Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Genomic expression analysis of human monocyte-derived DCs and monocytes

ABSTRACT: To identify the functional non-coding RNAs in human DC development, we explored the gene expression profiles during DC development from peripheral monocytes. Using RNA-seq, we identified many annotated lncRNAs with markedly altered expressions during human DC differentiation and maturation. RNA samples from human peripheral blood monocytes and monocyte-derived DCs were collected. RNAs from 3 donors were pooled together to form monocyte sample or DC sample and then these two samples were subjected to RNA-seq and analysis.

ORGANISM(S): Homo sapiens

SUBMITTER: PIN WANG

PROVIDER: E-GEOD-54401 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

The STAT3-binding long noncoding RNA lnc-DC controls human dendritic cell differentiation.

Wang Pin P Xue Yiquan Y Han Yanmei Y Lin Li L Wu Cong C Xu Sheng S Jiang Zhengping Z Xu Junfang J Liu Qiuyan Q Cao Xuetao X

Science (New York, N.Y.) 20140401 6181

Long noncoding RNAs (lncRNAs) play important roles in diverse biological processes; however, few have been identified that regulate immune cell differentiation and function. Here, we identified lnc-DC, which was exclusively expressed in human conventional dendritic cells (DCs). Knockdown of lnc-DC impaired DC differentiation from human monocytes in vitro and from mouse bone marrow cells in vivo and reduced capacity of DCs to stimulate T cell activation. lnc-DC mediated these effects by activatin ...[more]

PMID: 24744378

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Project description:The immune system relies on the plasticity of its components to produce appropriate responses to frequent environmental challenges. Dendritic cells (DCs) are critical initiators of innate immunity and orchestrate the later and more specific adaptive immunity. The generation of diversity in transcriptional programs is central for effective immune responses. Alternative splicing is widely considered a key generator of transcriptional and proteomic complexity, but its role has been rarely addressed systematically in immune cells. Here we used splicing-sensitive arrays to assess genome-wide gene- and exon-level expression profiles in human DCs in response to a bacterial challenge. We find widespread alternative splicing events and splicing factor transcriptional signatures induced by an E. coli challenge to human DCs. Alternative splicing acts in concert with transcriptional modulation, but these two mechanisms of gene regulation affect primarily distinct functional gene groups. Alternative splicing is likely to have an important role in DC immunobiology because it affects genes known to be involved in DC development, endocytosis, antigen presentation and cell cycle arrest 6 samples were analysed. 3 were used as controls (non-stimulated replicates) and 3 were stimulated with E.coli for 3 distinct periods: (T1) 4 hours, to cover early changes that occur after the bacterial challenge; (T2) 8 hours, to assess intermediate changes; and (T3) 18 hours, to measure the late response stage. Microarray data pre-processing and summarization was performed using AltAnalyze application, which includes several rigorous statistical methods for data filtering, summarization and determination of differential splicing. Briefly, low quality probesets were removed according to detection above background (DABG) p-value and absolute expression values. Next, probesets were summarized with RMA algorithm and using the AltAnalyze annotations, derived from Ensembl [29] and UCSC databases [30]. Gene expression levels were determined using only probesets targeting constitutive exons. Determination of overall gene level variation was assessed using linear models and empirical Bayes methods s implemented in M-bM-^@M-^XlimmaM-bM-^@M-^Y package. The B-statistics gives the log odds of differential expression and it requires an M-bM-^@M-^Xa prioriM-bM-^@M-^Y value for the estimated proportion of differentially expressed genes. To determine this value, we visually inspected the volcano plot, which compares biological significance (represented by fold-changes) with statistical significance (B-values), finding the value which enabled genes to be distinguished from the majority (logFCM-bM-^IM-$~1.585). Additionally, we verified the P-values corresponding to moderated F-statistics. Using the Benjamini and Hochberg method, all genes selected as differentially expressed had adjusted P-values <0.05. . Detection of alternative splicing variations were determined using splicing-index and FIRMA approaches, calculating MiDAS and normalized intensity p-values (p-value <0.05).

Project description:Purpose: Next-generation sequencing (NGS) has revolutionized systems-based analysis of cellular pathways. The goals of this study were to compare signals from competent and abnormal human embryos impacted differently on the expression of endometrial receptivity genes in mouse uteri using transcriptome profiling (RNA-seq). Methods: Immature female (25 d C57BL/6) mice were given a hormone treatment of a single dose 1 mg progesterone and 10 mg/kg/day ÃÂ²-estradiol for a total of 3d to prime the uterus for embryo transfer. The uterine horns of control and study mice were injected with an equal volume (50 ÃÂ¼l) of either unconditioned embryo culture medium (ECM), serving as controls, or pooled conditioned media from competent (n = 9) or arresting embryos (n = 18). Uterine mRNA profiles of 25-day-old wild-type (WT) were generated by deep sequencing, in triplicate, using Illumina HiSeq 2000 platform. The sequence reads that passed quality filters were analyzed with the following methods: Bowtie Alignment followed by TopHat (splice juntions mapper) and Cufflinks (transcript abundance). qRTÃ¢ÂÂPCR validation was performed using SYBR Green assays. Results: Using an optimized data analysis workflow, we mapped about 30 million sequence reads per sample to the mouse genome (build mm9) and identified 16,014 transcripts in the uteri WT and NrlÃ¢ÂÂ/Ã¢ÂÂ mice with BWA workflow and 34,115 transcripts with TopHat workflow. RNA-seq data confirmed stable expression of 25 known housekeeping genes, and 12 of these were validated with qRTÃ¢ÂÂPCR. RNA-seq data had a linear relationship with qRTÃ¢ÂÂPCR for more than four orders of magnitude and a goodness of fit (R2) of 0.8798. Approximately 10% of the transcripts showed differential expression between the WT and NrlÃ¢ÂÂ/Ã¢ÂÂ retina, with a fold change Ã¢ÂÂ¥1.5 and p value <0.05. Altered expression of 25 genes was confirmed with qRTÃ¢ÂÂPCR, demonstrating the high degree of sensitivity of the RNA-seq method. Hierarchical clustering of differentially expressed genes uncovered several as yet uncharacterized genes that may contribute to retinal function. Data analysis with BWA and TopHat workflows revealed a significant overlap yet provided complementary insights in transcriptome profiling. Conclusions: Our study represents the first detailed analysis of maternal uterine reponse to signals from competent and abnormal human embryos, with biological replicates, generated by RNA-seq technology. Supernatant from competent and abnormal human embryos were flushed into the uterine horns of 25d old C57BL/6 female mice. Uterine mRNA profiles of 21-day old mice were generated by deep sequencing, in triplicate, using Illumina HiSeq 2000 platform (HiSeq).

Dataset Information

Genomic expression analysis of human monocyte-derived DCs and monocytes

Publications

The STAT3-binding long noncoding RNA lnc-DC controls human dendritic cell differentiation.

Similar Datasets

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