Modeling Retinoblastoma in adult Drosophila by reducing Rbf gene expression

Abstract

Retinoblastoma is an eye cancer that begins in the retina due to genetic mutations in the retinal cells, typically of the Rb1 tumor suppressor gene, which encodes for the Rb protein. Rb inhibits the E2F transcription factor, which drives the expression of genes needed for entry into S-phase of the cell cycle. Loss of function of Rb dysregulates E2F, causing uninhibited cell-cycle proliferation and tumor formation. Current treatments for retinoblastoma are highly invasive such as surgery to remove the eye or chemotherapy impacting patients' lives. If an animal model of retinoblastoma is constructed, less invasive therapies can be tested. Drosophila melanogaster have a short life cycle, are cost-effective, and have an Rb1 homologue called Rbf1, making them an efficient model. An RNAi system was used to knock down the expression of Rbf1 protein via the gsGAL4-UAS gene switch system of gene regulation. A fly line in which the gsGAL4 transcription factor is expressed under the eye-specific elav promoter was crossed with a line in which an siRNA targeting Rbf1 mRNA is expressed under the control of an upstream activating sequence (UAS). When the offspring of this cross were fed RU486, the normally cytoplasmic gsGAL4 entered the nucleus of the retinal cells in which it was expressed and bound to the UAS, driving the expression of the siRNA, which was expected to knock down levels of Rbf1 protein in these flies. RU486-treated flies exhibited a change in eye morphology, suggestive of tumor formation, as well as differential Kaplan-Meier survival curves.