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Robust partitioning of microRNA targets from downstream regulatory changes [Ribo-seq]

ABSTRACT: Robust partitioning of microRNA targets from downstream regulatory changes [Ribo-seq]

PROVIDER: PRJNA589466 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR10441364.fastq.gz	Fastqsanger.gz
	SRR10441365.fastq.gz	Fastqsanger.gz
	SRR10441366.fastq.gz	Fastqsanger.gz
	SRR10441367.fastq.gz	Fastqsanger.gz
	SRR10441368.fastq.gz	Fastqsanger.gz

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Robust partitioning of microRNA targets from downstream regulatory changes

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Robust partitioning of microRNA targets from downstream regulatory changes [Ribo-seq]

Project description:MicroRNA’s function is defined by the targets it regulates. Despite miRNAs’ crucial role in various cellular processes and disease states, the identification of their targets remains challenging. Existing methods suffer from high false-positive rates and are inadequate in differentiating direct targets from downstream regulatory changes. Here, we introduce a simple approach involving combined analysis of RNA-seq and Precise RunOn-seq (PRO-seq) (CARP) to effectively measure post-transcriptional regulation. We apply this approach to seven different miRNAs and robustly distinguish their direct targets from downstream changes. We validate the efficacy of our approach using argonaute-CLIP-seq. We show using CARP and ribosome-profiling analyses that accelerated mRNA decay is the major mode of target regulation by miRNAs. Additionally, our comparative analyses reveal that CARP aids in discovery of complex regulatory mechanisms and in capturing false-negatives of existing approaches. Furthermore, post-transcriptional changes measured using CARP uncover miRNA-dependent regulation of coding sites. Lastly, we demonstrate that PRO-seq provides mechanistic explanation of indirect targeting, unraveling miRNA regulatory networks. Finally, we believe that CARP is particularly suitable to study in vivo functions of miRNAs in primary cells.

2020-08-07 | GSE140366 | GEO

Robust partitioning of microRNA targets from downstream regulatory changes.

Project description:The biological impact of microRNAs (miRNAs) is determined by their targets, and robustly identifying direct miRNA targets remains challenging. Existing methods suffer from high false-positive rates and are unable to effectively differentiate direct miRNA targets from downstream regulatory changes. Here, we present an experimental and computational framework to deconvolute post-transcriptional and transcriptional changes using a combination of RNA-seq and PRO-seq. This novel approach allows us to systematically profile the regulatory impact of a miRNA. We refer to this approach as CARP: Combined Analysis of RNA-seq and PRO-seq. We apply CARP to multiple miRNAs and show that it robustly distinguishes direct targets from downstream changes, while greatly reducing false positives. We validate our approach using Argonaute eCLIP-seq and ribosome profiling, demonstrating that CARP defines a comprehensive repertoire of targets. Using this approach, we identify miRNA-specific activity of target sites within the open reading frame. Additionally, we show that CARP facilitates the dissection of complex changes in gene regulatory networks triggered by miRNAs and identification of transcription factors that mediate downstream regulatory changes. Given the robustness of the approach, CARP would be particularly suitable for dissecting miRNA regulatory networks in vivo.

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Robust partitioning of microRNA targets from downstream regulatory changes [miRNA-seq]

Project description:The biological impact of microRNAs is determined by their targets, and robustly identifying direct miRNA targets remains challenging. Existing methods suffer from high false-positive rates and are unable to effectively differentiate direct miRNA targets from downstream regulatory changes. Here, we present a simple approach to deconvolute post-transcriptional and transcriptional changes using PRO-seq with RNA-seq. In combination, these methods allow us to systematically profile the regulatory impact of a miRNA. We refer to this approach as CARP: Combined Analysis of RNA-seq and PRO-seq. We apply CARP to multiple miRNAs and show that it robustly distinguishes direct targets from downstream changes, while greatly reducing false positives. We validate our approach using Argonaute eCLIP-seq and ribosome profiling, demonstrating that CARP defines a comprehensive repertoire of targets. We identify miRNA-specific activity of target sites within the coding region. CARP facilitates the dissection of complex changes in gene regulatory networks triggered by miRNAs and identification of transcription factors that underlie downstream regulatory changes. Given the robustness of the approach, CARP is particularly suitable for dissecting miRNA regulatory networks in vivo.

2020-08-07 | GSE142898 | GEO