Study On The Mutation Of Parkin Gene In Chinese Early-Onset And Late-Onset Patients With Parkinson's Disease

Background: Parkinson's disease (PD) is a common neurodegenerative disease with rest tremor, bradykinesia, rigidity and postural instability. Pathological findings show that nigral dopamine neurons are greatly diminished and Lewy bodies are present in the remaining neurons. Most people with Parkinson's disease do not have a family history. About 5~ 10% of patients have a first-degree relative with the disease. The inheritance mode of Parkinson's disease includes autosomal recessive and autosomal dominant types. Parkin gene was cloned in an autosomal recessive juvenile Parkinsonism (AR-JP). Recently varied mutations of parkin gene were detected in sporadic and familial or early-onset and later-onset patients with Parkinson's disease. The study on the relationship between Parkinson's disease and mutations of parkin gene becomes hotspot in this domain.Human parkin gene encodes a 465 amino acid protein that has a molecular weight of~ 52 kDa. Parkin, an E3 ligase, is a component of the ubiqutin/proteosome pathway (UPP). The UPP can selectively degrade mutative or mistaken protein and injured protein after translation or oxidative stress through a three step fashion .Step 1,Ub is activated by E1 Ub activating enzyme;step 2, Ub is accepted by Ubconjugating enzyme E2;step 3, Ub is then transferred via the Ub-proteinligase E3 to the substrate. The carboxyterminal glycin of Ub can covalently combine with lysine of the substrate to finish the ubiquitination process. Repetitive ubiquitination turns to polyubiquitinate substrates. Once polyubiquitinated the targets are accepted as substrate for 26S multiprotein complex proteasome-mediated degradation. In this degradation, polyubiquitinate substrates can release ubiquitin by ubiquitin carboxyterminal hydrolase (UCH). Under normal condition the substrates are degraded by parkin protein through UPP to maintain the normal function of dopamine neuron. But it is not known how parkin protein works to those substrates. The parkin gene with nonsense, missense or insert mutation can cause stop codon which finish translation earlier or change coding amino acid sequence result in truncation or null protein. The homozygous, heterozygous exonic deletion or exon duplication of parkin gene can reduce the important domain of parkin protein. Thus the intracellular accumulation of substrates leads to neuron degeneraten because not be ubiquited.The overseas studies find varied mutation types in 12 exons of parkin gene including missense mutation, frameshift mutation and nonsense mutation, intraexonic insert and absence, exon homozygous and heterozygous deletion, exon duplication and compound heterozygous mutation. In addition, the SNPs in promoter, exon4 and exonlO are correlative with PD. Recently more than 70 mutation types are found in parkin gene. In familial PD (age onset less than 45 years old) the rate of parkin gene mutation is about 40% while 18% in sporadic PD (age onset less than 45 years old). Rate of parkin gene mutation in PD (age onset more than 50 years old) islower. That parkin gene mutation is rarely reported in China with fewer mutation types and lower mutation rate possibly relates to the method sensitivity and spectrum of exons.The mutation of parkin gene exists in sporadic and familial, early-onset and later-onset PD with varied imitative types. Parkin-linked PD is characterized by early onset of parkinsonian signs, slow disease progression;good response to L-dopa, and early levodopa-induced dyskinesias and motor fluctuations.lt is difficult to distinguish parkin-linked PD from idiopathic PD depending on clinical manifestation. PD with heterozygous mutation of parkin is the same as sporadic PD in clinical manifestation. Some familial PD with parkin mutation has an autosomal dominat inherited type. As E3 ligase, parkin protein plays a key role maintaining dopamine neuron's normal function. But the mechanism of reciprocity between parkin protein and correlative substrates is not clear-out. More study on the parkin gene mutation will definitude diagnoses of parkin-linked PD and PD diagnose before clinical manifestation.Objective: To observe the type, mode and rate of parkin gene mutation in Chinese early-onset patients and late-onset patients with Parkinson's disease (PD), illuminate the distribution of parkin gene mutation to Chinese sporadic patients with PD, and relationship between the inheritary mode of familial patients and parkin gene mutation, sustain the gene diagnosis of parkin-linked PD, add the spectrum of genotypes and phenotypes in PD.Methods: 27 early-onset patients and 19 late-onset patients were recruited according with diagnostic criterion of PD and were evaluated with unified Parkinson's diseaserating scale (UPDRS) and hoehn-yahr scale. Genomic DNA from those patients was extracted from peripheral blood leukocytes with standard procedures. The total 12 exons in parkin gene were detected by single strand conformation polymorphism analysis of polymerase chain reaction products (PCR-SSCP). The DNA sample with abnormal electrophoresis was sequenced to get realizing mutative type. Results'. ?Three early-onset patients were found to be carrier of exon homozygous deletions among them. The locations of deletion were on exon 3 (one case), exon 4 (two cases) .(2)V/L380 on exon 10 was found in four samples while three S/N167 on exon 4 that exhibited abnormal mobility shift on SSCP electrophoresis.? The mutative rate is 14% in 21 early-onset and nonfamilial patients with three mutative cases. Three single nucleotide polymorphisms were detected in 6 early-onset and familial patients while one in 21 early-onset and nonfamilial patients. Three single nucleotide polymorphisms were detected in 19 late-onset patients. Conclusion: ?The sporadic PD has some parkin gene mutation in China. (2)In the study of 21 early-onset and nonfamilial PD patients, two mutation types were found in three cases locating on exon 3 and exon4. The mutation rate in parkin gene was estimated at 14%. It suggests that mutations of parkin gene can play an important role in the pathogenesis of Chinese patients with PD. (3)S/N167 polymorphism can increase the risk of PD. ?whether V/L38O increases the risk ofPD need further studies.