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A long noncoding RNA lincRNA-EPS acts as a transcriptional brake to restrain inflammation

ABSTRACT: LincRNA-EPS is a long noncoding RNA that is expressed in macrophages and downregulated upon exposure to diverse microbial products. Analysis of macrophages from lincRNA-EPS-deficient mice revealed a specific role for this lincRNA in restraining immune response gene (IRG) expression. Mechanistically, lincRNA-EPS associates with chromatin at regulatory regions of IRGs to repress their transcription. To gain insight into the molecular function of lincRNA-EPS on chromatin, we generated ATAC-Seq libraries from wildtype and lincRNA-EPS-deficient macrophages stimulated with LPS.

ORGANISM(S): Mus musculus

PROVIDER: GSE78873 | GEO | 2016/05/08

SECONDARY ACCESSION(S): PRJNA314247

REPOSITORIES: GEO

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A long noncoding RNA lincRNA-EPS acts as a transcriptional brake to restrain inflammation

Project description:RNA-seq was performed in biological replicates (2 mice per group) of wild-type and lincRNA-EPS-/- BMDMs at 0, 2, and 6 hours after LPS treatment (100 ng/ml). RNA-seq libraries were prepared as described (Heyer et al., 2015). Briefly, ribosomal RNA (rRNA) was depleted using the Ribozero rRNA removal kit (Epicentre). Purified mRNAs were fragmented with RNA fragmentation reagent (Life Technologies) for 4 minutes and 30 seconds at 70C to obtain 100-150 nt long RNA fragments. After ethanol precipitation and washing, RNAs were re-suspended in 5 l of water and the 3 ends dephosphorylated using PNK (New England BioLabs) for 1 hr at 37C. RNA fragments with a 3 OH were then ligated to a preadenylated DNA adaptor using T4 RNA ligase 2, truncated K227Q (NEB). Following this, ligated RNAs were reverse-transcribed with Superscript III (Invitrogen) using a bar-coded reverse-transcription primer that anneals to the preadenylated adaptor. After reverse-transcription, gel purified cDNAs were circularized using CircLigase I (Epicentre) and PCR amplified using paired-end primers PE1.0 and PE2.0 (Illumina) for 14 cycles. PCR amplicons were gel purified and submitted for sequencing on the Illumina Hiseq2000. Tophat version 2.0.12 was used to align single sequence reads to version 73 of the Ensembl mouse genome (mm10) with options: --library-type fr-firststrand -g 1 -x 1 --read-mismatches 2. Cufflinks version 2.1.1 was used to estimate RNA abundances with Ensembl version 73 GTF and the --library-type fr-firststrand option. The Cuffdiff program was used to perform differential expression analysis between wild-type BMDMs and lincRNA-EPS-/- BMDMs at each corresponding time (0,2, and 6 hours after LPS treatment). To compute fold-change values for genes/transcripts with FPKM values of zero, a pseudocount of 1 was added to all FPKM values first.

2016-07-29 | E-MTAB-4088 | biostudies-arrayexpress

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A Long Noncoding RNA lincRNA-EPS Acts as a Transcriptional Brake to Restrain Inflammation.

Project description:Long intergenic noncoding RNAs (lincRNAs) are important regulators of gene expression. Although lincRNAs are expressed in immune cells, their functions in immunity are largely unexplored. Here, we identify an immunoregulatory lincRNA, lincRNA-EPS, that is precisely regulated in macrophages to control the expression of immune response genes (IRGs). Transcriptome analysis of macrophages from lincRNA-EPS-deficient mice, combined with gain-of-function and rescue experiments, revealed a specific role for this lincRNA in restraining IRG expression. Consistently, lincRNA-EPS-deficient mice manifest enhanced inflammation and lethality following endotoxin challenge in vivo. lincRNA-EPS localizes at regulatory regions of IRGs to control nucleosome positioning and repress transcription. Further, lincRNA-EPS mediates these effects by interacting with heterogeneous nuclear ribonucleoprotein L via a CANACA motif located in its 3' end. Together, these findings identify lincRNA-EPS as a repressor of inflammatory responses, highlighting the importance of lincRNAs in the immune system.

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A long noncoding RNA lincRNA-EPS acts as a transcriptional brake to restrain inflammation

Project description:A long noncoding RNA lincRNA-EPS acts as a transcriptional brake to restrain inflammation

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