Computational Approaches for Mining GRO-Seq Data to Identify and Characterize Active Enhancers.
Transcriptional enhancers are DNA regulatory elements that are bound by transcription factors and act to positively regulate the expression of nearby or distally located target genes. Enhancers have many features that have been discovered using genomic analyses. Recent studies have shown that active enhancers recruit RNA polymerase II (Pol II) and are transcribed, producing enhancer RNAs (eRNAs). GRO-seq, a method for identifying the location and orientation of all actively transcribing RNA polymerases across the genome, is a powerful approach for monitoring nascent enhancer transcription. Furthermore, the unique pattern of enhancer transcription can be used to identify enhancers in the absence of any information about the underlying transcription factors. Here, we describe the computational approaches required to identify and analyze active enhancers using GRO-seq data, including data pre-processing, alignment, and transcript calling. In addition, we describe protocols and computational pipelines for mining GRO-seq data to identify active enhancers, as well as known transcription factor binding sites that are transcribed. Furthermore, we discuss approaches for integrating GRO-seq-based enhancer data with other genomic data, including target gene expression and function. Finally, we describe molecular biology assays that can be used to confirm and explore further the function of enhancers that have been identified using genomic assays. Together, these approaches should allow the user to identify and explore the features and biological functions of new cell type-specific enhancers.
The Laboratory of Signaling and Gene Expression, Cecil H. and Ida Green Center for Reproductive Biology Sciences, University of Texas Southwestern Medical Center, 5323 Harry Hines Boulevard, Dallas, TX, 75390-8511, USA.|The Division of Basic Research, Department of Obstetrics and Gynecology, University of Texas Southwestern Medical Center, Dallas, TX, 75390-8511, USA.
Anusha
A
Nagari
The Laboratory of Signaling and Gene Expression, Cecil H. and Ida Green Center for Reproductive Biology Sciences, University of Texas Southwestern Medical Center, 5323 Harry Hines Boulevard, Dallas, TX, 75390-8511, USA.|The Division of Basic Research, Department of Obstetrics and Gynecology, University of Texas Southwestern Medical Center, Dallas, TX, 75390-8511, USA.|Program in Genetics, Development and Disease, Graduate School of Biomedical Sciences, University of Texas Southwestern Medical Center, Dallas, TX, 75390, USA.
Shino
S
Murakami
The Laboratory of Signaling and Gene Expression, Cecil H. and Ida Green Center for Reproductive Biology Sciences, University of Texas Southwestern Medical Center, 5323 Harry Hines Boulevard, Dallas, TX, 75390-8511, USA.|The Division of Basic Research, Department of Obstetrics and Gynecology, University of Texas Southwestern Medical Center, Dallas, TX, 75390-8511, USA.
Venkat S
VS
Malladi
The Laboratory of Signaling and Gene Expression, Cecil H. and Ida Green Center for Reproductive Biology Sciences, University of Texas Southwestern Medical Center, 5323 Harry Hines Boulevard, Dallas, TX, 75390-8511, USA. LEE.KRAUS@utsouthwestern.edu.|The Division of Basic Research, Department of Obstetrics and Gynecology, University of Texas Southwestern Medical Center, Dallas, TX, 75390-8511, USA. LEE.KRAUS@utsouthwestern.edu.|Program in Genetics, Development and Disease, Graduate School of Biomedical Sciences, University of Texas Southwestern Medical Center, Dallas, TX, 75390, USA. LEE.KRAUS@utsouthwestern.edu.
W Lee
WL
Kraus
D019295:Computational Biology / Q000379:methods*; D057225:Data Mining / Q000379:methods*; D004742:Enhancer Elements, Genetic; D006801:Humans; D012319:RNA Polymerase II / Q000378:metabolism; D062085:RNA, Long Noncoding / Q000235:genetics; D016415:Sequence Alignment; D017423:Sequence Analysis, RNA; D014158:Transcription, Genetic
D062085:RNA, Long Noncoding; D012319:RNA Polymerase II
Enhancer; Enhancer RNAs (eRNAs); Enhancer prediction; GRO-seq; Gene regulation; Looping; Motif; Motif search; Promoter; Response element; Transcription; Transcription factor; Transcription unit; groHMM
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