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Unlinking a lncRNA from its associated cis element [gene expression]

ABSTRACT: 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.

ORGANISM(S): Mus musculus

PROVIDER: GSE75449 | GEO | 2016/06/01

SECONDARY ACCESSION(S): PRJNA304311

REPOSITORIES: GEO

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Unlinking a lncRNA from its associated cis element [Capture-C]

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.

2016-06-01 | E-GEOD-75880 | biostudies-arrayexpress

Unlinking a lncRNA from its associated cis element [gene expression]

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 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 5â 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. We analyzed mouse G1E erythroid cell line clones with Control Lockd (C - 3 replicates) and with Lockd deletion with CRISPR (KO - 4 replicates) using Mouse Gene 2.0 ST Array platform (transcript version). Array data was processed by RMA algorithm.

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