Biochemical and Biophysical Research Communications
Mitochondrial DNA heteroplasmy rises in substantial nigra of aged PINK1 KO mice
Introduction
Parkinson’s disease (PD) is the most common age-related, neurodegenerative movement disorder, which causes a dramatic economic and social burden along with worldwide population aging [1]. Accompanying with the pathological hallmarks of dopaminergic neurons (DAs) degeneration in the pars compacta of the substantia nigra (SNc) and the formation of α-synuclein immunoreactive Lewy bodies in midbrain, PD patients exhibited progressive motor dysfunction including bradykinesia, rigidity, resting tremor and postural instability, as well as non-motor features such as dementia, depression, and anxiety [2].
Previous studies have shown that environmental and genetic factors both participate in PD progression [3]. Meanwhile, aging is the most important risk factor for PD, as the disease incidence increases 5–10 fold after the age of 60 [3,4]. Several pathological mechanisms are reported contributing to PD pathogenesis, especially mitochondrial dysfunction, as well as aberrant protein degradation, calcium imbalance, neuroinflammation, and oxidative stress et al. [5].
Mitochondrial dysfunction is one of the most crucial contributors to PD pathogenesis. Mitochondrial complex I defects and ATP production decreases were observed in PD patients’ brains [6]. Mitochondrial DNA (mtDNA), which encodes 13 essential components of electron transport chain (ETC) complexes, is also reported to be damaged both in brain of PD patients and aged persons [7,8].
MPTP is transformed into its toxic form MPP + by monoamine oxidase (MAO) in glial cells, the neurotoxin is transported into dopaminergic (DA) neurons and then inhibits the activity of mitochondrial respiratory chain complex Ⅰ and causes selective DA neurons degeneration and death in substantia nigra (SN) [9]. Some other pesticides including rotenone and paraquat, which lead to mitochondrial electron transport chain (ETC) dysfunctions, also increasing the risk of PD [10]. In addition, Patients with mutations in mtDNA polymerase POLG1 gene, which increased mtDNA mutation rate, exhibited dopaminergic neuron degeneration and Parkinsonism symptoms [11]. Increasing of somatic mtDNA mutations and cell degeneration have been observed in the substantia nigra of PD related pesticide rotenone treated rat [12], further supported the relationship between mtDNA mutations and PD.
The most common autosomal recessive causative genes PINK1 and Parkin associated with PD are both functioning in mitophagy and mitochondrial quality control. PINK1 is a serine/threonine protein kinase, which can be stabilized at the outer membrane of damaged mitochondria, then recruits and phosphorylates Parkin, a ubiquitin E3 ligase, which ubiquitinates its substrates and mediates mitophagy [13]. PINK1-null flies showed mitochondrial morphological change combining with mitochondrial dysfunction including decreases of ATP production, respiratory chain activity and mtDNA content [14]. Restore respiratory chain activity by expressing complexⅠ subunit-NdufA10 in PINK1-null flies could restore mitochondrial function [15]. Parkin KO and POLG1 proofreading domain mutation mouse model, but not POLG1 mutation alone, exhibited dopaminergic neuron loss and parkinsonism phenotype [16], further suggested that PINK1/Parkin might work together with mitochondrial DNA mutagenic stress and might contribute to PD pathogenesis. In addition, recent studies have shown that PINK1 can limit deleterious mitochondrial DNA mutations transmission by inhibiting local protein synthesis [17]. However, the exact influence of PINK1 to age-related somatic mtDNA mutations remains to be clarified.
In this study, we analyzed the mtDNA heteroplasmy, including SNV numbers (Single Nucleotide Variants), average variation frequency, variation pathogenicity and mtDNA copy numbers in PINK1 KO and Parkin KO mice. Our data revealed that the interaction between PINK1/Parkin pathway and the aging process, thus providing insight to elucidate the crosstalk of genetic and environmental factors in PD pathogenesis.
Section snippets
Materials and methods
Animals. The generation and genotyping of PINK1 KO and Parkin KO mice were previously described [18,19]. The experimental protocol was approved by the Ethics Review Committee for Animal Experimentation of Central South University. The genetic background of all the mouse models used in this study was C57BL/6J.
mtDNA preparation. Anesthetized mice were perfused with ice-cold 1X PBS and subsequently sacrificed by cervical dislocation. Brains were isolated and DNA was extracted from dissected
Total SNV numbers of mtDNA in brain tissues increased with aging
To maximize the sequencing depth of mitochondrial DNA, we utilized a rolling circle amplification method for mtDNA described in previous studies [20]. After the amplification, mtDNA proportion in sequencing data increased by tens to hundreds of times. Meanwhile, this amplification did not change the original mtDNA profile. As shown in (Fig. S2A), normalized depth of all 135 samples did not fluctuate dramatically. We further checked the transitions (Ti) and transversions (Tv) of all the
Discussion
The relationship between mtDNA mutations, aging process and PD is a mystery for decades. Previous studies have shown that mtDNA mutations are increased in brain of PD patients [27]. Meanwhile, mutations in Polymerase gamma 1 gene, the catalytic subunit of mtDNA replication polymerase, which leading to levodopa-responsive parkinsonian features in patients. In addition, mitochondria dysfunction is one of the main reasons for the progression of Parkinson’s disease. PINK1 and Parkin are the most
Declaration of competing interest
The authors declare that they have no conflict of interest.
Acknowledgments
This work was supported by the National Natural Science Foundation of China (NO. 31730036).
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