Christian Pifl

Neurotransmitter transporters determine the intensity and duration of signal transduction by controlling the rapid removal of transmitter molecules from the synaptic cleft. The importance of their function is further reflected by the medical and social implications of compounds that inhibit their activity such as the antidepressants and cocaine. Molecular characterization of these transporters has revealed that they are members of a large family of membrane proteins with 12 putative transmembrane domains. However, little information exists as to whether discrete domains of these proteins mediate the various defined functions of these transporters. In this study, we constructed a series of chimeras between two structurally related but pharmacologically distinct transporters, the dopamine and norepinephrine transporters. The properties of these chimeric transporters suggest that distinct regions of these molecules determine these individual functions. Regions from the amino-terminal through the first five transmembrane domains are likely to be involved in the uptake mechanisms and ionic dependence. Regions within transmembrane domains 6-8 determine tricyclic antidepressant binding and cocaine interactions, whereas the carboxyl-terminal region encompassing transmembrane domain 9 through the COOH-terminal tail appears to be responsible for the stereoselectivity and high affinity for substrates. The dissociation of the substrate uptake and cocaine binding properties of these transporters further raises the possibility that antagonists of cocaine action devoid of uptake blockade activity might be developed for the clinical management of cocaine addiction.

Mutations in the DJ-1 gene cause early-onset autosomal recessive Parkinson's disease (PD), although the role of DJ-1 in the degeneration of dopaminergic neurons is unresolved. Here we show that the major interacting-proteins with DJ-1 in dopaminergic neuronal cells are the nuclear proteins p54nrb and pyrimidine tract-binding protein-associated splicing factor (PSF), two multifunctional regulators of transcription and RNA metabolism. PD-associated DJ-1 mutants exhibit decreased nuclear distribution and increased mitochondrial localization, resulting in diminished co-localization with co-activator p54nrb and repressor PSF. Unlike pathogenic DJ-1 mutants, wild-type DJ-1 acts to inhibit the transcriptional silencing activity of the PSF. In addition, the transcriptional silencer PSF induces neuronal apoptosis, which can be reversed by wild-type DJ-1 but to a lesser extent by PD-associated DJ-1 mutants. DJ-1-specific small interfering RNA sensitizes cells to PSF-induced apoptosis. Both DJ-1 and p54nrb block oxidative stress and mutant α-synuclein-induced cell death. Thus, DJ-1 is a neuroprotective transcriptional co-activator that may act in concert with p54nrb and PSF to regulate the expression of a neuroprotective genetic program. Mutations that impair the transcriptional co-activator function of DJ-1 render dopaminergic neurons vulnerable to apoptosis and may contribute to the pathogenesis of PD.