1. Chromosomes and Gene Expression
  2. Genetics and Genomics
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Regulatory Networks: Reality check for transposon enhancers

  1. Julie Brind'Amour
  2. Dixie L Mager  Is a corresponding author
  1. University of British Columbia, Canada
  2. BC Cancer, Canada
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Cite this article as: eLife 2019;8:e47900 doi: 10.7554/eLife.47900
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Figures

Functional validation of putative TE+ enhancers.

Here, the long terminal repeats belonging to the RLTR13D6 family are used to demonstrate how putative enhancers can be validated. (A) Of the 805 copies of RLTR13D6 sequences (blue boxes) in the mouse genome, 76 have an ‘enhancer-like’ chromatin state in embryonic stem cells and bind at least one key transcription factor in this cell type (green stars). (B) High-throughput plasmid-based reporter assays work by inserting a potential enhancer sequence into a plasmid (black circle), and examining its impact on the expression of a reporter gene (dark purple box). Only a third of the long terminal repeats that show enhancer activity in these assays have an enhancer-like chromatin state in the genome (dark green fraction of the pie chart). (C) Promoter capture Hi-C experiments showed that about a third of the putative enhancers (dark green fraction of the pie chart) interacted with the promoter of at least one gene (purple boxes). (D) Disrupting RLTR13D6 long terminal repeats using CRISPR interference reduced the histone mark H3K27ac (a sign of enhancer function) by at least two fold for 34 of the 76 sequences (light green). Using RNA-seq after CRISPR interference showed that only three genes associated with an RLTR13D6 element were down-regulated by at least 1.5 fold. (E) In embryonic stem cells, CRISPR-Cas9 deletion (blue box disappearing) of four long terminal repeats with enhancer signatures reduced gene expression in only one case.

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