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Genomic sequencing efforts can implicate large numbers of genes and de novo mutations as potential disease risk factors. A high throughput in vivo model system to validate candidate gene association with pathology is therefore useful. We present such a system employing Drosophila to validate candidate congenital heart disease (CHD) genes. The protocols exploit comprehensive libraries of UAS-GeneX-RNAi fly strains that when crossed into a 4×Hand-Gal4 genetic background afford highly efficient cardiac-specific knockdown of endogenous fly orthologs of human genes. A panel of quantitative assays evaluates phenotypic severity across multiple cardiac parameters. These include developmental lethality, larva and adult heart morphology, and adult longevity. These protocols were recently used to evaluate more than 100 candidate CHD genes implicated by patient whole-exome sequencing (Zhu et al., 2017).
The use of the Drosophila model to elucidate molecular mechanisms underlying human diseases is well documented (Bier and Bodmer, 2004; Cagan, 2011; Zhang et al., 2013; Owusu-Ansah and Perrimon, 2014; Diop and Bodmer, 2015; Na et al., 2015), and 75% of human disease associated genes are represented by functional homologs in the fly genome (Reiter et al., 2001). While it is a challenge to link Drosophila developmental phenotypes directly to patient symptoms, Drosophila can be used as a very sophisticated and efficient platform to test and validate candidate disease gene function in development, and this can readily be scaled to evaluate large number of candidate genes identified from patient genomic sequencing efforts. Drosophila has been used to study genes related to CHD for over 20 years, based on evolutionarily conserved genetic mechanisms of heart development (Bier and Bodmer, 2004; Olson, 2006; Yi et al., 2006). We developed a highly efficient cardiac-targeted gene silencing approach in flies to examine effects on heart structure and function for fly homologs of candidate CHD genes (Zhu et al., 2017).
Z.H. was supported by grants from the National Institutes of Health (RO1-HL090801, RO1-NK098410).