Rapid Tumor Induction in Zebrafish by TALEN-Mediated Somatic Inactivation of the Retinoblastoma1 Tumor Suppressor rb1

Abstract

<p>Investigating the <em>in vivo</em> role of tumor suppressor genes in cancer is technically challenging due to their essential requirement during early animal development. To address this bottleneck, we generated genetic mosaic adult zebrafish using TALEN genome editing and demonstrate somatic inactivation of the tumor suppressor <em>retinoblastoma1</em> (<em>rb1</em>) induces tumorigenesis at high frequency. 11–33% of 1-cell stage embryos injected with TALEN mRNAs targeting <em>rb1</em> exon 2 or 3 develop tumors beginning as early as 3.5 months of age. Lesions predominantly arise in the brain and show features of neuroectodermal-like and glial-like tumors. Mutant allele analysis is consistent with tumor initiation due to somatic inactivation of <em>rb1</em>, revealing a conserved role for <em>rb1</em> in tumor suppression across vertebrates. In contrast to genetic mosaics, heterozygous <em>rb1−</em>/+ adults show no evidence of neoplasia, while homozygous mutant <em>rb1−/−</em> are larval lethal. This is the first demonstration that somatic inactivation of a tumor suppressor causes cancer in zebrafish, and highlights the utility of site-specific nucleases to create genetic mosaic zebrafish for tumor suppressor gene discovery. Somatic inactivation with site-directed nucleases in zebrafish presents a rapid and scalable strategy to study tumor suppressor gene function in cancer.</p>