Ji-feng Guo

Identification of PRRT2 as the causative gene of paroxysmal kinesigenic dyskinesias

Paroxysmal kinesigenic dyskinesias is a paroxysmal movement disorder characterized by recurrent, brief attacks of abnormal involuntary movements induced by sudden voluntary movements. Although several loci, including the pericentromeric region of chromosome 16, have been linked to paroxysmal kinesigenic dyskinesias, the causative gene has not yet been identified. Here, we identified proline-rich transmembrane protein 2 (PRRT2) as a causative gene of paroxysmal kinesigenic dyskinesias by using a combination of exome sequencing and linkage analysis. Genetic linkage mapping with 11 markers that encompassed the pericentromeric of chromosome 16 was performed in 27 members of two families with autosomal dominant paroxysmal kinesigenic dyskinesias. Then, the whole-exome sequencing was performed in three patients from these two families. By combining the defined linkage region (16p12.1–q12.1) and the results of exome sequencing, we identified an insertion mutation c.649_650InsC (p.P217fsX7) in one family and a nonsense mutation c.487C>T (p.Q163X) in another family. To confirm our findings, we sequenced the exons and flanking introns of PRRT2 in another three families with paroxysmal kinesigenic dyskinesias. The c.649_650InsC (p.P217fsX7) mutation was identified in two of these families, whereas a missense mutation, c.796C>T (R266W), was identified in another family with paroxysmal kinesigenic dyskinesias. All of these mutations completely co-segregated with the phenotype in each family. None of these mutations was identified in 500 normal unaffected individuals of matched geographical ancestry. Thus, we have identified PRRT2 as the first causative gene of paroxysmal kinesigenic dyskinesias, warranting further investigations to understand the pathogenesis of this disorder.

Mutation analysis of Parkin, PINK1, DJ-1 and ATP13A2 genes in Chinese patients with autosomal recessive early-onset Parkinsonism

Autosomal recessive early‐onset Parkinsonism (AREP) has been associated with mutations in the Parkin, PINK1, DJ‐1, and ATP13A2 genes. We studied the prevalence of mutations in all four genes in 29 Chinese unrelated families with AREP using direct sequencing analysis and real‐time quantitative PCR analysis assay. There are 14 families (48.3%) with mutations of Parkin gene, 2 families (6.9%) with mutations of PINK1 gene, and 1 family (3.4%) with mutation of DJ‐1 gene. No pathogenic mutations in ATP13A2 gene were found in these families. Three Parkin gene mutations (c.G859T, c.1069‐1074delGTGTCC, and c.T1422C) and one DJ‐1 gene mutation (c.T29C) have not been reported previously. In conclusion, Parkin gene mutation is the most common pathogenic factor in Chinese patients with AREP. Mutations of DJ‐1 and PINK1 gene are also found in Chinese families with AREP. Mutations in ATP13A2 gene may be rare in Chinese families with AREP.

Hypomethylation of SNCA in blood of patients with sporadic Parkinson's disease

SNCA is a pathogenic gene identified in rare familial PD, and over-expression of SNCA was suggested in the pathogenesis of familial and sporadic PD. Rep1 polymorphism of SNCA was associated with susceptibility to sporadic PD and SNCA expression in intro and in vivo. Hypomethylation in SNCA intron-1 was associated with increased SNCA expression and was observed in postmortem brains of patients with sporadic PD. We studied the methylation status of SNCA intron-1, SNCA mRNA levels and Rep1 genotypes in PBMCs of 100 sporadic PD patients and 95 controls and explored the relationship between DNA methylation, mRNA expression and Rep1 genotypes. Hypomethylation of SNCA intron-1 was detected in PBMCs of PD patients, and DNA methylation levels were associated with Rep1 polymorphism. The shorter allele was associated with higher level of SNCA intron-1 methylation, and genotypes carrying the shorter allele showed significantly higher methylation level of SNCA intron-1 than genotypes carrying the longer allele. However, SNCA mRNA levels were not associated with disease status, Rep1 polymorphism or DNA methylation of SNCA intron-1 in our study.

Identification of CHIP as a Novel Causative Gene for Autosomal Recessive Cerebellar Ataxia

Autosomal recessive cerebellar ataxias are a group of neurodegenerative disorders that are characterized by complex clinical and genetic heterogeneity. Although more than 20 disease-causing genes have been identified, many patients are still currently without a molecular diagnosis. In a two-generation autosomal recessive cerebellar ataxia family, we mapped a linkage to a minimal candidate region on chromosome 16p13.3 flanked by single-nucleotide polymorphism markers rs11248850 and rs1218762. By combining the defined linkage region with the whole-exome sequencing results, we identified a homozygous mutation (c.493CT) in CHIP (NM_005861) in this family. Using Sanger sequencing, we also identified two compound heterozygous mutations (c.389AT/c.441GT; c.621C>G/c.707GC) in CHIP gene in two additional kindreds. These mutations co-segregated exactly with the disease in these families and were not observed in 500 control subjects with matched ancestry. CHIP colocalized with NR2A, a subunit of the N-methyl-D-aspartate receptor, in the cerebellum, pons, medulla oblongata, hippocampus and cerebral cortex. Wild-type, but not disease-associated mutant CHIPs promoted the degradation of NR2A, which may underlie the pathogenesis of ataxia. In conclusion, using a combination of whole-exome sequencing and linkage analysis, we identified CHIP, encoding a U-box containing ubiquitin E3 ligase, as a novel causative gene for autosomal recessive cerebellar ataxia.

Glucocerebrosidase gene L444P mutation is a risk factor for Parkinson's disease in Chinese population.

An association between mutations in the glucocerebrosidase (GBA) gene and Parkinsons disease (PD) has been reported in several populations. We searched for four common GBA mutations (L444P, F213I, R353W, and N370S) in 402 Chinese PD patients and 413 age‐ and sex‐matched controls. In the PD cohort, 11 patients were found carrying a heterozygous GBA mutation and all of them had the L444P mutation. Heterozygous GBA mutations were detected none in controls. The GBA gene L444P mutation was detected at a significantly higher frequency among PD patients (11/402 = 2.74%), when compared with the control group (0/413): P = 0.0007. To evaluate the possible role of the GBA gene L444P mutation in PD in Ashkenazi Jewish and non‐Jewish populations, we conducted a meta‐analysis on the topic. In the Chinese population, the GBA gene L444P mutation was detected at a significantly higher frequency among PD patients, when compared with the control group: Z = 3.83, P = 0.0001, OR = 8.42, confidence interval = 95%, 2.83–25.06. In the non‐Jewish populations, the difference was obviously significant: Z = 5.76, P < 0.00001, OR = 8.82, confidence interval = 95%, 4.21–18.48. The results suggest that the GBA gene L444P mutation appears to be a risk factor for PD in Chinese population.

Mutation analysis of Parkin , PINK1 and DJ - 1 genes in Chinese patients with sporadic early onset parkinsonism

Early onset parkinsonism (EOP) has been associated with mutations in the Parkin, PINK1, and DJ-1 genes. We studied the prevalence of mutations in all three genes in 127 unrelated Chinese patients with apparently sporadic EOP using direct sequencing analysis and real-time quantitative PCR analysis assay. There are 16 patients (12.6%) with mutations of Parkin gene, four patients (3.1%) with mutations of PINK1 gene, and three patients (2.4%) with mutation of DJ-1 gene. In conclusion, Parkin gene mutation is the most common pathogenic factor in Chinese patients with sporadic EOP. Mutations of DJ-1 and PINK1 gene are also found in Chinese patients with sporadic EOP.

Polygenic determinants of Parkinson's disease in a Chinese population.

It has been reported that some single-nucleotide polymorphisms (SNPs) are associated with the risk of Parkinsons disease (PD), but whether a combination of these SNPs would have a stronger association with PD than any individual SNP is unknown. Sixteen SNPs located in the 8 genes and/or loci (SNCA, LRRK2, MAPT, GBA, HLA-DR, BST1, PARK16, and PARK17) were analyzed in a Chinese cohort consisting of 1061 well-characterized PD patients and 1066 control subjects from Central South of Mainland China. We found that Rep1, rs356165, and rs11931074 in SNCA gene; G2385R in LRRK2 gene; rs4698412 in BST1 gene; rs1564282 in PARK17; and L444P in GBA gene were associated with PD with adjustment of sex and age (p < 0.05) in the analysis of 16 variants. PD risk increased when Rep1 and rs11931074, G2385R, rs1564282, rs4698412; rs11931074 and G2385R, rs1564282, rs4698412; G2385R and rs1564282, rs4698412; and rs1564282 and rs4698412 were combined for the association analysis. In addition, PD risk increased cumulatively with the increasing number of variants (odds ratio for carrying 3 variants, 3.494). In summary, we confirmed that Rep1, rs356165, and rs11931074 in SNCA gene, G2385R in LRRK2 gene, rs4698412 in BST1 gene, rs1564282 in PARK17, and L444P in GBA gene have an independent and combined significant association with PD. SNPs in these 4 genes have a cumulative effect with PD.

Screening for two SNPs of LINGO1 gene in patients with essential tremor or sporadic Parkinson's disease in Chinese population

Two markers rs9652490 and rs11856808 both located in intron 3 of the LINGO1 gene have been nominated recently to be associated with essential tremor (ET). Although ET and Parkinsons disease (PD) are considered as different entities, they have many overlapping clinical and pathological features. We aimed to evaluate the role of rs9652490 and rs11856808 in the development of ET and PD. To this point, we sequenced the region involving the two markers in 109 ET cases, 425 sporadic Parkinsons disease (SPD) cases and 430 controls in Chinese population. After stratification by age, the rs9652490G allele suggested protective role in the early onset PD (EOPD, age at onset < or =50 years) group compared with age matched controls (OR=0.56, 95% CI: 0.35-0.90, p=0.015). No other significant association was found. We concluded that the two markers rs9652490 and rs11856808 were not strongly related to the development of ET or late onset SPD, but the rs9652490G allele might be a protective factor for EOPD in Chinese population.

Analysis of SCA2 and SCA3/MJD repeats in Parkinson's disease in mainland China: genetic, clinical, and positron emission tomography findings.

To investigate the prevalence and clinical feature(s) of Parkinsons disease (PD) patients with expanded (ATXN2 and MJD1) genes of spinocerebellar ataxia type 2 and 3 (SCA2 and SCA3/MJD) in a mainland Chinese population, CAG triplet repeat expansions of (SCA2 and SCA3/MJD) genes (ATXN2 and MJD1) were analyzed in a cohort of 452 PD patients, including 386 sporadic and 66 familial forms. Striatal dopamine transporter was evaluated in two SCA2 and two SCA3/MJD‐positive family members, an idiopathic PD patient and a healthy control using carbon (C11) [11C]‐radiolabeled‐CFT positron emission tomography (PET). We found two patients in one familial PD (FPD) family (1.5%) and two sporadic PD patients (0.5%) with expanded CAG repeats in the ATXN2 locus, four patients in two FPD families (3%) and another three sporadic PD patients (0.8%) in the MJD1 locus. [11C]‐CFT PET in detected members in SCA2 and SCA3/MJD families showed decrements of 11C‐CFT uptake. These findings suggest that a mutation in SCA2 or SCA3/MJD may be one of the genetic causes of PD.

Investigation of TREM2, PLD3, and UNC5C variants in patients with Alzheimer's disease from mainland China.

Recently, 3 rare coding variants significantly associated with Alzheimers disease (AD) risk have been identified in western populations using whole exome sequencing method, including p.R47H in TREM2, p.V232M in PLD3, and p.T835M in UNC5C. To examine whether these variants are genetic risk factors in patients with AD from mainland China, we sequenced exon 2 of TREM2, exon 9 of PLD3, and exon 15 of UNC5C in Chinese Han population including 360 patients with AD and 400 control individuals. As a result, none of these 3 variants were identified in all subjects, however, 1 novel variant (p.A130V) in TREM2 and 4 novel variants (p.Q860H, p.T837K, p.S843G, and p.V836V) in UNC5C were detected in unrelated patients with late-onset AD. These findings suggest the 3 rare coding variants might not play an important role in AD risk in mainland China.