Gene-breaking Transposon Mutagenesis Reveals an Essential Role for Histone H2afza in Zebrafish Larval Development

Mech Dev. 2006 Jul;123(7):513-29. doi: 10.1016/j.mod.2006.06.002. Epub 2006 Jun 9.

Abstract

We report a novel gene tagging, identification and mutagenicity ('gene-breaking') method for the zebrafish, Danio rerio. This modular approach consists of two distinct and separable molecular cassettes. The first is a gene-finding cassette. In this study, we employed a 3' gene-tagging approach that selectively 'traps' transcripts regardless of expression status, and we show that this cassette identifies both known and novel endogenous transcripts in transgenic zebrafish. The second is a transcriptional termination mutagenicity cassette assembled from a combination of a splice acceptor and polyadenylation signal to disrupt tagged transcripts upon integration into intronic sequence. We identified both novel and conserved loci as linked phenotypic mutations using this gene-breaking strategy, generating molecularly null mutations in both larval lethal and adult viable loci. We show that the Histone 2a family member z (H2afza) variant is essential for larval development through the generation of a lethal locus with a truncation of conserved carboxy-terminal residues in the protein. In principle this gene-breaking strategy is scalable for functional genomics screens and can be used in Sleeping Beauty transposon and other gene delivery systems in the zebrafish.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Animals, Genetically Modified
  • DNA Transposable Elements / genetics*
  • Histones / genetics*
  • Histones / physiology
  • Larva / genetics
  • Larva / growth & development
  • Mutagenesis, Insertional*
  • Zebrafish / embryology
  • Zebrafish / genetics*

Substances

  • DNA Transposable Elements
  • Histones
  • histone H2A.F-Z