A Drosophila model of Menkes disease reveals a role for DmATP7 in copper absorption and neurodevelopment

The trace metal copper is an essential nutrient that is absorbed mainly by the small intestine into the bloodstream and other tissues, where it acts as a cofactor for a number of key metabolic enzymes (Nelson, 1999; Phillips and Hilliker, 1990). Although essential for life, copper can be very toxic at high concentrations and can promote the production of highly reactive hydroxyl radicals through the Haber-Weiss/Fenton reaction (Phillips and Hilliker, 1990). Therefore, heavy metal homeostasis needs to be regulated tightly to ensure an adequate supply of metal ions for normal cellular metabolism while protecting cells from their toxic effects (Williams et al., 2000). In humans, the primary copper exporters are the ATP7B and ATP7A copper-transporting ATPases. In Wilson’s disease, mutations in the ATP7B gene are associated with copper accumulation in the liver and the brain, with subsequent development of progressive hepatic and neurological abnormalities (Lutsenko et al., 2007; Pfeiffer, 2007). Menkes disease and occipital horn syndrome (OHS) are allelic X-linked recessive disorders of copper deficiency that are caused by mutations in the ATP7A copper-transporting gene (Cox and Moore, 2002; de Bie et al., 2007; Kaler et al., 1994; Levinson et al., 1996). The ATP7A coppertransporting protein is located in the trans-Golgi network, where it delivers copper to the cellular secretory compartments for incorporation into various copper-dependent enzymes; in the presence of excess copper, ATP7A traffics to the plasma membrane and regulates copper homeostasis by exporting excess copper across the cellular membrane (Harris, 2000; Lutsenko et al., 2007; Kim and Petris, 2007; Voskoboinik and Camakaris, 2002). These properties of ATP7A are required for copper efflux from intestinal cells and, as a consequence, mutations in the ATP7A gene can disrupt copper release from the intestinal tract into the blood and restrict its supply to other tissues, particularly the brain (Harris, 2000; Kodama et al., 1999; Lutsenko et al., 2007). Thus, Wilson’s and Menkes disorders are opposite pathologies, in that the former disease is associated with copper accumulation, whereas the latter is associated with profound copper deficiency. Menkes disease patients display severe developmental, neurological and connective tissue abnormalities along with various other symptoms that are associated with a decreased function of copper-dependent enzymes (Lutsenko et al., 2007; Tomita et al., 1992). Patients with OHS, a milder form of Menkes disease, also exhibit connective tissue impairments but are spared of severe neurological abnormalities, presumably owing to residual expression of functional ATP7A (Das et al., 1995; Mercer, 1998; Møller et al., 2000; Tang et al., 2006; Tsukahara et al., 1994). The mottled mutant mice are a series of copper-deficient mice that possess mutations in the murine homologue of the Menkes disease gene; these mutants display a diversity of phenotypic severities ranging from prenatal lethality to an adult viable phenotype (Kim and Petris, 2007; La Fontaine et al., 1999; Llanos et al., 2006). The mottled-brindled (Mobr) mutant mouse is the closest animal model to human Menkes disease and has been used widely for copper therapy and gene interaction studies (Grimes et al., 1997; Kelly and Palmiter, 1996). Similar to Menkes patients, the Mobr copperdeficient mice exhibit profound neurological abnormalities and die early after birth. However, blotchy (Moblo) is a less severe allele of mottled that is characterized mainly by connective tissue defects, akin to human OHS (Mercer, 1998). During the last decade, studies on mottled mutant mice have greatly advanced our understanding of the mechanisms of Menkes disease and OHS pathogenesis. Studies in mice, however, can be costly and time consuming. In the present study, we have constructed a Drosophila model of Menkes disease by conditional silencing of DmATP7 (Southon et al., 2004), the sole Drosophila orthologue of the mammalian ATP7A and ATP7B genes, in the gut. An earlier study has shown that DmATP7 is required for normal development and adult pigmentation, presumably by facilitating RESEARCH ARTICLE