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Long non-coding RNAs and enhancer RNAs regulate the lipopolysaccharide-induced inflammatory response in human monocytes.

IIott NE, Heward JA, Roux B, Tsitsiou E, Fenwick PS, Lenzi L, Goodhead I, Hertz-Fowler C, Heger A, Hall N, Donnelly LE, Sims D, Lindsay MA - Nat Commun (2014)

Bottom Line: Early reports indicate that long non-coding RNAs (lncRNAs) are novel regulators of biological responses.We identify 76 enhancer RNAs (eRNAs), 40 canonical lncRNAs, 65 antisense lncRNAs and 35 regions of bidirectional transcription (RBT) that are differentially expressed in response to bacterial lipopolysaccharide (LPS).Crucially, we demonstrate that knockdown of nuclear-localized, NF-κB-regulated, eRNAs (IL1β-eRNA) and RBT (IL1β-RBT46) surrounding the IL1β locus, attenuates LPS-induced messenger RNA transcription and release of the proinflammatory mediators, IL1β and CXCL8.

View Article: PubMed Central - PubMed

Affiliation: 1] CGAT Programme, MRC Functional Genomics Unit, Department of Physiology, Anatomy, and Genetics, University of Oxford, Oxford OX1 3QX, UK [2].

ABSTRACT
Early reports indicate that long non-coding RNAs (lncRNAs) are novel regulators of biological responses. However, their role in the human innate immune response, which provides the initial defence against infection, is largely unexplored. To address this issue, here we characterize the long non-coding RNA transcriptome in primary human monocytes using RNA sequencing. We identify 76 enhancer RNAs (eRNAs), 40 canonical lncRNAs, 65 antisense lncRNAs and 35 regions of bidirectional transcription (RBT) that are differentially expressed in response to bacterial lipopolysaccharide (LPS). Crucially, we demonstrate that knockdown of nuclear-localized, NF-κB-regulated, eRNAs (IL1β-eRNA) and RBT (IL1β-RBT46) surrounding the IL1β locus, attenuates LPS-induced messenger RNA transcription and release of the proinflammatory mediators, IL1β and CXCL8. We predict that lncRNAs can be important regulators of the human innate immune response.

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LncRNAs can be distinguished by canonical promoter and enhancer chromatin signatures.(a) H3k4me3 and H3K4me1 binding across a 1 Kb interval centred on the transcription start site of expressed protein-coding genes. Profiles are sorted based on the height of the H3K4me3 peak. Also provided is a plot of the H3K4me1/H3K4me3 log2(ratio) at each TSS (mean over interval). (b) H3k4me3 and H3K4me1 binding across a 1 Kb interval centred on the transcription start site of differentially expressed lncRNAs. Profiles are sorted on the height of the H3K4me3 peak. Also provided is a plot of the H3K4me1/H3K4me3 log2(ratio) at each TSS (mean). (c) Example of a lncRNA with a canonical promoter-like chromatin signature (can-lncRNA, left) and a lncRNA with an enhancer signature (eRNA, right). Figures were produced using the UCSC genome browser.
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f2: LncRNAs can be distinguished by canonical promoter and enhancer chromatin signatures.(a) H3k4me3 and H3K4me1 binding across a 1 Kb interval centred on the transcription start site of expressed protein-coding genes. Profiles are sorted based on the height of the H3K4me3 peak. Also provided is a plot of the H3K4me1/H3K4me3 log2(ratio) at each TSS (mean over interval). (b) H3k4me3 and H3K4me1 binding across a 1 Kb interval centred on the transcription start site of differentially expressed lncRNAs. Profiles are sorted on the height of the H3K4me3 peak. Also provided is a plot of the H3K4me1/H3K4me3 log2(ratio) at each TSS (mean). (c) Example of a lncRNA with a canonical promoter-like chromatin signature (can-lncRNA, left) and a lncRNA with an enhancer signature (eRNA, right). Figures were produced using the UCSC genome browser.

Mentions: Previous studies have reported the presence of transcription at active enhancers marked by H3K4me1 (ref. 18). While the poised/active promoter-associated mark, H3K4me3, may also be present at distal enhancer loci24, the ratio of H3K4me1/H3K4me3 is commonly used as a discriminatory mark between enhancers and promoters (reviewed in refs 14, 17). To explore whether our differentially expressed lncRNAs represent transcription from enhancer-like regions, we utilized recently available histone modification ChIP-seq data from the ENCODE consortium. We downloaded alignments for H3K4me1 and H3K4me3 ChIP-seq data in CD14+ resting monocytes and assessed the read coverage over intervals surrounding the transcription start site (TSS, ±0.5 Kb) of differentially expressed lncRNAs. An equivalent analysis using differentially expressed protein-coding genes (n=530) provided a comparison set. To eliminate confounding influences of marks associated with protein-coding genes on lncRNA marks, we removed lncRNAs that either overlapped (that is, antisense) or shared a TSS interval (<2 Kb) with protein-coding genes. This resulted in the subsequent analysis of 132/221 lncRNAs (see Supplementary Table 4 online). We hypothesized that a subset of our lncRNAs would display canonical mRNA-like promoter histone signatures (H3K4me1/H3K4me3 low) and a subset, representing enhancer RNAs (eRNAs), would display enhancer-like signatures (H3K4me1/H3K4me3 high). As expected, the majority of differentially expressed protein-coding genes displayed punctate binding of H3k4me3 around the TSS (Fig. 2a). In contrast, there was a relatively weaker signal for this histone modification at lncRNA TSS intervals, with only a small subset displaying a high H3K4me3 density (Fig. 2b). There was a stronger H3K4me1 signal across intervals for lncRNAs than protein-coding genes, with more dispersed binding around the TSS than for H3K4me3 (Fig. 2a,b). To investigate transcripts derived from enhancer regions, we calculated the ratio of H3K4me1/H3K4me3. The distribution of H3K4me1/H3K4me3 for protein-coding genes was markedly different from that of lncRNAs (Fig. 2a,b). Protein-coding genes predominantly displayed characteristic promoter marks whereas a large proportion of the investigated lncRNAs were associated with a dominant H3K4me1 histone mark (Fig. 2a,b), suggestive of transcription occurring from enhancer regions. Using an H3K4me1/H3K4me3 ratio of >1.2 and <0.8 to define enhancer and promoter states, respectively, we were able to show that 76 lncRNAs (58%) were putative eRNAs, 40 (30%) had canonical promoter signatures and 16 (12%) could not be assigned to either group, that is, 0.8<H3K4me1/H3K4me3<1.2) (see Supplementary Table 4 online). Hereafter, we refer to those RNA transcripts that have higher levels of H3K4me1 compared with H3K4me3 as eRNAs, which differ from canonical lncRNAs (can-lncRNAs), where the reverse is true (Fig. 2c).


Long non-coding RNAs and enhancer RNAs regulate the lipopolysaccharide-induced inflammatory response in human monocytes.

IIott NE, Heward JA, Roux B, Tsitsiou E, Fenwick PS, Lenzi L, Goodhead I, Hertz-Fowler C, Heger A, Hall N, Donnelly LE, Sims D, Lindsay MA - Nat Commun (2014)

LncRNAs can be distinguished by canonical promoter and enhancer chromatin signatures.(a) H3k4me3 and H3K4me1 binding across a 1 Kb interval centred on the transcription start site of expressed protein-coding genes. Profiles are sorted based on the height of the H3K4me3 peak. Also provided is a plot of the H3K4me1/H3K4me3 log2(ratio) at each TSS (mean over interval). (b) H3k4me3 and H3K4me1 binding across a 1 Kb interval centred on the transcription start site of differentially expressed lncRNAs. Profiles are sorted on the height of the H3K4me3 peak. Also provided is a plot of the H3K4me1/H3K4me3 log2(ratio) at each TSS (mean). (c) Example of a lncRNA with a canonical promoter-like chromatin signature (can-lncRNA, left) and a lncRNA with an enhancer signature (eRNA, right). Figures were produced using the UCSC genome browser.
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f2: LncRNAs can be distinguished by canonical promoter and enhancer chromatin signatures.(a) H3k4me3 and H3K4me1 binding across a 1 Kb interval centred on the transcription start site of expressed protein-coding genes. Profiles are sorted based on the height of the H3K4me3 peak. Also provided is a plot of the H3K4me1/H3K4me3 log2(ratio) at each TSS (mean over interval). (b) H3k4me3 and H3K4me1 binding across a 1 Kb interval centred on the transcription start site of differentially expressed lncRNAs. Profiles are sorted on the height of the H3K4me3 peak. Also provided is a plot of the H3K4me1/H3K4me3 log2(ratio) at each TSS (mean). (c) Example of a lncRNA with a canonical promoter-like chromatin signature (can-lncRNA, left) and a lncRNA with an enhancer signature (eRNA, right). Figures were produced using the UCSC genome browser.
Mentions: Previous studies have reported the presence of transcription at active enhancers marked by H3K4me1 (ref. 18). While the poised/active promoter-associated mark, H3K4me3, may also be present at distal enhancer loci24, the ratio of H3K4me1/H3K4me3 is commonly used as a discriminatory mark between enhancers and promoters (reviewed in refs 14, 17). To explore whether our differentially expressed lncRNAs represent transcription from enhancer-like regions, we utilized recently available histone modification ChIP-seq data from the ENCODE consortium. We downloaded alignments for H3K4me1 and H3K4me3 ChIP-seq data in CD14+ resting monocytes and assessed the read coverage over intervals surrounding the transcription start site (TSS, ±0.5 Kb) of differentially expressed lncRNAs. An equivalent analysis using differentially expressed protein-coding genes (n=530) provided a comparison set. To eliminate confounding influences of marks associated with protein-coding genes on lncRNA marks, we removed lncRNAs that either overlapped (that is, antisense) or shared a TSS interval (<2 Kb) with protein-coding genes. This resulted in the subsequent analysis of 132/221 lncRNAs (see Supplementary Table 4 online). We hypothesized that a subset of our lncRNAs would display canonical mRNA-like promoter histone signatures (H3K4me1/H3K4me3 low) and a subset, representing enhancer RNAs (eRNAs), would display enhancer-like signatures (H3K4me1/H3K4me3 high). As expected, the majority of differentially expressed protein-coding genes displayed punctate binding of H3k4me3 around the TSS (Fig. 2a). In contrast, there was a relatively weaker signal for this histone modification at lncRNA TSS intervals, with only a small subset displaying a high H3K4me3 density (Fig. 2b). There was a stronger H3K4me1 signal across intervals for lncRNAs than protein-coding genes, with more dispersed binding around the TSS than for H3K4me3 (Fig. 2a,b). To investigate transcripts derived from enhancer regions, we calculated the ratio of H3K4me1/H3K4me3. The distribution of H3K4me1/H3K4me3 for protein-coding genes was markedly different from that of lncRNAs (Fig. 2a,b). Protein-coding genes predominantly displayed characteristic promoter marks whereas a large proportion of the investigated lncRNAs were associated with a dominant H3K4me1 histone mark (Fig. 2a,b), suggestive of transcription occurring from enhancer regions. Using an H3K4me1/H3K4me3 ratio of >1.2 and <0.8 to define enhancer and promoter states, respectively, we were able to show that 76 lncRNAs (58%) were putative eRNAs, 40 (30%) had canonical promoter signatures and 16 (12%) could not be assigned to either group, that is, 0.8<H3K4me1/H3K4me3<1.2) (see Supplementary Table 4 online). Hereafter, we refer to those RNA transcripts that have higher levels of H3K4me1 compared with H3K4me3 as eRNAs, which differ from canonical lncRNAs (can-lncRNAs), where the reverse is true (Fig. 2c).

Bottom Line: Early reports indicate that long non-coding RNAs (lncRNAs) are novel regulators of biological responses.We identify 76 enhancer RNAs (eRNAs), 40 canonical lncRNAs, 65 antisense lncRNAs and 35 regions of bidirectional transcription (RBT) that are differentially expressed in response to bacterial lipopolysaccharide (LPS).Crucially, we demonstrate that knockdown of nuclear-localized, NF-κB-regulated, eRNAs (IL1β-eRNA) and RBT (IL1β-RBT46) surrounding the IL1β locus, attenuates LPS-induced messenger RNA transcription and release of the proinflammatory mediators, IL1β and CXCL8.

View Article: PubMed Central - PubMed

Affiliation: 1] CGAT Programme, MRC Functional Genomics Unit, Department of Physiology, Anatomy, and Genetics, University of Oxford, Oxford OX1 3QX, UK [2].

ABSTRACT
Early reports indicate that long non-coding RNAs (lncRNAs) are novel regulators of biological responses. However, their role in the human innate immune response, which provides the initial defence against infection, is largely unexplored. To address this issue, here we characterize the long non-coding RNA transcriptome in primary human monocytes using RNA sequencing. We identify 76 enhancer RNAs (eRNAs), 40 canonical lncRNAs, 65 antisense lncRNAs and 35 regions of bidirectional transcription (RBT) that are differentially expressed in response to bacterial lipopolysaccharide (LPS). Crucially, we demonstrate that knockdown of nuclear-localized, NF-κB-regulated, eRNAs (IL1β-eRNA) and RBT (IL1β-RBT46) surrounding the IL1β locus, attenuates LPS-induced messenger RNA transcription and release of the proinflammatory mediators, IL1β and CXCL8. We predict that lncRNAs can be important regulators of the human innate immune response.

Show MeSH
Related in: MedlinePlus