Oxidative Status of DJ-1-dependent Activation of Dopamine Synthesis through Interaction of Tyrosine Hydroxylase and 4-Dihydroxy-l-phenylalanine (l-DOPA) Decarboxylase with DJ-1

Abstract

Parkinson disease (PD) is caused by loss of dopamine, which is synthesized from tyrosine by two enzymes, tyrosine hydroxylase (TH) and 4-dihydroxy-L-phenylalanine decarboxylase (DDC).DJ-1 is a causative gene for the familial form of PD, but little is known about the roles of DJ-1 in dopamine synthesis. In this study, we found that DJ-1 directly bound to TH and DDC and positively regulated their activities in human dopaminergic cells. Mutants of DJ-1 found in PD patients, including heterozygous mutants, lost their activity and worked as dominant-negative forms toward wild-type DJ-1. When cells were treated with H 2 O 2 , 6-hydroxydopamine, or 1-methyl-4-phenylpyridinium, changes in activities of TH and DDC accompanied by oxidation of cysteine 106 of DJ-1 occurred. It was found that DJ-1 possessing Cys-106 with SH and SOH forms was active and that DJ-1 possessing Cys-106 with SO 2 H and SO 3 H forms was inactive in terms of stimulation of TH and DDC activities. These findings indicate an essential role of DJ-1 in dopamine synthesis and contribution of DJ-1 to the sporadic form of PD. Parkinson disease (PD)2 is a neurodegenerative disease that occurs by reduction of the dopamine level through dopaminergic cell death in the substantia nigra. Genetic and environmental factors are thought to be triggers for the onset of PD, but the precise molecular mechanisms are still not known. Dopamine is synthesized by the following two steps: tyrosine is converted to L-DOPA by tyrosine hydroxylase (TH) and then L-DOPA is converted to dopamine by L-dopa decarboxylase (DDC). TH is therefore a key enzyme for dopamine synthesis and is used as a marker for dopaminergic neurons.DJ-1 was first identified by our group as a novel candidate of the oncogene that transformed mouse NIH3T3 cells in cooperation with activated ras (1). Deletion and point (L166P) mutations of DJ-1 have been shown to be responsible for onset of familial Parkinson disease, PARK7 (2), and other homozygous and heterozygous mutations of DJ-1 have been identified in patients with familial or sporadic Parkinson disease (3-6). DJ-1 is a multifunctional protein and plays roles in transcriptional regulation and anti-oxidative stress function, and loss of its functions is thought to lead to the onset of Parkinson disease and cancer. DJ-1 has three cysteine residues at positions 46, 53, and 106 (Cys-46, Cys-53, and Cys-106, respectively), and these cysteine residues are oxidized after cells receive oxidative stress, resulting in scavenging of reactive oxidative species (7-12). Although DJ-1 does not directly bind to DNA, DI-1 acts as a co-activator to activate various transcription factors, including the androgen receptor and p53 tumor suppressor, PSF and Nrf2, by sequestering their inhibitory factors (13-18). The anti-oxidative stress function of DJ-1 is therefore thought to be carried out both by self-oxidation of cysteine residues and by activation of redox-related genes.It has been reported that PSF, a transcription repressor, binds to the promoter region of the TH gene...

Stimulation of vesicular monoamine transporter 2 activity by DJ-1 in SH-SY5Y cells

“…Parkinson's disease, regulates TH and DDC activities: DJ-1 upregulates transcription of the TH gene in a human-specific manner [15] and enhances TH and DDC activities through protein-protein interaction [24]. As in the case of TH, the present study showed that DJ-1 activates VMAT2 expression and enhances its activity at the level of transcription and by protein-protein interaction.…”

“…We have reported that DJ-1 positively regulates TH mRNA expression and TH activity by sequestering PSF, a repressor for transcription of the TH gene, and by direct association of DJ-1 with TH, respectively [15,24]. It has been reported that TH and DDC are associated with VMAT2 [3].…”

“…Human SH-SY5Y and SH-SY5Y cells expressing wild-type or mutant forms of DJ-1 [24] were cultured in Dulbecco's modified Eagle's medium (DMEM) with 10% calf serum. The nucleotide sequences for siRNA targeting human DJ-1 were as follows:…”

“…Stable cell lines of SH-SY5Y cells expressing wild-type DJ-1 and mutant DJ-1, including M26I, C106S and L166P DJ-1, were established as described previously [24]. SH-SY5Y cells harboring a vector backbone were also established.…”

“…DJ-1 is a multi-functional protein and plays roles in transcriptional regulation [8][9][10][11][12][13][14][15] and in modulation of signaling cascades [16][17][18][19][20] through protein-protein interaction, leading to anti-oxidative stress function [21][22][23]. Furthermore, we have reported that DJ-1 activated TH and DDC through direct binding to TH and DDC in an oxidative status of DJ-1-dependent manner [24] and that human DJ-1 activates TH gene expression in cultured human dopaminergic cells [15]. DJ-1 is preferentially localized in the cytoplasm and nucleus [7] and a portion of DJ-1 is localized in the mitochondria [22,[25][26][27] and synapse [28].…”