Regular articleMicroRNA-494 reduces DJ-1 expression and exacerbates neurodegeneration
Introduction
Parkinson's disease (PD) is a neurodegenerative disease that affects approximately 1% of persons 65 years or older of age and 5% of those 85 years or older (Tanner et al., 1996). The featured pathological change of PD is the selective and progressive loss of dopaminergic (DA) neurons, but the molecular mechanism underlying neuronal death still has not been clarified (Lang et al., 1998). Nevertheless, increasing evidence has suggested that both environmental and genetic factors play important roles in the pathogenesis of the disease (Olanow et al., 1999).
Epidemiologic studies have suggested that increased PD is associated with overexposure to environmental factors such as iron, manganese, herbicides, and pesticides (de Lau and Breteler, 2006, Warner and Schapira, 2003). These neurotoxins are believed to induce oxidative stress, which is thought to be a significant cause of PD (Jenner, 1991, Jenner et al., 1992). Studies in hereditary PD revealed that genetic factors also played an important role in the development of PD. To date, more than 13 loci and 9 genes have been identified (Farrer, 2006). Among these genes, DJ-1 is thought to be a molecular chaperone (Lee et al., 2003, Tao and Tong, 2003) and oxidative sensor (Choi et al., 2006), participating in both familial and sporadic PD (Choi et al., 2006). Most researches have been focused on the anti-oxidative stress and molecular chaperone function of DJ-1. Our previous studies showed that DJ-1 regulated SOD1 expression through the Erk1/2–Elk1 pathway in its protective response to oxidative insult (Wang et al., 2011a), and that DJ-1 acted as a molecular chaperone to inhibit MAP1B aggregation which could lead to endoplasmic reticulum (ER) stress-induced neuronal apoptosis (Wang et al., 2011b). Nonetheless, DJ-1 gene regulation has not been extensively explored in the brain. Loss of function mutation of DJ-1 was thought to be the cause of autosomal recessive familial PD (Bonifati et al., 2004). The most frequent DJ-1 mutant L166P fails to form dimers, and monomeric mutant DJ-1 is unstable and degraded rapidly (Moore et al., 2003). Shen Yong and his colleagues discovered that the level of total DJ-1 protein was significantly reduced in the substantia nigra of sporadic PD patients. Moreover, in the PD cortex mitochondria fraction, the high-molecular-weight (HMW) complexes of DJ-1 are significantly lower than in the non-pathological controls (Nural et al., 2009). These results suggest that lower DJ-1 level in PD patients may contribute to the pathogenesis. However, the molecular mechanisms underlying the decreased DJ-1 level are not yet clear.
Regulation of gene expression is manifested mainly in the following aspects: (1) transcriptional regulation; (2) mature mRNA processing; and (3) translational regulation. In recent years, as a post-transcriptional regulation of target gene expression, the roles of microRNAs in the development and disease progression have received widespread concerns. MicroRNAs (miRNAs) are endogenous ∼23 nt non-coding RNAs that play important gene-regulatory roles in animals and plants by pairing to the 3′ untranslated region (3′UTR) of mRNAs of protein-coding genes to direct their post-transcriptional repression (Bartel, 2009). Mounting studies in recent years have shown that miRNAs play crucial roles in the regulation of diverse biological processes, such as development, inflammation, and tumorigenesis (Ambros, 2004). One specific microRNA may regulate the expression of many different mRNAs, and many different microRNAs can regulate the expression of a specific mRNA (Bartel, 2009). Therefore, we hypothesized that the regulation of the DJ-1 expression by microRNAs may be involved in the pathogenesis of sporadic PD.
In the present study, we identified DJ-1 as a target of miR-494; and overexpression of miR-494 decreased DJ-1 expression, rendered cells more susceptible to oxidative stress, and resulted in loss of DA neurons in mice. Our data also suggest that miR-494 contributes to exacerbation of oxidative stress and loss of DA neurons in vitro or in vivo.
Section snippets
Antibodies and reagents
The following antibodies were used: rabbit polyclonal anti–PARK7/DJ-1 antibody (Abcam, ab18257, Cambridge, UK), rabbit polyclonal anti–tyrosine hydroxylase antibody (Millipore, AB152, Massachusetts, America), mouse monoclonal anti–β-actin antibody (Sigma-Aldrich, Clone AC-15, St. Louis, MO, USA), horseradish peroxidase (HRP)–conjugated goat anti-mouse IgG, goat anti-rabbit IgG, Alexa Fluor 594–AffiniPure goat anti-rabbit IgG (Jackson ImmunoResearch Laborataries, PA, USA).
DJ-1 as a target of miR-494
Using two miRNA databases (TargetScan.org and microRNA.org), we identified several miRNA candidates that may potentially target DJ-1, including miR-23a, miR-23b, miR-329, miR-362-3p, miR-378, miR-466g, miR-494, miR-670, miR-672, miR-743b-5p, and miR-883a-5p. To determine which of these microRNAs were capable of repressing the expression level of DJ-1 protein, transient transfections with microRNA mimics were performed in 3T3 cells, the endogenous DJ-1 expression level of which was high. As a
Discussion
DJ-1 is believed to play an important protective role against oxidative insult in the brains of PD. It was reported that DJ-1 mutations could cause instability and a loss function of the protein. However, what caused the reduced DJ-1 in brains of sporadic PD patients still remains a puzzle.
This study revealed a previously unreported mechanism by which DJ-1 level was regulated in the nervous system. Specifically, one brain-enriched miRNA, miR-494, was involved in the regulation of DJ-1
Disclosure statement
All authors certify that they do not have any actual or potential conflicts of interest in regard to this work.
Acknowledgements
This work was supported by the National Program of Basic Research (2010CB945200, 2011CB504104) of China, Natural Science Fund (30971031, 81200980, 81129018, 81371407), The twelfth Five-year National Science and Technology Support Program (2012BAI10B03), and Shanghai Key Project of Basic Science Research (10411954500).
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R.X. and Z.W. contributed equally to this work.