Multiplexed massively parallel SELEX for characterization of human transcription factor binding specificities

Research output: Journal Publications and ReviewsRGC 21 - Publication in refereed journalpeer-review

330 Scopus Citations
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Author(s)

  • Arttu Jolma
  • Teemu Kivioja
  • Jarkko Toivonen
  • Lu Cheng
  • Gonghong Wei
  • Martin Enge
  • Mikko Taipale
  • Juan M. Vaquerizas
  • Mikko J. Sillanpää
  • Martin Bonke
  • Kimmo Palin
  • Shaheynoor Talukder
  • Timothy R. Hughes
  • Nicholas M. Luscombe
  • Esko Ukkonen
  • Jussi Taipale

Detail(s)

Original languageEnglish
Pages (from-to)861-873
Journal / PublicationGenome Research
Volume20
Issue number6
Online published8 Apr 2010
Publication statusPublished - Jun 2010
Externally publishedYes

Abstract

The genetic code-the binding specificity of all transfer-RNAs-defines how protein primary structure is determined by DNA sequence. DNA also dictates when and where proteins are expressed, and this information is encoded in a pattern of specific sequence motifs that are recognized by transcription factors. However, the DNA-binding specificity is only known for a small fraction of the ∼1400 human transcription factors (TFs). We describe here a high-throughput method for analyzing transcription factor binding specificity that is based on systematic evolution of ligands by exponential enrichment (SELEX) and massively parallel sequencing. The method is optimized for analysis of large numbers of TFs in parallel through the use of affinity-tagged proteins, barcoded selection oligonucleotides, and multiplexed sequencing. Data are analyzed by a new bioinformatic platform that uses the hundreds of thousands of sequencing reads obtained to control the quality of the experiments and to generate binding motifs for the TFs. The described technology allows higher throughput and identification of much longer binding profiles than current microarray-based methods. In addition, as our method is based on proteins expressed in mammalian cells, it can also be used to characterize DNA-binding preferences of full-length proteins or proteins requiring post-translational modifications. We validate the method by determining binding specificities of 14 different classes of TFs and by confirming the specificities for NFATC1 and RFX3 using ChIP-seq. Our results reveal unexpected dimeric modes of binding for several factors that were thought to preferentially bind DNA as monomers.

Citation Format(s)

Multiplexed massively parallel SELEX for characterization of human transcription factor binding specificities. / Jolma, Arttu; Kivioja, Teemu; Toivonen, Jarkko et al.
In: Genome Research, Vol. 20, No. 6, 06.2010, p. 861-873.

Research output: Journal Publications and ReviewsRGC 21 - Publication in refereed journalpeer-review