Abstract
Dystonia represents the third most common movement disorder in humans. At least 15 genetic loci (DYT1-15) have been identified and some of these genes have been cloned. TOR1A (formally DYT1), the gene responsible for the most common primary hereditary dystonia, encodes torsinA, an AAA ATPase family protein. However, the function of torsinA has yet to be fully understood. Here, we have generated and characterized a complete loss-of-function mutant for dtorsin, the only Drosophila ortholog of TOR1A. Null mutation of the X-linked dtorsin was semi-lethal with most male flies dying by the pre-pupal stage and the few surviving adults being sterile and slow moving, with reduced cuticle pigmentation and thin, short bristles. Third instar male larvae exhibited locomotion defects that were rescued by feeding dopamine. Moreover, biochemical analysis revealed that the brains of third instar larvae and adults heterozygous for the loss-of-function dtorsin mutation had significantly reduced dopamine levels. The dtorsin mutant showed a very strong genetic interaction with Pu (Punch: GTP cyclohydrolase), the ortholog of the human gene underlying DYT14 dystonia. Biochemical analyses revealed a severe reduction of GTP cyclohydrolase protein and activity, suggesting that dtorsin plays a novel role in dopamine metabolism as a positive-regulator of GTP cyclohydrolase protein. This dtorsin mutant line will be valuable for understanding this relationship and potentially other novel torsin functions that could play a role in human dystonia.
Highlights
Dystonia is a movement disorder characterized by sustained muscle contraction
Our findings indicate that disrupted dopamine metabolism is at least one consequence of this null mutation and that dtorsin is a novel positive-regulator of GTP cyclohydrolase (GTPCH) protein in Drosophila
PCR analysis with dtorsin gene-specific primers demonstrated that males from each of the seven semi-lethal lines lacked the dtorsin gene, while males of the seven viable lines retained the dtorsin gene
Summary
Dystonia is a movement disorder characterized by sustained muscle contraction. While 15 different gene loci have been implicated in primary hereditary dystonia (DYT1-DYT15) [1], the most common and severe form, early-onset dystonia ( known as DYT1 dystonia) is due to mutation in TOR1A (formerly DYT1) and displays dominant inheritance with reduced penetrance [2]. TOR1A encodes torsinA, a 332 amino acid protein from the AAA ATPase family. Most cases of DYT1 dystonia are caused by a single mutation, a 3 bp (GAG) deletion that results in the loss of a glutamic acid residue in the carboxyl terminal region of torsinA [3]. The dtorsinencoded protein, Dtorsin, comprises 339 amino acids with 31.9% identity to human torsinA and displays the characteristic features of the AAA ATPase gene family [5]. Misexpression in Drosophila of the mutant form of human torsinA, pan-neuronally or in monoaminergic cells results in a locomotor defect [6] and aberrant bouton morphologies at synapses [7], though the function of Dtorsin is unclear. Muraro and Moffat [8] have reported that targeted down-regulation of the dtorsin gene in the Drosophila eye caused progressive degeneration of the retina, but no loss-offunction mutants of dtorsin have previously been characterized
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