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In Vivo Enhancer Analysis Chromosome 16 Conserved Noncoding Sequences

  • Author(s): Pennacchio, Len A.
  • Ahituv, Nadav
  • Moses, Alan M.
  • Nobrega, Marcelo
  • Prabhakar, Shyam
  • Shoukry, Malak
  • Minovitsky, Simon
  • Visel, Axel
  • Dubchak, Inna
  • Holt, Amy
  • Lewis, Keith D.
  • Plajzer-Frick, Ingrid
  • Akiyama, Jennifer
  • De Val, Sarah
  • Afzal, Veena
  • Black, Brian L.
  • Couronne, Olivier
  • Eisen, Michael B.
  • Rubin, Edward M.
  • et al.
Abstract

The identification of enhancers with predicted specificities in vertebrate genomes remains a significant challenge that is hampered by a lack of experimentally validated training sets. In this study, we leveraged extreme evolutionary sequence conservation as a filter to identify putative gene regulatory elements and characterized the in vivo enhancer activity of human-fish conserved and ultraconserved1 noncoding elements on human chromosome 16 as well as such elements from elsewhere in the genome. We initially tested 165 of these extremely conserved sequences in a transgenic mouse enhancer assay and observed that 48 percent (79/165) functioned reproducibly as tissue-specific enhancers of gene expression at embryonic day 11.5. While driving expression in a broad range of anatomical structures in the embryo, the majority of the 79 enhancers drove expression in various regions of the developing nervous system. Studying a set of DNA elements that specifically drove forebrain expression, we identified DNA signatures specifically enriched in these elements and used these parameters to rank all ~;3,400 human-fugu conserved noncoding elements in the human genome. The testing of the top predictions in transgenic mice resulted in a three-fold enrichment for sequences with forebrain enhancer activity. These data dramatically expand the catalogue of in vivo-characterized human gene enhancers and illustrate the future utility of such training sets for a variety of iological applications including decoding the regulatory vocabulary of the human genome.

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