A Novel Transgenic Zebrafish Line Reporting Dynamic Epigenetic Changes

Abstract:

Currently, there is no other whole-animal reporter for epigenetic regulation established in any vertebrate. 

The inventors generated this novel zebrafish line using a transgene construct containing dazl gene silencing sequences (CpG island) fused to a destabilized GFPd2 gene driven by the ubiquitously expressing ef1alpha promoter. The resulting transgenic line permits detailed tissue- or cellular- level visualization of dynamic changes in GFPd2 expression in response to changes in DNA methylation or downstream epigenetic regulation in developing or adult animals. GFPd2 is “off” in the fertilized egg but turns on during early development, peaking at approximately 24 hours post-fertilization, and is then rapidly and ubiquitously silenced. The reporter is “off’ in adults, except in particular stages of germline development in the gonads.  Experimental treatments or genetic mutants that interfere with epigenetic silencing result in global or tissue-specific “reactivation” of GFPd2 expression. The reporter is also reactivated in regenerating tissues.

The “EpiTag” zebrafish transgenic line accurately reports changes in epigenetic silencing due to altered epigenetic regulation. The line can provide a whole-animal platform for identifying and studying tissue- or even cellular- level function of genes involved in mediating epigenetic regulation, for carrying out screens for potential therapeutics targeting epigenetic regulatory mechanisms, or for studying the epigenetic effects of environmental toxins. This new model represents the first vertebrate reporter for studying epigenetic regulation in an intact, living animal, and potentially has wide applications in academic research as well as in drug development. 

Competitive Advantages:

• This new model represents the first vertebrate reporter for studying epigenetic regulation in an intact, living animal, and potentially has wide applications in academic research as well as in drug development
• This line incorporates the benefits of using zebrafish as a model organism in research or drug development, including high-throughput chemical or genetic screening with an intact animal model

Commercial Applications:

• Use of the transgenic zebrafish line:
  • Examine tissue-specific gene silencing during normal or abnormal development 
  • Use of ENU (N-ethyl-N-nitrosourea) mutagenesis screens to identify global- or tissue-specific epigenetic regulators 
  • Study the tissue- and cellular-level functions of identified epigenetic regulatory genes and proteins
  • Screen for therapeutic compounds that potentially modulate epigenetic regulation in certain clinical applications
  • Screen for environmental toxins that potentially affect epigenetic regulation
  • Visualize and study regenerating tissues and changes in epigenetic regulation that accompany regeneration