The origin of esterase activity of Parkinson's disease causative factor DJ-1 implied by evolutionary trace analysis of its prokaryotic homolog HchA.

DJ-1, a causative gene for hereditary recessive Parkinsonism, is evolutionarily conserved across eukaryotes and prokaryotes. Structural analyses of DJ-1 and its homologs suggested the 106th Cys is a nucleophilic cysteine functioning as the catalytic center of hydratase or hydrolase activity. Indeed, DJ-1 and its homologs can convert highly electrophilic α-oxoaldehydes such as methylglyoxal into α-hydroxy acids as hydratase in vitro, and oxidation-dependent ester hydrolase (esterase) activity has also been reported for DJ-1. The mechanism underlying such plural activities, however, has not been fully characterized. To address this knowledge gap, we conducted a series of biochemical assays assessing the enzymatic activity of DJ-1 and its homologs. We found no evidence for esterase activity in any of the Escherichia coli DJ-1 homologs. Furthermore, contrary to previous reports, we found that oxidation inactivated rather than facilitated DJ-1 esterase activity. The E. coli DJ-1 homolog HchA possesses phenylglyoxalase and methylglyoxalase activities but lacks esterase activity. Since evolutionary trace analysis identified the 186th H as a candidate residue involved in functional differentiation between HchA and DJ-1, we focused on H186 of HchA and found that an esterase activity was acquired by H186A mutation. Introduction of reverse mutations into the equivalent position in DJ-1 (A107H) selectively eliminated its esterase activity without compromising α-oxoaldehyde hydratase activity. The obtained results suggest that differences in the amino acid sequences near the active site contributed to acquisition of esterase activity in vitro and provide an important clue to the origin and significance of DJ-1 esterase activity.

Parkinson's disease variant detection and disclosure: PD GENEration, a North American study.

Variants in seven genes (LRRK2, GBA1, PRKN, SNCA, PINK1, PARK7 and VPS35) have been formally adjudicated as causal contributors to Parkinson's disease; however, individuals with Parkinson's disease are often unaware of their genetic status since clinical testing is infrequently offered. As a result, genetic information is not incorporated into clinical care, and variant-targeted precision medicine trials struggle to enrol people with Parkinson's disease. Understanding the yield of genetic testing using an established gene panel in a large, geographically diverse North American population would help patients, clinicians, clinical researchers, laboratories and insurers better understand the importance of genetics in approaching Parkinson's disease. PD GENEration is an ongoing multi-centre, observational study (NCT04057794, NCT04994015) offering genetic testing with results disclosure and genetic counselling to those in the US (including Puerto Rico), Canada and the Dominican Republic, through local clinical sites or remotely through self-enrolment. DNA samples are analysed by next-generation sequencing including deletion/duplication analysis (Fulgent Genetics) with targeted testing of seven major Parkinson's disease-related genes. Variants classified as pathogenic/likely pathogenic/risk variants are disclosed to all tested participants by either neurologists or genetic counsellors. Demographic and clinical features are collected at baseline visits. Between September 2019 and June 2023, the study enrolled 10 510 participants across >85 centres, with 8301 having received results. Participants were: 59% male; 86% White, 2% Asian, 4% Black/African American, 9% Hispanic/Latino; mean age 67.4 ± 10.8 years. Reportable genetic variants were observed in 13% of all participants, including 18% of participants with one or more 'high risk factors' for a genetic aetiology: early onset (<50 years), high-risk ancestry (Ashkenazi Jewish/Basque/North African Berber), an affected first-degree relative; and, importantly, in 9.1% of people with none of these risk factors. Reportable variants in GBA1 were identified in 7.7% of all participants; 2.4% in LRRK2; 2.1% in PRKN; 0.1% in SNCA; and 0.2% in PINK1, PARK7 or VPS35 combined. Variants in more than one of the seven genes were identified in 0.4% of participants. Approximately 13% of study participants had a reportable genetic variant, with a 9% yield in people with no high-risk factors. This supports the promotion of universal access to genetic testing for Parkinson's disease, as well as therapeutic trials for GBA1 and LRRK2-related Parkinson's disease.

Parkinson's disease protein PARK7 prevents metabolite and protein damage caused by a glycolytic metabolite.

Cells are continuously exposed to potentially dangerous compounds. Progressive accumulation of damage is suspected to contribute to neurodegenerative diseases and aging, but the molecular identity of the damage remains largely unknown. Here we report that PARK7, an enzyme mutated in hereditary Parkinson's disease, prevents damage of proteins and metabolites caused by a metabolite of glycolysis. We found that the glycolytic metabolite 1,3-bisphosphoglycerate (1,3-BPG) spontaneously forms a novel reactive intermediate that avidly reacts with amino groups. PARK7 acts by destroying this intermediate, thereby preventing the formation of proteins and metabolites with glycerate and phosphoglycerate modifications on amino groups. As a consequence, inactivation of PARK7 (or its orthologs) in human cell lines, mouse brain, and Drosophila melanogaster leads to the accumulation of these damaged compounds, most of which have not been described before. Our work demonstrates that PARK7 function represents a highly conserved strategy to prevent damage in cells that metabolize carbohydrates. This represents a fundamental link between metabolism and a type of cellular damage that might contribute to the development of Parkinson's disease.

FKBP3 aggravates the malignant phenotype of diffuse large B-cell lymphoma by PARK7-mediated activation of Wnt/ß-catenin signalling.

Diffuse large B-cell lymphoma (DLBCL) is difficult to treat due to the high recurrence rate and therapy intolerance, so finding potential therapeutic targets for DLBCL is critical. FK506-binding protein 3 (FKBP3) contributes to the progression of various cancers and is highly expressed in DLBCL, but the role of FKBP3 in DLBCL and its mechanism are not clear. Our study demonstrated that FKBP3 aggravated the proliferation and stemness of DLBCL cells, and tumour growth in a xenograft mouse model. The interaction between FKBP3 and parkinsonism associated deglycase (PARK7) in DB cells was found using co-immunoprecipitation assay. Knockdown of FKBP3 enhanced the degradation of PARK7 through increasing its ubiquitination modification. Forkhead Box O3 (FOXO3) belongs to the forkhead family of transcription factors and inhibits DLBCL, but the underlying mechanism has not been reported. We found that FOXO3 bound the promoter of FKBP3 and then suppressed its transcription, eventually weakening DLBCL. Mechanically, FKBP3 activated Wnt/ß-catenin signalling pathway mediated by PARK7. Together, FKBP3 increased PARK7 and then facilitated the malignant phenotype of DLBCL through activating Wnt/ß-catenin pathway. These results indicated that FKBP3 might be a potential therapeutic target for the treatment of DLBCL.

Succinylation of Park7 activates a protective metabolic response to acute kidney injury.

Acute kidney injury (AKI) is extremely prevalent among hospitalizations and presents a significant risk for the development of chronic kidney disease and increased mortality. Ischemia caused by shock, trauma, and transplant are common causes of AKI. To attenuate ischemic AKI therapeutically, we need a better understanding of the physiological and cellular mechanisms underlying damage. Instances of ischemia are most damaging in proximal tubule epithelial cells (PTECs) where hypoxic signaling cascades, and perhaps more rapidly, posttranslational modifications (PTMs), act in concert to change cellular metabolism. Here, we focus on the effects of the understudied PTM, lysine succinylation. We have previously shown a protective effect of protein hypersuccinylation on PTECs after depletion of the desuccinylase sirtuin5. General trends in the results suggested that hypersuccinylation led to upregulation of peroxisomal activity and was protective against kidney injury. Included in the list of changes was the Parkinson's-related deglycase Park7. There is little known about any links between peroxisome activity and Park7. In this study, we show in vitro and in vivo that Park7 has a crucial role in protection from AKI and upregulated peroxisome activity. These data in combination with published results of Park7's protective role in cardiovascular damage and chronic kidney disease lead us to hypothesize that succinylation of Park7 may ameliorate oxidative damage resulting from AKI and prevent disease progression. This novel mechanism provides a potential therapeutic mechanism that can be targeted.NEW & NOTEWORTHY Succinylation is an understudied posttranslational modification that has been shown to increase peroxisomal activity. Furthermore, increased peroxisomal activity has been shown to reduce oxidative stress and protect proximal tubules after acute kidney injury. Analysis of mass spectrometry succinylomic and proteomic data reveals a novel role for Parkinson's related Park7 in mediating Nrf2 antioxidant response after kidney injury. This novel protection pathway provides new insights for kidney injury prevention and development of novel therapeutics.

A novel compound alleviates oxidative stress via PKA/CREB1-mediated DJ-1 upregulation.

Oxidative stress is one of the major culprits causing dopaminergic neuron loss in Parkinson's disease (PD). DJ-1 is a protein with multiple actions against oxidative stress, apoptosis, neuroinflammation, etc. DJ-1 expression is decreased in sporadic PD, therefore increasing DJ-1 expression might be beneficial in PD treatment. However, drugs known to upregulate DJ-1 are still lacking. In this study, we identified a novel DJ-1-elevating compound called ChemJ through luciferase assay-based high-throughput compound screening in SH-SY5Y cells and confirmed that ChemJ upregulated DJ-1 in SH-SY5Y cell line and primary cortical neurons. DJ-1 upregulation by ChemJ alleviated MPP+-induced oxidative stress. In exploring the underlying mechanisms, we found that the transcription factor CREB1 bound to DJ-1 promoter and positively regulated its expression under both unstressed and 1-methyl-4-phenylpyridinium-induced oxidative stress conditions and that ChemJ promoted DJ-1 expression via activating PKA/CREB1 pathway in SH-SY5Y cells. Our results demonstrated that ChemJ alleviated the MPP+-induced oxidative stress through a PKA/CREB1-mediated regulation of DJ-1 expression, thus offering a novel and promising avenue for PD treatment.

PARK7/DJ-1 deficiency impairs microglial activation in response to LPS-induced inflammation.

BACKGROUND: Specific microglia responses are thought to contribute to the development and progression of neurodegenerative diseases, including Parkinson's disease (PD). However, the phenotypic acquisition of microglial cells and their role during the underlying neuroinflammatory processes remain largely elusive. Here, according to the multiple-hit hypothesis, which stipulates that PD etiology is determined by a combination of genetics and various environmental risk factors, we investigate microglial transcriptional programs and morphological adaptations under PARK7/DJ-1 deficiency, a genetic cause of PD, during lipopolysaccharide (LPS)-induced inflammation. METHODS: Using a combination of single-cell RNA-sequencing, bulk RNA-sequencing, multicolor flow cytometry and immunofluorescence analyses, we comprehensively compared microglial cell phenotypic characteristics in PARK7/DJ-1 knock-out (KO) with wildtype littermate mice following 6- or 24-h intraperitoneal injection with LPS. For translational perspectives, we conducted corresponding analyses in human PARK7/DJ-1 mutant induced pluripotent stem cell (iPSC)-derived microglia and murine bone marrow-derived macrophages (BMDMs). RESULTS: By excluding the contribution of other immune brain resident and peripheral cells, we show that microglia acutely isolated from PARK7/DJ-1 KO mice display a distinct phenotype, specially related to type II interferon and DNA damage response signaling, when compared with wildtype microglia, in response to LPS. We also detected discrete signatures in human PARK7/DJ-1 mutant iPSC-derived microglia and BMDMs from PARK7/DJ-1 KO mice. These specific transcriptional signatures were reflected at the morphological level, with microglia in LPS-treated PARK7/DJ-1 KO mice showing a less amoeboid cell shape compared to wildtype mice, both at 6 and 24 h after acute inflammation, as also observed in BMDMs. CONCLUSIONS: Taken together, our results show that, under inflammatory conditions, PARK7/DJ-1 deficiency skews microglia towards a distinct phenotype characterized by downregulation of genes involved in type II interferon signaling and a less prominent amoeboid morphology compared to wildtype microglia. These findings suggest that the underlying oxidative stress associated with the lack of PARK7/DJ-1 affects microglia neuroinflammatory responses, which may play a causative role in PD onset and progression.

cPGA hydrolase assay of DJ-1 in crude cell lysates: Implications for sensing of oxidative stress.

Cyclic 3-phosphosphoglyceric anhydride (cPGA), a side product of glycolysis, acylates cellular amines and thiols to form amides and thioesters, respectively. Since these acylation reactions are harmful, organisms rely on a protein, known as DJ-1 in humans, to inactivate cPGA. Inactivation of cPGA likely plays a significant role in cytoprotection by DJ-1, but further progress in this direction is hampered by the lack of quantitative assays to measure the cPGA hydrolase activity of DJ-1 in biological samples. Here we report an optimized procedure for preparation of cPGA which is then used as a substrate to quantify enzymatic activity of DJ-1. The end-point assay for cPGA hydrolase uses dilute cell lysates to hydrolyze cPGA for 0.5-3.5 min followed by conversion of the remaining cPGA into thioester for spectrophotometric quantitation. We illustrate the utility of this assay by showing that higher levels of cPGA hydrolase activity result in better protection from acylation by cPGA. Moreover, the decrease of cPGA hydrolase activity due to oxidation of the catalytic cysteine of DJ-1 under oxidative stress and its subsequent recovery can be monitored using the assay. This relatively simple assay allows functional characterization of DJ-1 in biological samples through quantitative assessment of its cPGA hydrolase activity.

DJ-1 preserves ischemic postconditioning-induced cardioprotection in STZ-induced type 1 diabetic rats: role of PTEN and DJ-1 subcellular translocation.

BACKGROUND: Ischemic postconditioning (IPostC) has been reported as a promising method for protecting against myocardial ischemia-reperfusion (MI/R) injury. Our previous study found that the infarct-limiting effect of IPostC is abolished in the heart of diabetes whose cardiac expression of DJ-1 (also called PARK7, Parkinsonism associated deglycase) is reduced. However, the role and in particular the underlying mechanism of DJ-1 in the loss of sensitivity to IPostC-induced cardioprotection in diabetic hearts remains unclear. METHODS: Streptozotocin-induced type 1 diabetic rats were subjected to MI/R injury by occluding the left anterior descending artery (LAD) and followed by reperfusion. IPostC was induced by three cycles of 10s of reperfusion and ischemia at the onset of reperfusion. AAV9-CMV-DJ-1, AAV9-CMV-C106S-DJ-1 or AAV9-DJ-1 siRNA were injected via tail vein to either over-express or knock-down DJ-1 three weeks before inducing MI/R. RESULTS: Diabetic rats subjected to MI/R exhibited larger infarct area, more severe oxidative injury concomitant with significantly reduced cardiac DJ-1 expression and increased PTEN expression as compared to non-diabetic rats. AAV9-mediated cardiac DJ-1 overexpression, but not the cardiac overexpression of DJ-1 mutant C106S, restored IPostC-induced cardioprotection and this effect was accompanied by increased cytoplasmic DJ-1 translocation toward nuclear and mitochondrial, reduced PTEN expression, and increased Nrf-2/HO-1 transcription. Our further study showed that AAV9-mediated targeted DJ-1 gene knockdown aggravated MI/R injury in diabetic hearts, and this exacerbation of MI/R injury was partially reversed by IPostC in the presence of PTEN inhibition or Nrf-2 activation. CONCLUSIONS: These findings suggest that DJ-1 preserves the cardioprotective effect of IPostC against MI/R injury in diabetic rats through nuclear and mitochondrial DJ-1 translocation and that inhibition of cardiac PTEN and activation of Nrf-2/HO-1 may represent the major downstream mechanisms whereby DJ-1 preserves the cardioprotective effect of IPostC in diabetes.

Extracellular DJ-1 induces sterile inflammation in the ischemic brain.

Inflammation is implicated in the onset and progression of various diseases, including cerebral pathologies. Here, we report that DJ-1, which plays a role within cells as an antioxidant protein, functions as a damage-associated molecular pattern (DAMP) and triggers inflammation if released from dead cells into the extracellular space. We first found that recombinant DJ-1 protein induces the production of various inflammatory cytokines in bone marrow-derived macrophages (BMMs) and dendritic cells (BMDCs). We further identified a unique peptide sequence in the αG and αH helices of DJ-1 that activates Toll-like receptor 2 (TLR2) and TLR4. In the ischemic brain, DJ-1 is released into the extracellular space from necrotic neurons within 24 h after stroke onset and makes direct contact with TLR2 and TLR4 in infiltrating myeloid cells. Although DJ-1 deficiency in a murine model of middle cerebral artery occlusion did not attenuate neuronal injury, the inflammatory cytokine expression in infiltrating immune cells was significantly decreased. Next, we found that the administration of an antibody to neutralize extracellular DJ-1 suppressed cerebral post-ischemic inflammation and attenuated ischemic neuronal damage. Our results demonstrate a previously unknown function of DJ-1 as a DAMP and suggest that extracellular DJ-1 could be a therapeutic target to prevent inflammation in tissue injuries and neurodegenerative diseases.

DJ-1 depletion prevents immunoaging in T-cell compartments.

Decline in immune function during aging increases susceptibility to different aging-related diseases. However, the underlying molecular mechanisms, especially the genetic factors contributing to imbalance of naïve/memory T-cell subpopulations, still remain largely elusive. Here, we show that loss of DJ-1 encoded by PARK7/DJ-1, causing early-onset familial Parkinson's disease (PD), unexpectedly diminished signs of immunoaging in T-cell compartments of both human and mice. Compared with two gender-matched unaffected siblings of similar ages, the index PD patient with DJ-1 deficiency showed a decline in many critical immunoaging features, including almost doubled non-senescent T cells. The observation was further consolidated by the results in 45-week-old DJ-1 knockout mice. Our data demonstrated that DJ-1 regulates several immunoaging features via hematopoietic-intrinsic and naïve-CD8-intrinsic mechanisms. Mechanistically, DJ-1 depletion reduced oxidative phosphorylation (OXPHOS) and impaired TCR sensitivity in naïve CD8 T cells at a young age, accumulatively leading to a reduced aging process in T-cell compartments in older mice. Our finding suggests an unrecognized critical role of DJ-1 in regulating immunoaging, discovering a potent target to interfere with immunoaging- and aging-associated diseases.

TrkB inhibition of DJ-1 degradation promotes the growth and maintenance of cancer stem cell characteristics in hepatocellular carcinoma.

Although TrkB may be associated with the pathogenesis of various cancer by upregulation, how upregulation of TrkB led to tumor progression in hepatocellular carcinoma (HCC) and the signaling mechanisms by which TrkB induces motility, invasion, metastasis, drug resistance, and acquisition of self-renewal traits has remained unclear. Here, we demonstrated that TrkB was significantly upregulated in highly metastatic HCC cells and HCC patients. Also, the increased TrkB levels were significantly correlated with tumor stages and poor survival of HCC patients. Furthermore, the upregulated TrkB expression enhances the metastatic ability of HCC cells through reduced anoikis sensitivity, induced migration, and colony formation. Most strikingly, TrkB markedly enhances the activation of STAT3 by preventing DJ-1 degradation through the formation of the TrkB/DJ-1 complex. This signaling mechanism is responsible for triggering cellular traits of highly aggressive HCC. The activation of the EMT program of HCC via increasing DJ-1 stability by TrkB induces the gain of cancer stem cell states and chemoresistance via the upregulation of stem cells cell markers and ABC transporters. Also, TrkB-mediated inhibition of DJ-1 degradation promotes tumor formation and metastasizes to other organs in vivo. Our observations illustrate that TrkB is a prognostic and therapeutic targeting in promoting aggressiveness and metastasis of HCC.

Diallyl disulfide antagonizes DJ-1 mediated proliferation, epithelial-mesenchymal transition, and chemoresistance in gastric cancer cells.

Diallyl disulfide (DADS), an organic component of allicin abstracted from garlic, possesses multi-target antitumor activity. DJ-1 performs a vital function in promoting AKT aberrant activation via down-regulating phosphatase and tensin homologue (PTEN) in tumors. It is unknown the involvement of DJ-1 in epithelial-mesenchymal transition (EMT) of gastric cancer (GC) cells. The purpose of this study is to investigate whether diallyl disulfide (DADS) intervenes in the role of DJ-1 in GC. Based on the identification that the correlation between high DJ-1 and low PTEN expression in GC was implicated in clinical progression, we illuminated that down-regulation of DJ-1 by DADS aided in an increase in PTEN expression and a decrease in phosphorylated AKT levels, which was in line with the results manifested in the DJ-1 knockdown and overexpressed cells, concurrently inhibiting proliferation, EMT, migration, and invasion. Furthermore, the antagonistic effects of DADS on DJ-1 were observed in in vivo experiments. Additionally, DADS mitigated the DJ-1-associated drug resistance. The current study revealed that DJ-1 is one of potential targets for DADS, which hopefully provides a promising strategy for prevention and adjuvant chemotherapy of GC.

Parkinson biomarker determination with an Au microflower-enhanced electrochemical immunosensor using non-Faradaic capacitance measurements.

This work comprehends the development and characterization of a carbon black-based electrode modified with Au microflowers to increase its effect as a capacitance biosensor for the determination of PARK7/DJ-1. Due to its high surface-to-volume ratio and biocompatibility, Au particles are suitable for antibody binding, and by monitoring surface capacitance, it is possible to identify the immune-pair interaction. Au microflowers allowed the adequate immobilization of Parkinsonian-related proteins: PARK7/DJ-1 and its antibody. The protein is associated with several antioxidant mechanisms, but its abnormal concentrations or mutations can be the cause of the loss of dopaminergic neurons, leading to Parkinson's disease. The device was characterized by scanning electron microscopy and cyclic voltammetry, revealing the flower-like structures and the electrochemically-interest enhancements they provide, such as increased heterogeneous electron transfer rate coefficient and electroactive area. The self-assembled monolayers of different molecules were optimized with the aid of 22 central composite experiments and a linear calibration curve was obtained between 0.700 and 120 ng mL-1 of PARK7/DJ-1, with a limit of detection of 0.207 ng mL-1. The data confirms that the addition of Au microflowers enhanced the electrochemical signal of the device, as well as allowed for the determination of an early stage Parkinson's disease biomarker with appreciable analytical performance.

Peptide Activator Stabilizes DJ-1 Structure and Enhances Its Activity.

DJ-1 is a vital enzyme involved in the maintenance of mitochondrial health, and its mutation has been associated with an increased risk of Parkinson's disease (PD). Effective regulation of DJ-1 activity is essential for the well-being of mitochondria, and DJ-1 is thus a potential target for PD drug development. In this study, two peptides (15EEMETIIPVDVMRRA29 and 47SRDVVICPDA56) were utilized with the aim of enhancing the activity of DJ-1. The mechanisms underlying the activity enhancement by these two peptides were investigated using hydrogen/deuterium exchange mass spectrometry (HDXMS). The HDXMS results revealed distinct mechanisms. Peptide 1 obstructs the access of solvent to the dimer interface and stabilizes the α/ß hydrolase structure, facilitating substrate binding to a stabilized active site. Conversely, peptide 2 induces a destabilization of the α/ß hydrolase core, enhancing substrate accessibility and subsequently increasing DJ-1 activity. The binding of these two peptides optimizes the activity site within the dimeric structure. These findings offer valuable insights into the mechanisms underlying the activity enhancement of DJ-1 by the two peptides, potentially aiding the development of new drugs that can enhance the activity of DJ-1 and, consequently, advance PD treatment.

Knockdown of DJ-1 Resulted in a Coordinated Activation of the Innate Immune Antiviral Response in HEK293 Cell Line.

PARK7, also known as DJ-1, plays a critical role in protecting cells by functioning as a sensitive oxidation sensor and modulator of antioxidants. DJ-1 acts to maintain mitochondrial function and regulate transcription in response to different stressors. In this study, we showed that cell lines vary based on their antioxidation potential under basal conditions. The transcriptome of HEK293 cells was tested following knockdown (KD) of DJ-1 using siRNAs, which reduced the DJ-1 transcripts to only 12% of the original level. We compared the expression levels of 14k protein-coding transcripts and 4.2k non-coding RNAs relative to cells treated with non-specific siRNAs. Among the coding genes, approximately 200 upregulated differentially expressed genes (DEGs) signified a coordinated antiviral innate immune response. Most genes were associated with the regulation of type 1 interferons (IFN) and the induction of inflammatory cytokines. About a quarter of these genes were also induced in cells treated with non-specific siRNAs that were used as a negative control. Beyond the antiviral-like response, 114 genes were specific to the KD of DJ-1 with enrichment in RNA metabolism and mitochondrial functions. A smaller set of downregulated genes (58 genes) was associated with dysregulation in membrane structure, cell viability, and mitophagy. We propose that the KD DJ-1 perturbation diminishes the protective potency against oxidative stress. Thus, it renders the cells labile and responsive to the dsRNA signal by activating a large number of genes, many of which drive apoptosis, cell death, and inflammatory signatures. The KD of DJ-1 highlights its potency in regulating genes associated with antiviral responses, RNA metabolism, and mitochondrial functions, apparently through alteration in STAT activity and downstream signaling. Given that DJ-1 also acts as an oncogene in metastatic cancers, targeting DJ-1 could be a promising therapeutic strategy where manipulation of the DJ-1 level may reduce cancer cell viability and enhance the efficacy of cancer treatments.

DJ-1 Molecular Chaperone Activity Depresses Tau Aggregation Propensity through Interaction with Monomers.

Tau aggregate-bearing lesions are pathological markers and potential mediators of tauopathic neurodegenerative diseases, including Alzheimer's disease. The molecular chaperone DJ-1 colocalizes with tau pathology in these disorders, but it has been unclear what functional link exists between them. In this study, we examined the consequences of tau/DJ-1 interaction as isolated proteins in vitro. When added to full-length 2N4R tau under aggregation-promoting conditions, DJ-1 inhibited both the rate and extent of filament formation in a concentration-dependent manner. Inhibitory activity was low affinity, did not require ATP, and was not affected by substituting oxidation incompetent missense mutation C106A for wild-type DJ-1. In contrast, missense mutations previously linked to familial Parkinson's disease and loss of α-synuclein chaperone activity, M26I and E64D, displayed diminished tau chaperone activity relative to wild-type DJ-1. Although DJ-1 directly bound the isolated microtubule-binding repeat region of tau protein, exposure of preformed tau seeds to DJ-1 did not diminish seeding activity in a biosensor cell model. These data reveal DJ-1 to be a holdase chaperone capable of engaging tau as a client in addition to α-synuclein. Our findings support a role for DJ-1 as part of an endogenous defense against the aggregation of these intrinsically disordered proteins.

PARK7 Catalyzes Stereospecific Detoxification of Methylglyoxal Consistent with Glyoxalase and Not Deglycase Function.

The protein PARK7 (also known as DJ-1) has been implicated in several diseases, with the most notable being Parkinson's disease. While several molecular and cellular roles have been ascribed to DJ-1, there is no real consensus on what its true cellular functions are and how the loss of DJ-1 function may contribute to the pathogenesis of Parkinson's disease. Recent reports have implicated DJ-1 in the detoxification of several reactive metabolites that are produced during glycolytic metabolism, with the most notable being the α-oxoaldehyde species methylglyoxal. While it is generally agreed that DJ-1 is able to metabolize methylglyoxal to lactate, the mechanism by which it does so is hotly debated with potential implications for cellular function. In this work, we provide definitive evidence that recombinant DJ-1 produced in human cells prevents the stable glycation of other proteins through the conversion of methylglyoxal or a related alkynyl dicarbonyl probe to their corresponding α-hydroxy carboxylic acid products. This protective action of DJ-1 does not require a physical interaction with a target protein, providing direct evidence for a glutathione-free glyoxalase and not a deglycase mechanism of methylglyoxal detoxification. Stereospecific liquid chromatography-mass spectrometry (LC-MS) measurements further uncovered the existence of nonenzymatic production of racemic lactate from MGO under physiological buffer conditions, whereas incubation with DJ-1 predominantly produces l-lactate. Collectively, these studies provide direct support for the stereospecific conversion of MGO to l-lactate by DJ-1 in solution with negligible or no contribution of direct protein deglycation.

Parkinsonism-Associated Protein DJ-1 Is an Antagonist, Not an Eraser, for Protein Glycation.

Advanced glycation end-products (AGEs) are irreversible protein modifications that are strongly associated with aging and disease. Recently, the Parkinsonism-associated protein DJ-1 has been reported to exhibit deglycase activity that erases early glycation intermediates and stable AGEs from proteins. In this work, we use mass spectrometry and western blot to demonstrate that DJ-1 is not a deglycase and cannot remove AGEs from protein or peptide substrates. Instead, our studies revealed that DJ-1 antagonizes glycation through glyoxalase activity that detoxifies the potent glycating agent methylglyoxal (MGO) to lactate. We further show that attenuated glycation in the presence of DJ-1 can be attributed solely to its ability to decrease the available concentration of MGO. Our studies also provide evidence that DJ-1 is allosterically activated by glutathione. Together, this work reveals that although DJ-1 is not a genuine deglycase, it still harbors the ability to prevent AGE formation and can be used as a valuable tool to investigate metabolic stress.

Oncoprotein DJ-1 interacts with mTOR complexes to effect transcription factor Hif1α-dependent expression of collagen I (α2) during renal fibrosis.

Proximal tubular epithelial cells respond to transforming growth factor ß (TGFß) to synthesize collagen I (α2) during renal fibrosis. The oncoprotein DJ-1 has previously been shown to promote tumorigenesis and prevent apoptosis of dopaminergic neurons; however, its role in fibrosis signaling is unclear. Here, we show TGFß-stimulation increased expression of DJ-1, which promoted noncanonical mTORC1 and mTORC2 activities. We show DJ-1 augmented the phosphorylation/activation of PKCßII, a direct substrate of mTORC2. In addition, coimmunoprecipitation experiments revealed association of DJ-1 with Raptor and Rictor, exclusive subunits of mTORC1 and mTORC2, respectively, as well as with mTOR kinase. Interestingly, siRNAs against DJ-1 blocked TGFß-stimulated expression of collagen I (α2), while expression of DJ-1 increased expression of this protein. In addition, expression of dominant negative PKCßII and siRNAs against PKCßII significantly inhibited TGFß-induced collagen I (α2) expression. In fact, constitutively active PKCßII abrogated the effect of siRNAs against DJ-1, suggesting a role of PKCßII downstream of this oncoprotein. Moreover, we demonstrate expression of collagen I (α2) stimulated by DJ-1 and its target PKCßII is dependent on the transcription factor hypoxia-inducible factor 1α (Hif1α). Finally, we show in the renal cortex of diabetic rats that increased TGFß was associated with enhanced expression of DJ-1 and activation of mTOR and PKCßII, concomitant with increased Hif1α and collagen I (α2). Overall, we identified that DJ-1 affects TGFß-induced expression of collagen I (α2) via an mTOR-, PKCßII-, and Hif1α-dependent mechanism to regulate renal fibrosis.