Dataset Information

FIREWACh: high-throughput functional detection of transcriptional regulatory modules in mammalian cells.

ABSTRACT: Promoters and enhancers establish precise gene transcription patterns. The development of functional approaches for their identification in mammalian cells has been complicated by the size of these genomes. Here we report a high-throughput functional assay for directly identifying active promoter and enhancer elements called FIREWACh (Functional Identification of Regulatory Elements Within Accessible Chromatin), which we used to simultaneously assess over 80,000 DNA fragments derived from nucleosome-free regions within the chromatin of embryonic stem cells (ESCs) and identify 6,364 active regulatory elements. Many of these represent newly discovered ESC-specific enhancers, showing enriched binding-site motifs for ESC-specific transcription factors including SOX2, POU5F1 (OCT4) and KLF4. The application of FIREWACh to additional cultured cell types will facilitate functional annotation of the genome and expand our view of transcriptional network dynamics.

SUBMITTER: Murtha M

PROVIDER: S-EPMC4020622 | biostudies-literature | 2014 May

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

FIREWACh: high-throughput functional detection of transcriptional regulatory modules in mammalian cells.

Murtha Matthew M Tokcaer-Keskin Zeynep Z Tang Zuojian Z Strino Francesco F Chen Xi X Wang Yatong Y Xi Xiangmei X Basilico Claudio C Brown Stuart S Bonneau Richard R Kluger Yuval Y Dailey Lisa L

Nature methods 20140323 5

Promoters and enhancers establish precise gene transcription patterns. The development of functional approaches for their identification in mammalian cells has been complicated by the size of these genomes. Here we report a high-throughput functional assay for directly identifying active promoter and enhancer elements called FIREWACh (Functional Identification of Regulatory Elements Within Accessible Chromatin), which we used to simultaneously assess over 80,000 DNA fragments derived from nucleo ...[more]

PMID: 24658142

Similar Datasets

Project description:Post-transcriptional gene regulation controls the amount of protein produced from an individual mRNA by altering rates of decay and translation. Many sequence elements that direct post-transcriptional regulation have been found; in mammals, most such elements are located within the 3' untranslated regions (3'UTRs). Comparative genomic studies demonstrate that mammalian 3'UTRs contain extensive conserved sequence tracts, yet only a small fraction corresponds to recognized elements, implying that many additional novel elements exist. Despite a variety of computational, molecular, and biochemical approaches, identifying functional 3'UTRs elements remains difficult.We created a high-throughput cell-based screen that enables identification of functional post-transcriptional 3'UTR regulatory elements. Our system exploits integrated single-copy reporters, which are expressed and processed as endogenous genes. We screened many thousands of short random sequences for their regulatory potential. Control sequences with known effects were captured effectively using our approach, establishing that our methodology was robust. We found hundreds of functional sequences, which we validated in traditional reporter assays, including verifying their regulatory impact in native sequence contexts. Although 3'UTRs are typically considered repressive, most of the functional elements were activating, including ones that were preferentially conserved. Additionally, we adapted our screening approach to examine the effect of elements on RNA abundance, revealing that most elements act by altering mRNA stability.We developed and used a high-throughput approach to discover hundreds of post-transcriptional cis-regulatory elements. These results imply that most human 3'UTRs contain many previously unrecognized cis-regulatory elements, many of which are activating, and that the post-transcriptional fate of an mRNA is largely due to the actions of many individual cis-regulatory elements within its 3'UTR.

Dataset Information

FIREWACh: high-throughput functional detection of transcriptional regulatory modules in mammalian cells.

Publications

FIREWACh: high-throughput functional detection of transcriptional regulatory modules in mammalian cells.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets