Systematic dissection and optimization of inducible enhancers in human cells using a massively parallel reporter assay

Alexandre Melnikov, Anand Murugan, Xiaolan Zhang, Tiberiu Tesileanu, Li Wang, Peter Rogov, Soheil Feizi, Andreas Gnirke, Curtis Gove Callan, Justin B. Kinney, Manolis Kellis, Eric S. Lander, Tarjei S. Mikkelsen

Research output: Contribution to journalArticlepeer-review

310 Scopus citations

Abstract

Learning to read and write the transcriptional regulatory code is of central importance to progress in genetic analysis and engineering. Here we describe a massively parallel reporter assay (MPRA) that facilitates the systematic dissection of transcriptional regulatory elements. In MPRA, microarray-synthesized DNA regulatory elements and unique sequence tags are cloned into plasmids to generate a library of reporter constructs. These constructs are transfected into cells and tag expression is assayed by high-throughput sequencing. We apply MPRA to compare >27,000 variants of two inducible enhancers in human cells: a synthetic cAMP-regulated enhancer and the virus-inducible interferon-β enhancer. We first show that the resulting data define accurate maps of functional transcription factor binding sites in both enhancers at single-nucleotide resolution. We then use the data to train quantitative sequence-activity models (QSAMs) of the two enhancers. We show that QSAMs from two cellular states can be combined to design enhancer variants that optimize potentially conflicting objectives, such as maximizing induced activity while minimizing basal activity.

Original languageEnglish (US)
Pages (from-to)271-277
Number of pages7
JournalNature biotechnology
Volume30
Issue number3
DOIs
StatePublished - Mar 2012

All Science Journal Classification (ASJC) codes

  • Biotechnology
  • Bioengineering
  • Applied Microbiology and Biotechnology
  • Molecular Medicine
  • Biomedical Engineering

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