PARK2與PARK13分別與體染色體隱性遺傳及偶發性帕金森氏症(Parkinson's disease；簡稱PD)相關。PARK2基因產物Parkin具ubiquitin E3 ligase功能，而PARK13基因產物HTRA2則與細胞凋亡相關。本研究延續先前的PARK2 cDNA定序(管, 2007)，分析台灣帕金森氏症患者Parkin的基因變異。結果共發現二個缺失突變(Ex2-3del、Ex5del)、一個點突變(R334C)、兩個多型性(S167N、V380L)及一個新穎的內含子9插入突變(c.1084intron+)，此插入突變與內含子9上的-6g>a (IVS9 g>a)的多型性點相關。進一步對所蒐集的病人與性別、年齡相當的正常人進行S167N、V380L的病例-對照組分析，結果顯示V380L C等位基因頻率在病人族群中明顯較正常人族群低，且和低帕金森氏症感受性相關。Ex5del缺失突變與正常人淋巴細胞的分析結果顯示，病人淋巴細胞的Caspase-3活性顯著高於正常人，粒線體膜電位分析結果亦顯示病人淋巴細胞的細胞凋亡顯著高於正常人，在staurosporine或去血清處理下，病人淋巴細胞存活率顯著低於正常人。在HTRA2基因分析方面，HTRA2 cDNA定序結果發現二個新穎的點突變(R36W與T215M)。進一步建構EGFP標記的cDNA質體，表現於HEK-293T及SK-N-SH細胞，次細胞分層、西方轉漬、螢光顯微鏡分析結果顯示，R36W與T215M蛋白皆有成熟型蛋白生成且座落於粒線體，但R36W蛋白成熟型表現量較野生型低，T215M蛋白則表現異常分子量的前軀及成熟蛋白。

PARK2 and PARK13 are involved in autosomal recessive juvenile parkinsonism and sporadic Parkinson’s disease (PD), respectively. The PARK2 gene product Parkin has ubiquitin E3 ligase activity, whereas PARK13 gene product HTRA2 is located in the mitochondrial intermembrane space and released into the cytosol during apoptosis. In the present study PARK2 mutations were analyzed in a cohort of Taiwanese PD patients by using direct cDNA sequencing. Two deletions (Ex2-3del and Ex5del), one point mutation (R334C), one insertion (c.1084intron+) and two reported SNPs (S167N and V380L) in Parkin were identified. The c.1084intron+ was due to a novel g-a transition SNP at position -6 of a cryptic splice acceptor site within IVS9 (-6g>a). The association of Parkin polymorphisms S167N and V380L with PD were analyzed using a case-control study. Although the difference is not significant, the V380L C allele was notably lower in PD patients than the controls, and a trend toward decrease in risk of developing PD was evident. In lymphoblastoid cells, caspase-3 activity and loss of mitochondrial membrane potential in cells with Ex5del were significantly higher than that of the control cells. Treatment of staurosporine significantly increases cell death in the cells with e Ex5del. Screening of the HTRA2 cDNA revealed two novel point mutations (R36W and T215M). The EGFP-tagged HTRA2 constructs were prepared for transient expression in HEK-293T and SK-N-SH cells. Subcellular fractionation, Western blot and fluorescence microscopy examination revealed that both R36W and T215M mature proteins localized to mitochondria. However, the amount of R36W mature protein is less than that of wild type. Also unusual precursor and mature proteins were observed with T215M mutation.

管郡潔(2007)。Parkin、DJ-1基因突變及粒線體DNA多型性與台灣帕金森氏症的分子遺傳研究。國立台灣師範大學生命科學系九十五學年度碩士論文。
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