Unlinking a lncRNA from its associated cis element
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ABSTRACT:
PROVIDER: PRJNA305598 | ENA |
REPOSITORIES: ENA
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Project description:Capture-C using probes at the Cdkn1b promoter and the Lockd promoter Many long non-coding (lnc) RNAs are reported to regulate gene expression and protein functions. However, the proportion of lncRNAs with biological activities among the thousands expressed in mammalian cells is controversial. We studied Lockd (LncRNA downstream of Cdkn1b), a 434 bp polyadenylated lncRNA originating 4 kb 3â to the Cdkn1b gene. Heterozygous and homozygous deletion of the 25 kb Lockd locus reduced Cdkn1b transcription by approximately 35 and 70% respectively in a mouse erythroid cell line. In contrast, homozygous insertion of a polyadenylation cassette 80 bp downstream of the Lockd transcription start site reduced the entire lncRNA transcript level by > 90%, but had no effect on Cdkn1b transcription. The promoter of the Lockd gene contains a DNase hypersensitive site, binds numerous transcription factors (TFs), and physically associates with the Cdkn1b promoter in chromosomal conformation capture (NG Capture-C) studies. Thus, the Lockd gene positively regulates Cdkn1b transcription through an enhancer-like cis element, while the lncRNA itself is dispensable. These findings demonstrate that the biological activities of a lncRNA cannot be inferred from phenotypes that arise after deleting the corresponding gene. Rather, the model of an inert transcript arising from a functional genomic cis element should be considered while investigating the biology of any lncRNA.
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Project description:Transcriptome analysis of effect of Lockd knockout on cells Many long non-coding (lnc) RNAs are reported to regulate gene expression and protein functions. However, the proportion of lncRNAs with biological activities among the thousands expressed in mammalian cells is controversial. We studied Lockd (Downstream of p27), a 434 bp polyadenylated lncRNA originating 4 kb 3’ to the Cdkn1b gene. Heterozygous and homozygous deletion of the 25 kb Lockd locus reduced Cdkn1b transcription by approximately 35 and 70% respectively in a mouse erythroid cell line. In contrast, homozygous insertion of a polyadenylation cassette 80 bp downstream of the Lockd transcription start site reduced the entire lncRNA transcript level by > 90%, but had no effect on Cdkn1b transcription. The 5’ region of the Lockd gene contains a DNase hypersensitive site, binds numerous transcription factors (TFs), and physically associates with the Cdkn1b promoter in chromosomal conformation capture (NG Capture-C) studies. Thus, the Lockd gene positively regulates Cdkn1b transcription through an enhancer-like cis element and not via the lncRNA transcript. These findings demonstrate that the biological functions of a lncRNA cannot be inferred simply from phenotypes that arise after deleting the corresponding genomic locus.
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Project description:Innate lymphoid cells (ILCs) comprise three groups of recently identified tissue resident immune cell lineages that play critical roles in protective immune responses and tissue homeostasis. While significant progress has been made in defining the key protein mediators of ILC development and function, how cis-acting epigenetic regulatory elements or long non-coding RNAs (lncRNAs) regulate ILCs is unknown. Herein, we describe a cis-regulatory element demarcated by a novel lncRNA that controls the maturation, function and lineage identity of group 1 ILCs while being dispensable for early ILC development and homeostasis of mature ILC2s and ILC3s. We named this ILC1-restricted lncRNA Rroid. The Rroid locus controls the functional specification and lineage identity of ILC1 by promoting chromatin accessibility and STAT5 deposition at the promoter of its neighboring gene, Id2, in response to the ILC1-specific cytokine IL-15.
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Project description:ILC1 lineage identity is determined by a cis-regulatory element marked by a novel lncRNA
| PRJNA394398 | ENA
Project description:Thymic central tolerance is essential to preventing autoimmunity. In medullary thymic epithelial cells (mTECs), the Autoimmune regulator (Aire) gene plays an essential role in this process by driving the expression of a diverse set of tissue-specific antigens (TSAs), which are presented and help tolerize self-reactive thymocytes. Interestingly, Aire has a highly tissue-restricted pattern of expression, with only mTECs and peripheral extrathymic Aire-expressing cells (eTACs) known to express detectable levels in adults. Despite this high level of tissue specificity, the cis-regulatory elements that control Aire expression have remained obscure. We used sequence conservation analysis and ChIP-seq against the enhancer-associated histone mark H3K27ac to identify a candidate Aire cis-regulatory element. There is enrichment of H3K27ac near this element, ACNS1, in mTECs and the element also has characteristics of being NF-κB-responsive. Finally, we find that this element is essential for Aire expression in vivo and necessary to prevent spontaneous autoimmunity, reflecting the importance of this regulatory DNA element in promoting immune tolerance.
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