAR signaling pathway, were hypermethylated at P5 and P15, respectively. There was common hypermethylation of 1,205 genes at both P5 and P15. In addition, genes that were hypermethylated at P5 (CPEB1, GMPPA, CDKN1A, TBX2, SMAD9 and MCM2) showed lower mRNA expression than did those hypermethylated at P15, whereas genes that were hypermethylated at P15 (MAML2, FEN1 and CDK4) showed lower mRNA expression than did those that were hypermethylated at P5, demonstrating that hypermethylation at DNA promoter regions inhibited gene expression and that hypomethylation increased gene expression. In the case of hypermethylation on miRNA, 27 miRNAs were hypermethylated at P5, whereas 44 miRNAs were hypermethylated at P15.

Conclusion: These results show that hypermethylation increases at genes related to DNA replication, cell cycle and adipogenic differentiation due to long-term culture, which may in part affect MSC senescence.

overall_design: Analysis of DNA methylation changes of MSCs at passage 5 and passage 15 using Human 2.1M Deluxe Promoter Arrays (NimbleGen)

first: MiRan

last: Choi

first: Young-Ho

last: In

first: Jungsun

last: Park

first: Taesung

first: Kyoung

last: Jung

first: Nando

middle: D

last: Das

first: MiKyung

last: Chung

first: Sunghoon

last: Kim

first: Young

middle: G

last: Chai


title: *MMP1* bimodal expression and differential response to inflammatory mediators is linked to promoter polymorphisms

first: Muna

last: Affara

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

overall_design: Refer to individual Series


title: Post-transcriptional generation of miRNA variants by multiple nucleotidyl transferases contributes to miRNA transcriptome complexity (Validation 1)

summary: Modification of microRNA sequences by the 3' addition of nucleotides to generate so-called “isomiRs” adds to the complexity of miRNA function, with recent reports showing that 3' modifications can influence miRNA stability and efficiency of target repression. Here we show that the 3' modification of miRNAs is a physiological and common post-transcriptional event that shows selectivity for specific miRNAs and is observed across species ranging from C. elegans to human. The modifications result predominantly from adenylation and uridylation, and are seen across tissue types, disease states, and developmental stages. To quantitatively profile 3' nucleotide additions, we developed and validated a novel assay based on NanoString Technologies' nCounter platform. For certain miRNAs, the frequency of modification was altered by processes such as cell differentiation, indicating that 3' modification is a biologically regulated process.  To investigate the mechanism of 3' nucleotide additions, we used RNA interference to screen a panel of eight candidate miRNA nucleotidyl transferases for 3' miRNA modification activity in human cells. Multiple enzymes, including PAPD1, PAPD4, PAPD5, ZCCHC6, ZCCHC11, and TUT1, were found to govern 3' nucleotide addition to miRNAs in a miRNA-specific manner. Three of these enzymes–PAPD1, ZCCHC6 and TUT1–have not previously been known to modify miRNAs.  Collectively, our results indicate that 3' modification observed in next generation small RNA sequencing data is a biologically relevant process, and identify enzymatic mechanisms that may lead to new approaches for modulating miRNA activity in vivo.

overall_design: We validated the specificity of the platform by assaying three pools of synthetic RNA oligonucleotides representing canonical, variant 1, and variant 2 versions of 5 miRNAs (miR-15a, miR-15b, miR-125a-5p, miR-143 and miR-221).Each bridge pool (i.e., specific to canonical, variant 1 or variant 2) was used to assay the three mixtures of synthetic miRNA oligonucleotides.  In order to assess accuracy, we also assayed mixtures of 5 synthetic miRNAs containing 60% canonical miRNA, 30% variant 1, and 10% variant 2. Triplicate technical replicates of each sample were performed.


title: Post-transcriptional generation of miRNA variants by multiple nucleotidyl transferases contributes to miRNA transcriptome complexity (hESC)

overall_design: RNA was isolated from undifferentiated or differentiated H1 human embryonic cells as previously described (Bar et al, 2008). Two biological replicates of undifferentiated and differentiated cell RNA was profiled on the NanoString nCounter miRNA expression assay adapted to discriminate miRNA 3' variants.


title: Expression data from early luteal bovine endometrium

status: Public on Jun 10 2011

city: Grange

first: aideen

middle: patricia

last: killeen

state: Co. Meath

address: Dunsany

department: Animal and Bioscience Research Department

country: Ireland

institute: Teagasc

summary:
In both beef and dairy cattle, the majority of embryo loss occurs in the first 14 days following insemination. During this period, the embryo is completely dependent on its maternal uterine environment for development, growth and ultimately survival, therefore an optimum uterine environment is critical to embryo survival.
We used microarrays to assess endometrial gene expression in high and low fertility heifers during the mid-luteal phase of the estrous cycle.

overall_design: Charlaois × Limousin heifers were artificially inseminated on 4 successive occasions and heifers were subsequently characterized as either high or low fertility (HF; LF) based on the presence of an embryo on day 28 of pregnancy on all four inseminations or on one occasion only, respectively. From this population of animals, HF and LF heifers were slaughtered on day 7 of a synchronized estrous cycle and global gene expression in uterine endometrial tissue was determined using the Affymetrix 23K Bovine GeneChip.

first: Aideen

last: Killeen

first: Dermot

last: Morris

first: David

last: Kenny

first: Michael

last: Diskin

first: Sinead

last: Waters


title: Post-transcriptional generation of miRNA variants by multiple nucleotidyl transferases contributes to miRNA transcriptome complexity (normal prostate and prostate cells)

status: Public on Jun 13 2011

first: Muneesh

last: Tewari

type: Non-coding RNA profiling by high throughput sequencing

summary: Modification of microRNA sequences by the 3' addition of nucleotides to generate so-called isomiR adds to the complexity of miRNA function, with recent reports showing that 3' modifications can influence miRNA stability and efficiency of target repression. Here we show that the 3' modification of miRNAs is a physiological and common post-transcriptional event that shows selectivity for specific miRNAs and is observed across species ranging from C. elegans to human. The modifications result predominantly from adenylation and uridylation, and are seen across tissue types, disease states, and developmental stages. To quantitatively profile 3' nucleotide additions, we developed and validated a novel assay based on NanoString Technologies' nCounter platform. For certain miRNAs, the frequency of modification was altered by processes such as cell differentiation, indicating that 3' modification is a biologically regulated process. To investigate the mechanism of 3' nucleotide additions, we used RNA interference to screen a panel of eight candidate miRNA nucleotidyl transferases for 3' miRNA modification activity in human cells. Multiple enzymes, including MTPAP, PAPD4, PAPD5, ZCCHC6, ZCCHC11, and TUT1, were found to govern 3' nucleotide addition to miRNAs in a miRNA-specific manner. Three of these enzymes (MTPAP, ZCCHC6 and TUT1) have not previously been known to modify miRNAs. Collectively, our results indicate that 3' modification observed in next generation small RNA sequencing data is a biologically relevant process, and identify enzymatic mechanisms that may lead to new approaches for modulating miRNA activity in vivo.

overall_design:
Normal human prostate tissue was sequenced in three lanes by Illumina sequencing
454 sequencing of two prostate cell culture samples, one epithelial, one stromal

first: Stacia

last: Wyman


title: Post-transcriptional generation of miRNA variants by multiple nucleotidyl transferases contributes to miRNA transcriptome complexity

sample_organism:
Homo sapiens
synthetic construct


title: Gene expression profiles of dental follicle cells after 48 hours of transfection in vitro with a DLX3 plasmid and an empty vector

status: Public on Jun 07 2011

city: Regensburg

first: Christian

last: Morsczeck

address: Franz-Josef Strauss Allee 11

institute: University of Regensburg

summary: We analysed gene expression profiles in dental follicle cells after 48 hours of overexpression with the transcription factor DLX3.

overall_design: Total RNAs were isolated from dental follicle cells after 48 hours of transfection with a DLX3 expressions plasmid and for control with an empty vector.

first: Sandra

last: Viale-Bouroncle

first: Oliver

last: Felthaus

first: Gottfried

last: Schmalz

first: Torsten

last: Reichert


title: A biophysical DNA binding model for the LEAFY transcription factor reveals the evolutionary fluidity of its binding sites

status: Public on May 13 2011

city: Tuebingen

first: Felix

last: Ott

address: Spemannstr. 35-39

institute: Max Planck Institute for Developmental Biology

summary: Despite great advances in sequencing capacity, generating functional information for non-model organisms remains a challenge. One solution lies in an improved ability to predict genetic circuits based on primary DNA sequence combined with the characterization of regulatory molecules from model species. Here, we focus on the LEAFY (LFY) transcription factor, a conserved master regulator of floral development. Starting with biochemical and structural information, we built a biophysical model describing LFY DNA binding specificity in vitro that accurately predicts in vivo LFY binding sites in the Arabidopsis thaliana genome. Extending the model to other species, we show that it can correctly identify functional homologs of known LFY targets from Arabidopsis thaliana in other angiosperms, even if a functional shift between orthologs and paralogs has occurred. Moreover, this model demonstrates the evolutionary fluidity of the link between LFY and one of its target genes, underlining how this regulatory interaction can be conserved despite changes in position, sequence and affinity of the LFY binding sites. Our study shows that the cis-element fluidity recently illustrated in animals also exists in plants, and that it can be detected without any experimental work in each individual species, using a biophysical transcription factor model.

overall_design: A. thaliana LEAFY ChIP-seq w control, 2 replicates

first: Edwige

last: Moyroud

first: Eugenio

last: Minguet

first: Levi

last: Yant

last: Posé

first: Sandrine

last: Blanchet

first: Marie

last: Monniaux

first: Olivier

last: Bastien

first: Emmanuel

last: Thévenon

first: Detlef

last: Weigel

first: Markus

last: Schmid

first: Francois

last: Parcy


title: Coexpression of Normally Incompatible Developmental Pathways in Retinoblastoma Genesis [single tumor cell data]

city: Memphis

first: David

last: Finkelstein

state: TN

address: 332 N. Lauderdale St.

department: Computational Biology

institute: St Jude Children's Research Hospital

summary: It is widely believed that the molecular and cellular features of a tumor reflect its cell-of-origin and can thus provide clues about treatment targets. The retinoblastoma cell-of-origin has been debated for over a century.  Here we report that human and mouse retinoblastomas have molecular, cellular, and neurochemical features of multiple cell classes, principally amacrine/horizontal interneurons, retinal progenitor cells, and photoreceptors. Importantly, single-cell gene expression array analysis showed that these multiple cell type–specific developmental programs are coexpressed in individual retinoblastoma cells, which creates a progenitor/neuronal hybrid cell. Importantly, neurotransmitter receptors, transporters, and biosynthetic enzymes are expressed in human retinoblastoma, and targeted disruption of these pathways reduces retinoblastoma growth in vivo and in vitro.   Our finding that retinoblastoma tumor cells express multiple neuronal differentiation programs that are normally incompatible in development suggests that the pathways that control retinal development and establish distinct cell types are perturbed during tumorigenesis. Therefore, the cell-of-origin for retinoblastoma cannot be inferred from the features of the tumor cells themselves. However, we now have a detailed understanding of the neuronal pathways that are deregulated in retinoblastoma and targeting the catecholamine and indolamine receptors or downstream components could provide useful therapeutic approaches in future studies. This example highlights the importance of comprehensive molecular, cellular and physiological characterization of human cancers with single cell resolution as we incorporate molecular targeted therapy into treatment regimens.

overall_design: 20 single cells isolated from primary pediatric retinoblastoma tumors were assayed to asses the within tumor consistency of expression signals

first: Justina

last: McEvoy

first: Jacqueline

last: Flores-Otero

first: Jiakun

last: Zhang

first: Katie

last: Nemeth

first: Rachel

last: Brennan

first: Cori

last: Bradley

first: Fred

last: Krafcik

first: Carlos

last: Rodriguez-Galindo

first: Matthew

last: Wilson

first: Shunbin

last: Xiong

first: Guillermina

last: Lozano

first: Julien

last: Sage

first: Ligia

last: Fu

first: Lotfi

last: Louhibi

first: Jeff

last: Trimarchi

first: Amar

last: Pani

first: Richard

last: Smeyne

first: Dianna

last: Johnson

first: Michael

last: Dyer


title: Coexpression of Normally Incompatible Developmental Pathways in Retinoblastoma Genesis [human tumor/cell line data]

first: David

last: Finkelstein

overall_design: 55 primary pediatric retinoblastoma tumors were collected and assayed and compared to with 3 passaged xenografts and 4 RB cell lines

first: Justina

last: McEvoy

first: Jacqueline

last: Flores-Otero

first: Jiakun

last: Zhang


title: Coexpression of Normally Incompatible Developmental Pathways in Retinoblastoma Genesis [mouse model data]

overall_design: 132 mouse model primary retinoblastoma tumor Samples

first: Katie

last: Nemeth

first: Rachel

last: Brennan

first: Cori

last: Bradley

first: Fred

last: Krafcik


title: Identification of androgen-regulated genes in RWPE-1-AR cells

status: Public on May 12 2011

city: OULLINS

first: Virginie

last: Vlaeminck-Guillem

address: Chemin du Grand Revoyet

department: Faculté de Médecine Lyon Sud

country: France

institute: Institut de Génomique Fonctionnelle de Lyon

summary: Transcriptional profiling of RWPE-1 cells stably expressing human androgen receptor (as described in Altintas et al., Mol Cell Endocrinol 2011) treated with a non-metabolisable androgen, R1881

overall_design: RWPE-1-AR cells were treated with R1881 during 3h or 24h and compared to control not treated cells. Three independent cell culture experiments for each treatment condition (vehicle or R1881 for 3h and 24h).

last: Vlaeminck-guillem

first: Dogus

last: Altintas

first: Jacques

last: Samarut


title: Coexpression of Normally Incompatible Developmental Pathways in Retinoblastoma Genesis

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus


title: Post-transcriptional generation of miRNA variants by multiple nucleotidyl transferases contributes to miRNA transcriptome complexity (KD)

first: Muneesh

last: Tewari

overall_design: Total RNA was isolated from HCT-116 cells 48 hours post transfection with siRNAs targeting a panel of nucleotidyl transferases, and profiled with a version of the NanoString nCounter miRNA expression assay adapted to discriminate miRNA 3' variants. Four biological replicates of cells treated with the negative control siRNA directed against Cyclophilin B were profiled.  Two biological replicates of cells treated individually with siRNAs targeting each of eight nucleotidyl transferases were profiled.

first: Stacia

last: Wyman


title: LEAFY target genes reveal a link between flower development and biotic stimulus response

status: Public on May 11 2011

city: Philadelphia

first: Cara

middle: Michele

last: Winter

address: 421 Curie Blvd.

institute: University of Pennsylvania

type: Genome binding/occupancy profiling by genome tiling array


title: LEAFY target genes reveal a link between flower development and biotic stimulus response (II)

summary:
The transition from vegetative growth to flower formation is critical for the survival of flowering plants. The plant-specific transcription factor LEAFY (LFY) has central, evolutionarily conserved roles in this process, both in the formation of the first flower and later in floral patterning. We performed genome-wide binding and expression studies to elucidate the molecular mechanisms by which LFY executes these roles. Our study reveals that LFY directs an intricate regulatory network in control of floral homeotic gene expression and, unexpectedly, controls the expression of genes regulating the response to external stimuli in Arabidopsis. We further show that LFY dampens responses to a bacterial MAMP (microbe-associated molecular pattern) and to pathogen challenge. Our findings suggest a molecular mechanism for the coordination of reproductive stage development and disease response programs in plants. Regulation of these distinct survival programs by a single transcription factor may ensure optimal allocation of plant resources for reproductive fitness.
Genome binding/occupancy profiling by genome tiling array used to identity genes bound by endogenous LFY in inflorescences.

overall_design: ChIP-chip of 19-day-old WT (Landsberg erecta) inflorescences using a LFY antibody. Five biological replicate immunoprecipated samples were compared to their corresponding genomic input DNA (taken prior to immunoprecipitation). As a negative control, one 19-day-old Ify6 (Landsberg erecta) IP and its corresponding Input sample were also hybridized.

first: Doris

last: Wagner


title: LEAFY target genes reveal a link between flower development and biotic stimulus response (I)

summary:
The transition from vegetative growth to flower formation is critical for the survival of flowering plants. The plant-specific transcription factor LEAFY (LFY) has central, evolutionarily conserved roles in this process, both in the formation of the first flower and later in floral patterning. We performed genome-wide binding and expression studies to elucidate the molecular mechanisms by which LFY executes these roles. Our study reveals that LFY directs an intricate regulatory network in control of floral homeotic gene expression and, unexpectedly, controls the expression of genes regulating the response to external stimuli in Arabidopsis. We further show that LFY dampens responses to a bacterial MAMP (microbe-associated molecular pattern) and to pathogen challenge. Our findings suggest a molecular mechanism for the coordination of reproductive stage development and disease response programs in plants. Regulation of these distinct survival programs by a single transcription factor may ensure optimal allocation of plant resources for reproductive fitness.
Genome binding/occupancy profiling by genome tiling array used to identity genes bound by induced LFY in 9-day-old seedlings.

overall_design: ChIP-chip of 9-day-old 35S::LFY-GR dexamethasone-treated seedlings using a LFY antibody. Three biological replicate immunoprecipated samples were compared to their corresponding genomic input DNA (taken prior to immunoprecipitation). As a negative control, three biological replicates of 9-day-old WT (Landsberg erecta) IP and their corresponding Input samples were also hybridized (9-day-old Ler seedlings express very low levels of LFY).


title: Peripheral blood RNA expression profiling in illicit methcathinone users reveals effect on immune system

status: Public on Apr 19 2011

city: Perth

first: Sulev

last: Koks

state: Western Australia

address: 90 South Street

department: Centre for Molecular Medicine and Innovative Therapeutics

country: Australia

institute: Murdoch University

summary: Methcathinone (ephedrone) is relatively easily accessible for abuse. Its users develop an extrapyramidal syndrome and it is not known if this is caused by methcathinone itself, by side-ingredients (manganese), or both. In the present study we aimed to clarify molecular mechanisms underlying this condition. We used microarrays to analyze whole genome gene expression patterns of peripheral blood from 20 methcathinone users and 20 matched controls. Gene expression profile data were analyzed by Bayesian modelling and functional annotation. Of 28,869 genes on the microarrays, 326 showed statistically significant differential expression with FDR adjusted p-values below 0.05. Quantitative RT-PCR confirmed differential expression for the most of the genes selected for validation. Functional annotation and network analysis indicated activation of a gene network that included immunological disease, cellular movement and cardiovascular disease functions (enrichment score 42). As HIV and HCV infections were confounding factors, we performed additional stratification of patients. A similar functional activation of the “immunological disease” category was evident when we compared patients according to injection status (past versus current users, balanced for HIV and HCV infection). However, this difference was not large therefore the major effect was related to the HIV status of the patients. Mn-methcathinone abusers have blood RNA expression patterns that mostly reflect their HIV and HCV infections. However, despite the strong confounding effect from infection, some modest drug abuse effects on gene expression were detected.

overall_design: 40 samples, 20 healthy volunteers and 20 illicit methcathinone users

first: Katrin

last: Sikk

last: Kõks

first: Ursel

last: Soomets

first: Leonard

middle: C

last: Schalkwyk

first: Cathy

last: Fernandes

last: Haldre

first: Sten-Magnus

last: Aquilonius

first: Pille

last: Taba


title: Accumulating mitochondrial DNA mutations drive premature hematopoietic aging phenotypes molecularly distinct from physiologic stem cell aging

status: Public on May 06 2011

city: Lund

first: Gudmundur

last: Norddahl

address: Tornavagen 10

country: Sweden

institute: Lund University

summary:
Somatic stem cells mediate tissue maintenance for the lifetime of an organism. Despite the well-established longevity that is a prerequisite for such function, accumulating data argue for compromised stem cell function with age. Identifying the mechanisms underlying age-dependent stem cell dysfunction is therefore key to understand the aging process.
Using a model that carries a proofreading defective mitochondrial DNA polymerase, we demonstrate hematopoietic defects reminiscent of premature HSC aging including anemia, lymphopenia and myeloid lineage skewing. However, in contrast to physiologic stem cell aging, rapidly accumulating mitochondrial DNA mutations displayed little involvement of the hematopoietic stem cell pool but rather with distinct differentiation blocks and/or disappearance of downstream progenitors.

overall_design: Hematopoietic stem cells (HSC) has been sorted out from midaged wildtype and mutator mice and compared with stem cells sorted from young and and old wt mice

middle: L

first: Cornelis

last: Pronk

first: Martin

last: Wahlestedt

first: Gerd

last: Sten

first: Jens

middle: M

last: Nygren

first: Amol

last: Ugale

first: Mikael

last: Sigvardsson

first: David

last: Bryder


title: Expression profile of etoposide-resistant MCF7 (MCF7VP) cells

status: Public on Apr 08 2011

city: Frederick

first: Kate

middle: McGee

last: Im

state: MD

address: 1050 Boyles Street

institute: National Cancer Institute at Frederick

summary: The second leading cause of cancer death for women in the U.S. is breast cancer.  Moreover, a significant number of patients with breast tumors acquire resistance to drugs during therapy. To develop targeted therapeutic strategies to combat drug resistance it is essential to understand the basic molecular mechanisms through which cancer cells control sensitivity to chemotherapeutics. To identify new candidate genes and facilitate the discovery of novel drug resistance pathways, we have generated a resistance profile or  ‘resistome’ of  etoposide resistant MCF7 breast cancer cells. Differential expression of over 5000 genes (fold change > 2, P value < 0.05) indicate that several drug resistance mechanisms may be operating in these cells, including up-regulation of ABC transporter genes, down-regulation of the drug target and down-regulation of apoptotic genes. Several transcription factors such as RUNX2, SOX9, ETS1 and SMAD3 were up-regulated in the drug resistant cells. Targeted RUNX2 knockdown in the resistant cells using siRNA increased sensitivity to etoposide and also upregulated expression of pro-apoptotic genes indicating that RUNX2 could be a molecular target against etoposide resistance. Differential miRNA (microRNA) expression was observed among the drug resistant and sensitive cells suggesting that miRNA may also play a role in regulation of drug resistance. Hsa-miR-218, which targets ABCC6, was down-regulated in the drug resistant cell line. Transfection of a miR-218 mimic could down-regulate the expression of the efflux pump ABCC6 by 65% in drug resistant cells suggesting that regulation of miRNA may play an important role in etoposide resistance.

overall_design: RNA from MCF7 and etoposide-resistant MCF7 (MCF7VP) cell lines were hybridized to Affymetrix microarrays. Both samples were run in triplicate

first: Karobi

last: Moitra

first: Katy

last: Limpert

first: Julie

last: Sawitzke

first: Rob

last: Robey

first: Naoya

last: Yuhki

first: Ram

last: Savan

first: Da

middle: W

last: Huang

first: Howard

last: Young

first: Richard

last: Lempiki

first: Susan

last: Bates

first: Michael

last: Dean


title: Latexin is downregulated in human upper digestive track tumors and exhibits tumor suppressor potential

status: Public on Apr 14 2011

first: Jianqing

last: Zhao

address: 18 Life Science Parkway

institute: Capitalbio. Crop

summary: Latexin (Lxn) was originally isolated as a tissue specific marker of rat neuron. It is expressed in various tissues of humans and many other vertebrates. Lxn inhibits human carboxypeptidase A4 (hCPA4), whose expression is induced by histone deacetylase inhibitors in prostate cancer cells, and is associated with cancer progression. Structural analysis has shown significant similarity between Lxn protein and the tumor suppressor protein TIG1. Recent reports have demonstrated Lxn functions in the negative control of hematopoietic stem cell numbers (HSC) in mice. In the present study, an anti-Lxn monoclonal antibody 1G11 was generated and Western blot analysis showed that Lxn was expressed mainly in the cytoplasm of numerous human cell lines. Immunohistochemical analysis in gastric cancer tissues and the corresponding adjacent normal tissues showed that Lxn expression was lower in cancer tissues as compared to normal tissues. In addition, the Lxn gene was exogenously introduced into a Lxn null gastric cancer cell line, MGC803. Results of colony formation assay, soft agar assay and tumor growth in nude mice showed that over-expression of Lxn inhibits the proliferation and tumorigenicity of MGC803 cells expressing Lxn. Thirty-seven genes, whose expression were altered in response to Lxn expression were identified by microarray analysis. Methylation analysis of the central promoter region of the Lxn gene in several cell lines suggested that Lxn expression was silenced by CpG hypermethylation. Taken together, our results indicate that Lxn is a potential tumor suppressor and plays a role in negative control of tumor cell growth.

overall_design: We analyzed gene expression differece between Latexin (Lxn) postive cell line and negtive cells to find the active pathway and related biology function.

first: Yong

last: Li


title: Deciphering the Cancer Cell Resistome: Gene and microRNA Expression Signatures reveal Unique Molecular Targets for Therapeutic Intervention in Etoposide Resistant Breast Cancer

summary:
The second leading cause of cancer death for women in the U.S. is breast cancer.  Moreover, a significant number of patients with breast tumors acquire resistance to drugs during therapy. To develop targeted therapeutic strategies to combat drug resistance it is essential to understand the basic molecular mechanisms through which cancer cells control sensitivity to chemotherapeutics. To identify new candidate genes and facilitate the discovery of novel drug resistance pathways, we have generated a resistance profile or  ?resistome? of  etoposide resistant MCF7 breast cancer cells. Differential expression of over 5000 genes (fold change > 2, P value < 0.05) indicate that several drug resistance mechanisms may be operating in these cells, including up-regulation of ABC transporter genes, down-regulation of the drug target and down-regulation of apoptotic genes. Several transcription factors such as RUNX2, SOX9, ETS1 and SMAD3 were up-regulated in the drug resistant cells. Targeted RUNX2 knockdown in the resistant cells using siRNA increased sensitivity to etoposide and also upregulated expression of pro-apoptotic genes indicating that RUNX2 could be a molecular target against etoposide resistance. Differential miRNA (microRNA) expression was observed among the drug resistant and sensitive cells suggesting that miRNA may also play a role in regulation of drug resistance. Hsa-miR-218, which targets ABCC6, was down-regulated in the drug resistant cell line. Transfection of a miR-218 mimic could down-regulate the expression of the efflux pump ABCC6 by 65% in drug resistant cells suggesting that regulation of miRNA may play an important role in etoposide resistance.

This SuperSeries is composed of the SubSeries listed below.

overall_design: Refer to individual Series.


title: Real-time quantitative PCR analysis of mouse RAW264.7 macrophages

status: Public on Sep 08 2011

city: Boston

first: Shitao

state: MA

address: 77 Ave Louis Pasteur

department: MBIB

institute: Harvard Medical School

summary: Mouse RAW264.7 macrophages were treated with LPS, IFNb, poly(rI:rC), poly(dA:dT), VSV, HSV, Sendai virus. Genes identified by Human Innate Immunity Interactome for type I Interferon (HI5) were examined for expression.

overall_design: qPCR gene expression profiling. RAW264.7 macrophages were used and treated separately as indicated in the summary. Equal amount total RNA from each group was  used for gene expression analysis.

first: Lingyan

last: Wang

last: Berman

first: Martin

last: Dorf


title: mRNA expression profile in PCa And BPH from Chinese patients

status: Public on Mar 29 2011

city: Guangzhou

first: Yifeng

last: Luo

address: 58 Zhongshan 2nd Road

department: Institute of Respiratory Diseases of Sun Yat-sen University

institute: The First Affiliated Hospital of Sun Yat-sen University

summary: To study the miRNA modulating network and different expressed miRNA lists in prostate cancer, we use hight-throught method to genomically detect the expression profile of miRNA, mRNA and protein.

overall_design: Agilent microarray products for miRNA and mRNA expression profile are choosed. As possible as we can to use the same sample for miRNA, mRNA and protein experiment. 4 prostate cancer and 4 normal tissue are applied to mRNA microarray experiment. The mRNA microaaray experiment service are supplied by ShanghaiBio Corporation(Shanghai,China).

first: Weide

last: Zhong

first: Huichan

last: He

first: Xiyin

last: Liu

first: Qishan

last: Dai

first: Funeng

last: Jiang

first: Jin

first: Dongjiang

last: Liao

first: Xinbo

last: Lu


title: Expression profile of etoposide-resistant MCF7 (MCF7VP) cells with targeted RUNX2 knockdown

first: Kate

last: Im

summary: The second leading cause of cancer death for women in the U.S. is breast cancer, moreover, a significant number of patients with breast tumors acquire resistance to drugs during therapy. To develop targeted therapeutic strategies to combat drug resistance it is essential to understand the basic molecular mechanisms through which cancer cells control sensitivity to chemotherapeutics. To identify new candidate genes and facilitate the discovery of novel drug resistance pathways, we have generated a resistance profile or  ‘resistome’ of  etoposide resitant MCF7 breast cancer cells. Differential expression of over 5000 genes (fold change > 2, P value < 0.05) indicate that several drug resistance mechanisms may be operating in these cells, including up-regulation of ABC transporter genes, down-regulation of the drug target and down-regulation of apoptotic genes. Several transcription factors such as RUNX2, SOX9, ETS1 and SMAD3 were up-regulated in the drug resistant cells. Targeted RUNX2 knockdown in the resistant cells using siRNA increased sensitivity to etoposide and also upregulated expression of pro-apoptotic genes indicating that RUNX2 could be a molecular target against etoposide resistance. Differential miRNA (microRNA) expression was observed among the drug resistant and sensitive cells suggesting that miRNA may also play a role in regulation of drug resistance. Hsa-miR-218, which targets ABCC6, was down-regulated in the drug resistant cell line. Transfection of a miR-218 mimic could down-regulate the expression of the efflux pump ABCC6 by 65% in drug resistant cells suggesting that regulation of miRNA may play an important role in etoposide resistance.

overall_design: RNA from etoposide-resistant MCF7 (MCF7VP) cell lines were hybridized to Affymetrix microarrays.

first: Karobi

last: Moitra

first: Katy

last: Limpert

first: Julie

last: Sawitzke

first: Rob

last: Robey

first: Naoya

last: Yuhki

first: Ram

last: Savan

first: Da

last: Huang

first: Howard

last: Young

first: Richard


title: Expression data from CXCR4+/FLK1+ and CXCR4-/FLK1-  cells isolated by flow cytometry from Day 5 embryoid bodies

status: Public on Mar 11 2011

city: Rochester

first: Anca

last: Chiriac

state: MN

address: Stabile 5, 200 First Street SW

department: Molecular Pharmacology and Experimental Therapeutics

institute: Mayo Clinic

summary:
A dual biomarker signature extracted a stage-specific cytotype according to cell surface expression of CXCR4/FLK-1 after 5 days of spontaneous differentiation from pluripotent stem cells. Genome-wide microarray analysis revealed a high degree of similarity between CXCR4+/FLK-1+ and CXCR4-/FLK-1- subpopulations at day 5, yet the divergent gene expression profile represents more than 700 unique transcripts. Functional analysis of the 294 up-regulated and 440 down-regulated transcripts that distinguished CXCR4+/Flk-1+ from CXCR4-/Flk-1- subpopulations identified an overt ontologic prioritization of “Cardiovascular Development”-IPA 7.0, 2009.Thus, a biomarker-selected subpopulation from spontaneously differentiated pluripotent stem cells identifies a pool of genes that non-stochastically integrate into a blueprint providing instructions for cardiac lineage-specification.

Keywords: Comparison of day 5 embryonic stem cell progenity: CXCR4/FLK-1 biomarker positive versus biomarker negative cells

overall_design: Differentiating embryonic stem cells were FACS sorted at day 5 based on a dual CXCR4/FLK-1 biomarker signature and double positive and double negative progeny were thus collected. Day 5 sorted progeny were independently collected to provide raw material for three biological replicates for each experimental condition. In this manner, three CXCR4/FLK-1 double positive biological samples, and three CXCR4/FLK-1 double negative biological samples were obtained. Total RNA was extracted from each of the samples and RNA pools were profiled on Affymetrix Mouse 430 2.0 Arrays to identify global gene expression changes between double positive and double negative progeny at day 5 of differentiation.

first: Timothy

last: Nelson

first: Randolph

last: Faustino

first: Atta

last: Behfar

first: Andre

last: Terzic


title: Identification and functional analysis of 9p24 amplified genes in human breast cancer

status: Public on Apr 29 2011

city: Detroit

first: Zeng-Quan

last: Yang

state: MI

address: 4100 John R HWCRC 815

department: Oncology

institute: Karmanos Cancer Institute

type: Genome variation profiling by genome tiling array

summary: For the purpose of characterization of the 9p24 amplicon, we carried out high-resolution array CGH (Agilent 244K chip) analysis of four cancer cell lines, including three breast cancer cell lines, Colo824, HCC1954 and HCC70, and one esophageal cancer cell line, KYSE150.

overall_design: For each array, female DNA (Promega, Madison, WI) was used as a reference sample and labelled with Cy-3. The samples of interest were each labelled with Cy-5.


title: Signal-dependent incorporation of MyoD-BAF60c into Brg1-based SWI/SNF chromatin-remodeling complex

status: Public on Mar 07 2011

city: La Jolla

first: Roy

last: Williams

state: CA

address: 3030 Science Park Road

department: Regenerative Medicine

institute: The Scripps Research Institute

summary: Tissue-specific transcription factors initiate differentiation toward a specialized cell type by inducing transcription-permissive chromatin modifications at target gene promoters, through the recruitment of the SWI/SNF chromatin-remodeling complex (1, 2). The molecular mechanism that regulates the chromatin re-distribution of SWI/SNF in response to differentiation signals is currently unknown. Here we show that the muscle determination factor MyoD and the SWI/SNF structural sub-unit, BAF60c (SMARCD3), form a complex on the regulatory elements of MyoD-target genes in undifferentiated myoblasts, prior to the activation of gene expression. MyoD-BAF60c complex is devoid of the ATP-dependent enzymatic sub-units Brg1 and Brm, is required for stable MyoD binding to Ebox sequences, and marks the chromatin for signal-dependent recruitment of the SWI/SNF core complex to muscle loci. BAF60c phosphorylation on a conserved threonine by differentiation-activated p38 signalling promotes the incorporation of MyoD-BAF60c into a Brg1-based SWI/SNF complex, which is competent to remodel the chromatin and activates transcription of MyoD-target genes. Our data support an unprecedented two-step model, by which pre-assembled BAF60c-MyoD complex directs the SWI/SNF complex chromatin re-distribution to muscle loci in response to differentiation cues.

overall_design: Differentiation of C2C12 cells individually interfered for BRG1, BAF60B, BAF60C


title: Gene expression induced by TSA in undifferentiated human primary myoblasts versus terminally differentiated human myotubes

status: Public on Mar 01 2011

summary: Gene expression analysis in human skeletal myoblasts (undifferentiated mononucleated cells, cultured in growth medium - 15% FBS) and human skeletal myotubes (pre-formed myotubes, cultured in differentiation medium – 2% horse serum) exposed to the HDACi TSA (50nM) for 24 hours

overall_design: TSA treated undifferentiated human primary myoblasts and terminally differentiated human myotubes

first: Pier

last: Puri

first: Giulia

last: Minetti


title: Wheat response to powdery mildew infection and heat stress

status: Public on Feb 17 2011

city: BEIJING

first: mingming

last: xin

address: YUAN MING YUAN WEST ROAD 2, HAIDIAN DISTRICT

department: plant genetics and breeding

institute: China Agricultrual University

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

overall_design: Refer to individual Series


title: Expression data from HTR-8/Svneo side population (SP) and non-side population (NSP) cells

status: Public on Mar 12 2011

city: Fukuoka

first: Tomoka

last: Takao

address: Maidashi3-1-1, Higashi-ku, Fukuoka

institute: Kyushu University

summary: Immature cell populations, including stem cells and progenitor cells, can be found in “side-population (SP)” cells. Although SP cells isolated from some adult tissues have been reported elsewhere, isolation and characterization of human trophoblast SP cells remained to be reported. We used microarrays to detail the global program of gene expression underlying cell differentiation and identified up-regulated genes of SP cells.

overall_design: HTR-8/SVneo SP cells or NSP cells were isolated using a cell sorter for RNA extraction and hybridization on Affymetrix microarrays. To obtain many SP or NSP cells, we cultured a large amount of HTR-8/SVneo cells (1×10^7 cells). The cells were labeled with 2.5µg/ml Hoechst 33342 dye (Molecular Probes, Eugene, OR) for 120 minutes at 37 ℃, either alone or in combination with 50 µmol/L verapamil (Sigma-Aldrich, ST Louis, MO). Finally, the cells were then passed through EPICS ALTRA HyPerSort (Beckman Coulter, Fullerton, CA) using dual wavelength analysis (blue, 424-444 nm; red, 675 nm) after excitation with 350 nm UV light.

first: Norio

last: Wake


title: Expression data in wheat (T. aestivum L.) near isogenic lines in response to powdery mildew infection

status: Public on Feb 15 2011

summary: We used two wheat genotypes, the susceptible wheat cultivar ‘8866 ’(S) and its near isogenic line with single powdery mildew resistance gene ‘pm30’ (R), to investigate gene expression changes in response to powdery mildew infection by using Wheat Genome Array

overall_design: wheat young leveas of near isogenic lines before or 12 hours after powdery mildew infection were selected for RNA extraction and hybridization on Affymetrix microarrays.The leaf samples were harvested from three independent biological replicates, and the leaves without inoculation were regarded as control.

first: Mingming

last: Xin

first: Qixin

last: Sun


title: NESG1-mediated differential expression genes in nasopharyngeal carcinoma (NPC)

city: guangzhou

first: weiyi

last: fang

address: tonghe road

institute: cancer institute

summary: Analysis of differential expression genes in NESG1-overexpressed and negative nasopharyngeal carcinoma. NESG1 is a candidate tumor suppressor in NPC. Results provide insight into the molecular pathogenesis of NESG1 in NPC.

overall_design: NESG1 cDNA was transfected into NESG1-negative NPC 5-8F cells and affymetrix microarrays HG-U133_Plus_2 were used to analyze the differential genes in NESG1-overexpressed NPC cells and their control NESG1-negative NPC cells.

first: Weiyi

last: Fang


title: Expression analysis of Trichoderma reesei QM9414 and delta-phlp1, delta-gnb1 and delta-gng1 in light and darkness.

status: Public on Mar 02 2011

city: Vienna

first: Doris

last: Tisch

state: Vienna

address: Gumpendorferstraße 1a

department: Chemical Engineering

country: Austria

institute: Technical University of Vienna

summary:
Investigation of whole genome gene expression level changes in response to different light conditions of the T. reesei QM9414 parental strain and the deletion strains delta-phlp1, delta-gnb1 and delta gng1, cultivated on 1 % microcrystalline cellulose.
The mutants analyzed in this study are further described in Tisch et al. 2011: Carbohydrate degradation is significantly regulated by light and the phosducin like protein PhLP1 in Trichoderma reesei (Hypocrea jecorina).

overall_design: We used two biological replicates of four T. reesei strains (QM9414, delta-phlp1, delta-gnb1 and delta-gng1), cultivated in constant light (LL, 1800 lux) or constant darkness (DD) on microcrystalline cellulose.

first: Monika

last: Schmoll


title: Identification of the regulatory circuit of PhoB in Bacteroides fragilis strain YCH46

status: Public on Feb 22 2011

city: tokushima

first: Haruyuki

last: Nakayama-Imaohji

address: 3-18-15 kuramoto-cho

institute: Tokushima University

summary: Comparison of transcriptional profiles of the wild-type (WT) and the phoB-mutant strain in B. fragilis strain YCH46 during phosphate (Pi) starvation.

sample_organism:
Bacteroides fragilis
Bacteroides fragilis YCH46

overall_design: A four chip study using total RNA isolated from the WT culture and the phoB mutant culture under Pi-excess (6.6 mM) or Pi-limiting (0.0066mM) condition. Each sample contains duplicated data.

first: Tomomi

last: Kuwahara

first: Hideki

last: Hirakawa


title: Genome-wide mapping of BES1 binding sites in Arabidopsis thaliana

status: Public on Jan 02 2011

city: Ames

first: Xiaofei

last: Yu

address: 1035D Roy J Carver Co-Lab

institute: Iowa state Univeristy

summary: Brassinosteroids (BRs) are important regulators for plant growth and development. BRs signal to control the activities of the BES1 and BZR1 family transcription factors. In order to further understand the mechanism by which BES1/BZR1 regulates downstream genes, we performed chromatin immunoprecipitation coupled with tiling arrays (ChIP-chip) to identify BES1 binding sites in the genome. By combining ChIP-chip data with gene expression microarray data, we are able to discover genes that are directly regulated by BES1 (i.e. BES1 target genes).

overall_design: Chromatin from bes1-D Arabidopsis seedlings are immunoprecipitated by anti-BES1 antibody with anti-GFP antibody as control.

first: Yanhai

last: Yin


title: Gene expression profiles after traumatic brain injury

status: Public on Dec 07 2010

city: Chuo

last: Shojo

state: Yamanashi

address: 1110 shimokato

department: Graduate School of Medical Science

institute: University of Yamanashi

summary:
Time dependent-profiles in the gene expression level following lateral moderate fluid percussion injury in the rat brain
We used microarray to elucidate relationship between the alteration of gene expression levels and the progression of brain damages following traumatic brain injury.

overall_design: To examine the levels of gene expression in the early phase of traumatic brain injury, we analyzed the gene expression at 3, 6, 12, and 48 h after trauma using the lateral moderate fluid percussion TBI model. The ratios of the gene expression level were compared between chips corresponding to the 3, 6 and 12 h fluid percussion groups and the sham group chips. On the other hand, the rations of gene expression level after 48 h FPI were compared with 48 h sham chip, because the gene expression levels of 48 h sham chip were distinct from sham group chips (3, 6 and 12 h) in the cluster and principal components analyses.


title: Sox10 controls migration of melanoma cells through multiple regulatory target genes

status: Public on Nov 20 2010

city: SEOUL

first: IK JOO

last: SEONG

address: Seodeamungu Deahyundong

department: Life Science

institute: Ewha woman university

summary: It is believed that the inherent differentiation program of melanocytes during embryogenesis predisposes melanoma cells to high frequency of metastasis. Sox10, a transcription factor expressed in neural crest stem cells and a subset of progeny lineages, plays a key role in the development of melanocytes. We show that B16F10 melanoma cells transfected with siRNA specific for Sox10 display reduced migratory activity which in turn indicated that a subset of transcriptional regulatory target genes of Sox10 are likely to be involved in migration and metastasis of melanoma cells. We carried out microarray-based gene expression profiling using Sox10-specific siRNA to identify regulatory targets and found that multiple genes including melanocortin-1 receptor (Mc1R) partake in the regulation of migration. We provide evidences that a significant portion of the effect of Sox10 on migration is mediated by Mitf, a transcription factor downstream to Sox10. The involvement of Mc1R in migration was studied in detail in vivo using a murine metastasis model. Specifically, B16F10 melanoma cells treated with a specific siRNA showed reduced tendency in metastasizing to and colonizing the lung after being injected in the tail vein. These data reveal a cadre of novel regulators and mediators involved in migration and metastasis of melanoma cells that represent potential targets of therapeutic intervention.

overall_design: Chemically synthesized siRNA duplex (WT1-Sox10) was used to knock-down the transcription factor Sox10 in murine melanoma cell line B16F10. For the control, siRNA containing 5 nucleotide alterations (MT1-Sox10) were used. Total RNA was subsequently prepared to synthesize probes for microarray screening. A total of three pairs of replicate samples were generated each from separate transfection followed by RNA preparation. Expression values were determined and compared within each pair of WT1-Sox10 and MT1-Sox10 transfections. Genes showing over 2 fold changes in all three replicate pairs were subjected to subsequent analyses.

first: Ik

last: Seong

first: Hyun

last: Min

first: Chang-Yeol

last: Yeo

first: Dong

last: Kang

first: Jae

last: Kim

first: Eun

last: Hwang

first: Eok-Soo

last: Oh


title: Transcriptional profile of Salmonella enterica serovars Kentucky and Enteritidis in response to acidic conditions

status: Public on Feb 01 2011

city: Irvine

first: Michael

last: McClelland

address: 132 Med Surge I

department: Microbiology & Molecular Genetics

institute: University of California, Irvine

summary: S. enterica sv Kentucky 3795 and S. enterica sv Enteritidis NalR were grown to mid-log phase in TSB, then subjected to three different acidic conditions generated by two different acids: HCl to pH4.5, HCl to pH5.5 and CH3COOH to pH5.5. After 10min, total RNA was harvested und compared to total RNA harvested from identical control cultures grown in TSB without the pH alteration.

sample_organism: Salmonella enterica subsp. enterica serovar Enteritidis

sample_organism: Salmonella enterica subsp. enterica serovar Kentucky

overall_design: At least three biological replicates were performed for each strain and acidic condition. Total RNAs were harvested, and labeled by the conventional protocol of Brown et al, then hybridized onto a Salmonella-specific PCR product array that covered over 97% of the Enteritidis genome.

first: Steffen

last: Porwollik

first: Rolf

last: Joerger

first: Jonathan

last: Frye


title: DEVELOPMENTAL MicroRNA EXPRESSION PROFILING OF MURINE EMBRYONIC OROFACIAL TISSUE

status: Public on Dec 01 2010

city: louisville

first: Guy

last: Brock

state: KY

address: 485 E. Gray St

department: Bioinformatics and Biostatistics

institute: University of Louisville

summary:
BACKGROUND: Orofacial development is a multifaceted process involving precise, spatio-temporal expression of a panoply of genes.  MicroRNAs (miRNAs) constitute the largest family of noncoding RNAs involved in gene silencing, and represent critical regulators of cell and tissue differentiation. MicroRNA gene expression profiling is an effective means of acquiring novel and valuable information regarding the expression and regulation of genes, under the control of miRNA, involved in mammalian orofacial development.
RESULTS: To identify differentially expressed miRNAs during mammalian orofacial ontogenesis, miRNA expression profiles from gestation day (GD) -12, -13 and -14 murine orofacial tissue were compared utilizing miRXplore™ microarrays from Miltenyi Biotech GmbH. TaqManTM quantitative Real-Time PCR was utilized for validation of gene expression changes.  Cluster analysis of the microarray data was conducted with the clValid R package and the UPGMA (hierarchical) clustering method.  Functional relationships between selected miRNAs were investigated using Ingenuity Pathway Analysis. Expression of over 26% of the approximately 588 murine miRNA genes examined was detected in murine orofacial tissues from GD 12, 13 and 14.  Among these expressed genes several clusters were seen to be developmentally regulated. Differential expression of genes encoding miRNAs within such clusters were shown to target genes encoding proteins involved in cell proliferation, cell adhesion, differentiation, apoptosis and epithelial-mesenchymal transformation, all processes critical for normal orofacial development. Functional relationships between miRNAs differentially expressed were investigated using Ingenuity Pathway Analysis (IPA; Ingenuity Systems).
CONCLUSIONS: Using miRNA microarray technology, unique gene expression signatures of hundreds of miRNAs in embryonic orofacial tissue were defined. Gene targeting and functional analysis revealed that the expression of numerous protein-encoding genes, crucial to normal orofacial ontogeny, may be regulated by specific miRNAs.

sample_organism:
Homo sapiens
Mus musculus
Rattus norvegicus

overall_design: Time-course experiment (Developmental Stages), ICR mice embryos on gestational days (GD) 12, 13 and 14.  Biological replicates: For each day of gestation, 3 independent pools of 15 to 20 staged embryos were used to procure embryonic orofacial tissues for preparation of 3 distinct pools of RNA that were independently processed and applied to individual miRXplore™ microRNA Microarray chips (Miltenyi Biotec GmbH).  Technology: 2-color spotted cDNA, Hy5 (experimental sample) vs. Hy3 (control - miRXplore Universal Reference).

first: Partha

last: Mukhopadhyay

middle: N

first: Vasyl

last: Pihur

first: Cynthia

last: Webb

first: M

last: Pisano

first: Robert

last: Greene


title: Interaction between circadian clocks and metabolic physiology: implications for reproductive fitness

status: Public on Oct 05 2010

city: Philadelphia

last: Hughes

state: PA

address: 421 Curie Bvd, Brb 835

institute: UPenn

summary: Circadian profiling of total RNA collected from wildtype and dnClk adult Drosophila fatbody

overall_design: 24 wild type and 12 dnClk samples, no replication

first: Kanyan

last: Xu

first: Justin

middle: R

last: DiAngelo

first: John

middle: B

last: Hogenesch

first: Amita

last: Sehgal


title: Drop-on-demand single cell isolation and total RNA analysis

status: Public on Oct 01 2010

first: Lingsheng

last: Dong

address: 75 Francis Street

department: Renal Division

institute: Brigham and Women's Hospital

summary: We present a microdroplet-based method to isolate and pattern single cells from heterogeneous cell suspensions (10% target cell mixture), preserve viability of the extracted cells (97.0 ± 0.8%), and obtain genomic information from isolated cells compared to the non-patterned controls.

overall_design: To evaluate the sensitivity and reproducibility of a functional genomic analysis of stem cell encapsulating droplets, we used DNA microarrays to measure gene expression levels in RNA samples extracted from mouse embryonic stem cells obtained by using drop-on-demand. We compared the results with RNA extracted from a control pool of mESCs, which were isolated with serial dilution and manual pipetting.

first: Sangjun

last: Moon

first: Yun-Gon

last: Kim

last: Lombardi

first: Edward

last: Haeggstrom

first: Roderick

middle: V

last: Jensen

first: Li-Li

last: Hsiao

first: Utkan

last: Demirci


title: Glucocorticoid target genes in 3T3-L1 adipocytes

status: Public on Sep 15 2010

city: Berkeley

first: Jen-Chywan

last: Wang

address: 315 Morgan Hall, UC Berkeley

department: Nutritional Sciences & Toxicology

institute: UC Berkeley

summary: Analysis of 3T3-L1 adipocytes treeated with dexamethasone (Dex) for 6 hours.  Dex is a synthetic glucorticoid (GC) receptor agonist. Results provide insight into the effect of glucocorticoids on adipocytes.

overall_design: Differentiated 3T3-L1 adipocytes  were cultured in DMEM supplemented with 5% stripped FBS for 24 hours and then treated with 500 nΜ Dex or an equal volume (0.05% v/v of media) of ethanol as a vehicle control for 6 hours at 37°C in DMEM with 190 5% stripped FBS. Total cellular RNA was isolated utilizing the NucleoSpin RNA II kit (Macherey-Nagel).RNA isolates were first quantified by standard spectrophotometry, and then qualitatively evaluated by capillary electrophoresis employing the Bio-Rad Experion system per the manufacturer’s instruction. The final labeled cRNA samples were hybridized overnight to Illumina mouseWG-6 BeadChip arrays, which was performed at UCSF Genomic Core.  All treatments were done in triplicates and the same batch of microarrays were used for all treatments. The Illumina expression arrays were pre-processed using lumi package. The differential expression analysis was performed using the Limma package.


title: Genome-wide sequencing small RNAs in response to powdery mildew and heat stress in wheat

first: mingming

last: xin

summary: To test whether non-coding RNAs play roles in regulating response to powdery mildew infection and heat stress in wheat, by using Solexa high-throughput sequencing and computational analysis and experimental approach we cloned the small RNAs and identified 125 putative long npcRNAs from wheat leaves infected by preponderant physiological strain Erysiphe graminis f. sp. tritici (Egt) or by heat stress treatment. Among long non-coding RNAs, some were precursors of small RNAs such as microRNAs and siRNAs, two long npcRNAs were identified as signal recognition particle (SRP) 7S RNA variants, and three were characterized as U3 snoRNAs. Wheat long npcRNAs showed tissue dependent expression patterns and were responsive to powdery mildew infection and heat stress.

overall_design: Examination non-coding RNAs of 2 near isogenic lines 8866 (Susceptible) and Pm30 (Resistant) in response to powdery milew and two genotypes CK (insensitive) and TAM107 (insensitive) to heat.  CK and TAM107 represent the same material in different treatments (no heat stress or 1hour after heat stress).

first: Mingming

last: Xin

first: Qixin

last: Sun


title: Tracheal occlusion conditioning in conscious rats modulates gene expression profile of medial thalamus

status: Public on Nov 05 2010

city: Gainesville

first: Vipa

last: Bernhardt

state: FL

address: 1600 Archer Rd

institute: University of Florida

summary: The thalamus may be the critical brain area involved in sensory gating and the relay of respiratory mechanical information to the cerebral cortex for the conscious awareness of breathing. We hypothesized that respiratory mechanical stimuli in the form of tracheal occlusions would modulate the gene expression profile of the thalamus. Specifically, it was reasoned that conditioning to the respiratory loading would induce a state change in the medial thalamus consistent with a change in sensory gating and the activation of molecular pathways associated with learning and memory. In addition, respiratory loading is stressful and thus should elicit changes in gene expressions related to stress, anxiety, and depression. Rats were instrumented with inflatable tracheal cuffs. Following surgical recovery, they underwent ten days (5 days/week) of transient tracheal occlusion conditioning. On day 10, the animals were sacrificed and the brains removed. The medial thalamus was dissected and microarray analysis of gene expression performed. Tracheal obstruction conditioning modulated a total of 661 genes (p < 0.05, log2 fold change ≥ 0.58), 250 genes were down-regulated and 411 up-regulated. There was a significant down-regulation of GAD1, GAD2 and HTR1A, HTR2A genes. CCK, PRKCG, mGluR4, and KCJN9 genes were significantly up-regulated. Some of these genes have been associated with anxiety and depression, while others have been shown to play a role in switching between tonic and burst firing modes in the thalamus and thus may be involved in gating of the respiratory stimuli. Furthermore, gene ontology and pathway analysis showed a significant modulation of learning and memory pathways. These results support the hypothesis that the medial thalamus is involved in the respiratory sensory neural pathway due to the state change of its gene expression profile following repeated tracheal occlusions.

overall_design: Four experimental animals that received tracheal occlusions and four surgical control animals were used. A reference design was used to compare all samples.

first: Mark

middle: T

last: Hotchkiss

first: Natalia

last: Garcia-Reyero

first: Nancy

last: Denslow

first: Paul

last: Davenport


title: Gene expression study identifies important protective pathways utilized by appendicitis and appendectomy for ameliorating colitis

status: Public on Sep 01 2010

city: Sydney

first: Rajkumar

last: Cheluvappa

state: NSW

address: Inflammation and Infection Research Centre, School of Medical Sciences, Wallace Wurth Building, University of New South Wales, Gate 9 High Street, Sydney, NSW 2052, Australia.

department: St George Clinical School

institute: University of New South Wales

summary:
BACKGROUND: Appendicitis followed by appendectomy (AA) at a young age protects against later inflammatory bowel disease (IBD). Using a novel murine appendicitis model we earlier demonstrated that AA proffered significant protection against subsequent experimental colitis.
AIM: To delineate genes and biological pathways involved in the protective effect of AA against subsequent colitis using gene set enrichment analysis (GSEA) of DNA microarray data.
METHODS:  Appendicitis and appendicectomy was done (5 week old male BALB/c mice) near the most proximal colon (caecal lymphoid follicles) and colonic samples were harvested from the most distal colon. Two consecutive laparotomies were done in control Sham-Sham (SS) mice. RNA was extracted (TRIzol®) from 4 individual colonic samples per group (AA group vs. SS group) with each sample taken independently through Affymetrix® microarray hybridization. For GSEA, data for more than 23,000 genes were exported from Partek and analyzed with GSEA software (with 2852 gene sets encoded) to establish correlates with phenotypes of the gene sets.
RESULTS: Distal colonic expression of 636 gene-sets were significantly upregulated in AA group samples (False Discovery Rates (FDR) values < 1 % and p value < 0.001; stringent statistical selection). These were validated by quantitative PCR of 14 selected genes across the immunological spectrum and over time-intervals of 3 days, 14 days and 28 days.
CONCLUSIONS:  Many key immunological, apoptosis-related and cellular function-associated gene-sets involved in the protective effect of AA in experimental colitis were identified. Further analysis of these profiles and biological pathways will assist utilizing these gene products and manipulating various aspects of these pathways to develop better therapeutic strategies in the management of intractable IBD.

overall_design:
Appendicitis and appendicectomy was done (5 week old male BALB/c mice) near the most proximal colon (caecal lymphoid follicles) and colonic samples were harvested from the most distal colon. Two consecutive laparotomies were done in control Sham-Sham (SS) mice. RNA was extracted (TRIzol®) from 4 individual colonic samples per group (AA group vs. SS group) with each sample taken independently through Affymetrix® microarray hybridization.
variable_protocol: appendicitis and appendectomy: AA1, AA2, AA3, AA4
variable_protocol: sham/sham surgery: SS1, SS2, SS3, SS4
repeat_biological replicate: AA1, AA2, AA3, AA4
repeat_biological replicate: SS1, SS2, SS3, SS4

Upregulated gene-set linked as supplementary file.

first: Annie

last: Luo

first: Clovis

last: Palmer

first: Warren

last: Kaplan

middle: J

last: Cowley

first: Michael

last: Grimm


title: Transcription in adult mouse tissues

status: Public on Nov 17 2010

city: Leuven

first: Lieven

last: Thorrez

address: Kasteelpark Arenberg 10

country: Belgium

institute: Katholieke Universiteit Leuven

summary:
We used Affymetrix Gene Arrays (1.0 ST) to compare gene expression across different murine tissues.
Arrays were used to find disallowed genes in tissues.

overall_design: Different tissues were dissected from 10-12 week old C57Bl6 mice for RNA extraction and hybridization on Affymetrix microarrays.

first: Leentje

last: Van Lommel

first: Katleen

last: Lemaire

first: Frans

last: Schuit


title: Genome-wide FLP/MYB88 binding sites in Arabidopsis thaliana

city: Columbus

first: Zidian

last: Xie

state: OH

address: 1060 Carmack Rd.

department: Dept. of Plant Cellular and Molecular Biology

institute: The Ohio State University

summary: FLP and MYB88 are two paralogous MYB proteins, regulating the symmetric division of guard mother cell during Arabidopsis stomatal development. To understand their molecular functions, we performed genome-wide identification of FLP/MYB88 binding targets using ChIP-chip with FLP/MYB88 antibody. By comparing ChIP-chip between wild-type and flp-1 myb88 lines, a total genes were identified as putative direct targets for FLP/MYB88.

overall_design: Ten-day old wild-type and flp-1 myb88 Arabidopsis green tissues were used for ChIP. Rabbit polyclonal antibody recognizing FLP and MYB88 proteins was used for chromatin immunoprecipitation

first: Erich

last: Grotewold


title: Aedes aegypti whole-genome gene expression

status: Public on Jul 05 2010

first: Osvaldo

last: Marinotti

address: University of California, Irvine

department: Molecular Biology & Biochemistry

summary: Sequencing of the Aedes aegypti genome has enabled genome-wide studies of gene expression in this mosquito.  The large quantities of data produced from such studies require efficient cataloguing in order for new insight to be made into gene expression patterns and the underlying molecular mechanisms for producing these patterns.  Our study provides a comprehensive catalogue of genes whose transcription products increase or decrease in abundance in adult females following blood feeding. We developed a publicly-accessible database and data-mining tool, aeGEPUCI, that integrates 1) stage-specific microarray analyses of gene expression in Ae. aegypti, 2) functional gene annotation, 3) genomic sequence data, and 4) computational sequence analysis tools.  The database is accessible from the address http://www.aegep.bio.uci.edu.

overall_design: 8 conditions, 3 replicates for each condition

first: Anthony

middle: A

last: James


status: Public on Jul 19 2010

city: Chapel Hill

first: Jen Jen

last: Yeh

state: NC

address: LCCC, 450 West Drive

department: Surgical Oncology

institute: University of North Carolina at Chapel Hill

summary: A six-gene signature predicts survival of patients with localized pancreatic ductal adenocarcinoma Pancreatic ductal adenocarcinoma (PDAC), comprising over 90% of all pancreatic cancers, remains a lethal disease with an estimated 232,000 new cases, 227,000 deaths per year worldwide, and a less than 5% five-year survival rate. Currently the standard of care for the 20% of patients with localized disease is surgery followed by chemotherapy, and in some cases radiation. Unfortunately despite the use of adjuvant therapy, median survival remains at best 23 months. It is important to note however, that up to 27% of patients with resected PDAC can survive for five years. However, in these studies examining actual long-term survivors, only two have found that adjuvant therapy was associated with improved survival. In addition, randomized controlled trials of gemcitabine-based chemotherapy demonstrate an improvement in median survival of at best 3 months. One possible conclusion from these studies is that tumor biology dictates outcome and that our current adjuvant therapy has only a modest impact on altering a patient's course.Hypothesizing that the dismal outcome of patients with localized disease is due to the presence of micrometastasic disease, current clinical investigation has focused on preoperative or neoadjuvant therapy. This approach, where patients who cannot tolerate the stress of therapy or develop metastatic disease during treatment are spared surgery, has demonstrated an overall survival of 34 months in this highly selected patient population. Therefore the ability to select patients who would most benefit from a neoadjuvant approach may be important. One way to do this is to define a prognostic gene signature that can identify patients with more aggressive tumor biology upfront.

overall_design:
reference x sample
The 30 Nebraska and UNC samples were not analyzed with clinical data so it is not provided. One of the PE samples is missing some clinical data.

first: Jeran

middle: K

last: Stratford

first: Jen


title: Expression data of multiple sclerosis patients receiving subcutaneous Interferon-beta-1b therapy [U133 A and B]

status: Public on Sep 29 2010

city: Rostock

first: Robert

last: Goertsches

address: Gehlsheimer Strasse 20

department: Neurology

institute: University

summary:
The purpose of this study was to characterize the transcriptional effects induced by subcutaneous IFN-beta-1b treatment (Betaferon, 250 µg every other day) in patients with relapsing-remitting form of multiple sclerosis (MS).
By using Affymetrix DNA microarrays, we obtained genome-wide expression profiles of peripheral blood mononuclear cells from 25 MS patients within the first two years of IFN-beta administration.

overall_design:
EDTA blood samples were drawn from all patients immediately before first, second, month 1, 12, and 24 IFN-beta injection.
Total RNA of Ficoll-isolated peripheral blood mononuclear cells from each sample was extracted, labeled and hybridized to Affymetrix Human Genome U133 A and B arrays to quantify the mRNA levels.

middle: H

last: Hecker

first: Dirk

last: Koczan

first: Uwe

last: Zettl