Drosophila Ctr1A Functions as a Copper Transporter Essential for Development

Michelle L Turski,Dennis J Thiele

doi:10.1074/jbc.m703792200

Abstract

Copper is an essential trace element required by all aerobic organisms as a cofactor for enzymes involved in normal growth, development, and physiology. Ctr1 proteins are members of a highly conserved family of copper importers responsible for copper uptake across the plasma membrane. Mice lacking Ctr1 die during embryogenesis from widespread developmental defects, demonstrating the need for adequate copper acquisition in the development of metazoan organisms via as yet uncharacterized mechanisms. Whereas the fruit fly, Drosophila melanogaster, expresses three Ctr1 genes, ctr1A, ctr1B, and ctr1C, little is known about their protein isoform-specific roles. Previous studies demonstrated that Ctr1B localizes to the plasma membrane and is not essential for development unless flies are severely copper-deficient or are subjected to copper toxicity. Here we demonstrate that Ctr1A also resides on the plasma membrane and is the primary Drosophila copper transporter. Loss of Ctr1A results in copper-remedial developmental arrest at early larval stages. Ctr1A mutants are deficient in the activity of copper-dependent enzymes, including cytochrome c oxidase and tyrosinase. Amidation of Phe-Met-Arg-Phe-amides, a group of cardiomodulatory neuropeptide hormones that are matured via the action of peptidylglycine alpha-hydroxylating monooxygenase, is defective in neuroendocrine cells of Ctr1A mutant larvae. Moreover, both the Phe-Met-Arg-Phe-amide maturation and heart beat rate defects observed in Ctr1A mutant larvae can be partially rescued by exogenous copper. These studies establish clear physiological distinctions between two Drosophila plasma membrane copper transport proteins and demonstrate that copper import by Ctr1A is required to drive neuropeptide maturation during normal growth and development.

Highlights

Melanin production [1,2,3]
Ctr1A Localizes to the Plasma Membrane and to Intracellular Vesicles—Earlier work established that the Drosophila Ctr1B copper transporter is inducibly expressed in response to copper deficiency, localizes to the plasma membrane, and has a physiologically important role in copper accumulation and in development under conditions of either copper limitation or copper excess. mRNA blotting experiments demonstrated that in contrast to Ctr1B, Ctr1A mRNA is expressed during all major stages of development and in adult flies [29]
Data reported here strongly suggest that Ctr1A is the primary copper transporter in Drosophila given its ubiquitous expression pattern throughout development and into adulthood, the fact that it shares the highest degree of homology to mammalian Ctr1 proteins, and its lethal loss of function phenotype

Summary

Introduction

Melanin production [1,2,3]. In addition to the canonical role of copper as a cofactor, data support novel roles for copper in the secretion of growth factors and in modulating neuronal function (4 – 6). The modest reduction in copper levels in both the heterozygous and hemizygous ctr1A25 mutant larvae can potentially be explained by the copper transporting activity of Ctr1B, which is expressed in 1st instar larvae, the stage at which metal content analysis was performed.

Results

Conclusion