帕金森氏症(PD)為僅次於阿茲海默氏症常見的神經退化性疾病。臨床症狀包括四肢及軀體顫抖、行動緩慢等現象，主要的病理特徵是多巴胺神經元在基底核與其他腦幹神經核有退化死亡的現象，尤其以黑質之緻密區與路易氏體(Lewy body)存在的神經元最為嚴重。LRRK2基因為目前引起顯性遺傳帕金森氏症病患重要的基因。近年來研究發現LRRK2基因上的G2385R與R1628P變異為亞洲人種所特有的危險因子。本研究經定序PD患者的LRRK2 cDNA，找到一個已報導的R1441H、兩個未被報導過的R767H與S885N突變，及5個改變胺基酸、8個未改變胺基酸的多型性或變異。進一步針對R1398H、G2385R與M2397T多型性，進行病例-對照組研究，發現G2385R多型性異型合子在PD患者的頻率要高於正常人(8.4 vs. 4.2%, odds ratio = 2.08, 95% CI: 1.05-4.41, P = 0.044)，且M2397T多型性同基因型會加重G2385R異基因型對PD的感受性(4.2 vs. 0.4%, odds ratio = 10.63, 95% CI: 2.08-194.26, P = 0.024)。由於近年研究發現GRN基因突變與額顳葉型失智症(FTD)相關，本研究亦針對41位病人的GRN基因做定序分析，發現兩個未被報導的突變L57M、T487I及兩個已被報導的多型性D128、3’ UTR C>T。本研究結果可提供臨床上診斷及諮詢的參考。

Abstract
Parkinson’s disease (PD), the second most common neurodegenerative disorder, is characterized by resting tremor, rigidity, bradykinesia, and postural instability. The symptoms of PD in pathology are neuronal loss in the basal ganglia, especially in the substantia nigra pars compacta, and presence of intracellular Lewy body inclusions in surviving neurons. Mutations in leucine-rich repeat kinase 2 (LRRK2, encoding dardarin) are the most frequent genetic cause of PD known nowadays. Mutations in different functional regions of LRRK2 have been reported. Studies from China, Singapore, and Japan indicate that the G2385R polymorphism is an ancient and common risk factor for sporadic PD in the Asian population. In this study, we screened LRRK2 mutations in PD patients and identified a reported (R1441H) and two novel (R767H and S885N) mutations, in addition to 5 nonsynonymous and 8 synonymous amino acid substitutions. In the case-control study, the frequency of the heterozygous G2385R genotype was higher in PD compared to controls (8.4 vs. 4.2%, odds ratio = 2.08, 95% CI: 1.05-4.41, P = 0.044). Moreover, M2397T acting synergistically with G2385R to play role in PD susceptibility (4.2 vs. 0.4%, odds ratio = 10.63, 95% CI: 2.08-194.26, P = 0.024). Granulin (GRN) has been identified as one of the gene responsible for frontotemporal dementia (FTD) and mutations in GRN have been reported. Using cDNA sequencing, we found two novel mutations L57M and T487I, additionally to a synonymous D128 and a C>T variation in 3’UTR. The studies may provide a tool for clinical diagnosis and counseling.

An, X.K., Peng, R., Li, T., Burgunder, J.M., Wu, Y., Chen, W.J., Zhang, J.H., Wang, Y.C., Xu, Y.M., Gou, Y.R., et al. (2008). LRRK2 Gly2385Arg variant is a risk factor of Parkinson's disease among Han-Chinese from mainland China. Eur J Neurol 15, 301-305.
Baker, M., Mackenzie, I.R., Pickering-Brown, S.M., Gass, J., Rademakers, R., Lindholm, C., Snowden, J., Adamson, J., Sadovnick, A.D., Rollinson, S., et al. (2006). Mutations in progranulin cause tau-negative frontotemporal dementia linked to chromosome 17. Nature 442, 916-919.
Belin, A.C., and Westerlund, M. (2008). Parkinson's disease: a genetic perspective. FEBS J 275, 1377-1383.
Berg, D., Schweitzer, K., Leitner, P., Zimprich, A., Lichtner, P., Belcredi, P., Brussel, T., Schulte, C., Maass, S., and Nagele, T. (2005). Type and frequency of mutations in the LRRK2 gene in familial and sporadic Parkinson's disease*. Brain 128, 3000-3011.
Bertram, C.P., Lemay, M., and Stelmach, G.E. (2005). The effect of Parkinson's disease on the control of multi-segmental coordination. Brain Cogn 57, 16-20.
Bhandari, V., Palfree, R.G., and Bateman, A. (1992). Isolation and sequence of the granulin precursor cDNA from human bone marrow reveals tandem cysteine-rich granulin domains. Proc Natl Acad Sci USA 89, 1715-1719.
Biskup, S., Moore, D.J., Celsi, F., Higashi, S., West, A.B., Andrabi, S.A., Kurkinen, K., Yu, S.W., Savitt, J.M., Waldvogel, H.J., et al. (2006). Localization of LRRK2 to membranous and vesicular structures in mammalian brain. Ann Neurol 60, 557-569.
Bonifati, V., Rizzu, P., Squitieri, F., Krieger, E., Vanacore, N., van Swieten, J.C., Brice, A., van Duijn, C.M., Oostra, B., Meco, G., et al. (2003). DJ-1( PARK7), a novel gene for autosomal recessive, early onset parkinsonism. Neurol Sci 24, 159-160.
Braak, H., Del Tredici, K., Rub, U., de Vos, R.A., Jansen Steur, E.N., and Braak, E. (2003). Staging of brain pathology related to sporadic Parkinson's disease. Neurobiol Aging 24, 197-211.
Bruni, A.C., Momeni, P., Bernardi, L., Tomaino, C., Frangipane, F., Elder, J., Kawarai, T., Sato, C., Pradella, S., Wakutani, Y., et al. (2007). Heterogeneity within a large kindred with frontotemporal dementia: a novel progranulin mutation. Neurology 69, 140-147.
Chen, R.C., Chang, S.F., Su, C.L., Chen, T.H., Yen, M.F., Wu, H.M., Chen, Z.Y., and Liou, H.H. (2001). Prevalence, incidence, and mortality of PD: a door-to-door survey in Ilan county, Taiwan. Neurology 57, 1679-1686.
Cheverud, J.M. (2001). A simple correction for multiple comparisons in interval mapping genome scans. Heredity 87, 52-58.
Conley, S.C., and Kirchner, J.T. (1999). Parkinson's disease--the shaking palsy. Underlying factors, diagnostic considerations, and clinical course. Postgrad Med 106, 39-42, 45-36, 49-50 passim.
Cruts, M., Gijselinck, I., van der Zee, J., Engelborghs, S., Wils, H., Pirici, D., Rademakers, R., Vandenberghe, R., Dermaut, B., Martin, J.J., et al. (2006). Null mutations in progranulin cause ubiquitin-positive frontotemporal dementia linked to chromosome 17q21. Nature 442, 920-924.
Daniel, R., He, Z., Carmichael, K.P., Halper, J., and Bateman, A. (2000). Cellular localization of gene expression for progranulin. J Histochem Cytochem 48, 999-1009.
Daniel, R., Daniels, E., He, Z., and Bateman, A. (2003). Progranulin (acrogranin/PC cell-derived growth factor/granulin-epithelin precursor) is expressed in the placenta, epidermis, microvasculature, and brain during murine development. Dev Dyn 227, 593-599.
Dauer, W., and Przedborski, S. (2003). Parkinson's disease: mechanisms and models. Neuron 39, 889-909.
Deng, H., Le, W., Huang, M., Xie, W., Pan, T., and Jankovic, J. (2007). Genetic analysis of LRRK2 P755L variant in Caucasian patients with Parkinson's disease. Neurosci Lett 419, 104-107.
Deng, J., Lewis, P.A., Greggio, E., Sluch, E., Beilina, A., and Cookson, M.R. (2008). Structure of the ROC domain from the Parkinson's disease-associated leucine-rich repeat kinase 2 reveals a dimeric GTPase. Proc Natl Acad Sci USA 105, 1499-1504.
Di Fonzo, A., Rohe, C.F., Ferreira, J., Chien, H.F., Vacca, L., Stocchi, F., Guedes, L., Fabrizio, E., Manfredi, M., Vanacore, N., et al. (2005). A frequent LRRK2 gene mutation associated with autosomal dominant Parkinson's disease. Lancet 365, 412-415.
Di Fonzo, A., Tassorelli, C., De Mari, M., Chien, H.F., Ferreira, J., Rohe, C.F., Riboldazzi, G., Antonini, A., Albani, G., Mauro, A., et al. (2006a). Comprehensive analysis of the LRRK2 gene in sixty families with Parkinson's disease. Eur J Hum Genet 14, 322-331.
Di Fonzo, A., Wu-Chou, Y.H., Lu, C.S., van Doeselaar, M., Simons, E.J., Rohe, C.F., Chang, H.C., Chen, R.S., Weng, Y.H., Vanacore, N., et al. (2006b). A common missense variant in the LRRK2 gene, Gly2385Arg, associated with Parkinson's disease risk in Taiwan. Neurogenetics 7, 133-138.
Eriksen, J.L., and Mackenzie, I.R. (2008). Progranulin: normal function and role in neurodegeneration. J Neurochem 104, 287-297.
Farrer, M.J. (2006). Genetics of Parkinson disease: paradigm shifts and future prospects. Nat Rev Genet 7, 306-318.
Farrer, M.J., Stone, J.T., Lin, C.H., Dachsel, J.C., Hulihan, M.M., Haugarvoll, K., Ross, O.A., and Wu, R.M. (2007). Lrrk2 G2385R is an ancestral risk factor for Parkinson's disease in Asia. Parkinsonism Relat Disord 13, 89-92.
Foster, N.L., Wilhelmsen, K., Sima, A.A., Jones, M.Z., D'Amato, C.J., and Gilman, S. (1997). Frontotemporal dementia and parkinsonism linked to chromosome 17: a consensus conference. Conference Participants. Ann Neurol 41, 706-715.
Funayama, M., Hasegawa, K., Kowa, H., Saito, M., Tsuji, S., and Obata, F. (2002). A new locus for Parkinson's disease (PARK8) maps to chromosome 12p11.2-q13.1. Ann Neurol 51, 296-301.
Funayama, M., Li, Y., Tomiyama, H., Yoshino, H., Imamichi, Y., Yamamoto, M., Murata, M., Toda, T., Mizuno, Y., and Hattori, N. (2007). Leucine-rich repeat kinase 2 G2385R variant is a risk factor for Parkinson disease in Asian population. Neuroreport 18, 273-275.
Gass, J., Cannon, A., Mackenzie, I.R., Boeve, B., Baker, M., Adamson, J., Crook, R., Melquist, S., Kuntz, K., Petersen, R., et al. (2006). Mutations in progranulin are a major cause of ubiquitin-positive frontotemporal lobar degeneration. Hum Mol Genet 15, 2988-3001.
Giasson, B.I., and Van Deerlin, V.M. (2008). Mutations in LRRK2 as a cause of Parkinson's disease. Neurosignals 16, 99-105.
Gibb, W.R., and Lees, A.J. (1991). Anatomy, pigmentation, ventral and dorsal subpopulations of the substantia nigra, and differential cell death in Parkinson's disease. J Neurol Neurosurg Psychiatry 54, 388-396.
Giulian, D., and Lachman, L.B. (1985). Interleukin-1 stimulation of astroglial proliferation after brain injury. Science 228, 497-499.
Goedert, M., Jakes, R., Spillantini, M.G., Hasegawa, M., Smith, M.J., and Crowther, R.A. (1996). Assembly of microtubule-associated protein tau into Alzheimer-like filaments induced by sulphated glycosaminoglycans. Nature 383, 550-553.
Greggio, E., Zambrano, I., Kaganovich, A., Beilina, A., Taymans, J.M., Daniels, V., Lewis, P., Jain, S., Ding, J., Syed, A., et al. (2008). The Parkinson disease-associated leucine-rich repeat kinase 2 (LRRK2) is a dimer that undergoes intramolecular autophosphorylation. J Biol Chem 283, 16906-16914.
Hatano, Y., Sato, K., Elibol, B., Yoshino, H., Yamamura, Y., Bonifati, V., Shinotoh, H., Asahina, M., Kobayashi, S., Ng, A.R., et al. (2004). PARK6-linked autosomal recessive early-onset parkinsonism in Asian populations. Neurology 63, 1482-1485.
He, Z., and Bateman, A. (2003). Progranulin (granulin-epithelin precursor, PC-cell-derived growth factor, acrogranin) mediates tissue repair and tumorigenesis. J Mol Med 81, 600-612.
He, Z., Ong, C.H., Halper, J., and Bateman, A. (2003). Progranulin is a mediator of the wound response. Nature medicine 9, 225-229.
Higashi, S., Biskup, S., West, A.B., Trinkaus, D., Dawson, V.L., Faull, R.L., Waldvogel, H.J., Arai, H., Dawson, T.M., Moore, D.J., et al. (2007a). Localization of Parkinson's disease-associated LRRK2 in normal and pathological human brain. Brain Res 1155, 208-219.
Higashi, S., Moore, D.J., Colebrooke, R.E., Biskup, S., Dawson, V.L., Arai, H., Dawson, T.M., and Emson, P.C. (2007b). Expression and localization of Parkinson's disease-associated leucine-rich repeat kinase 2 in the mouse brain. J Neurochem 100, 368-381.
Hoehn, M.M., and Yahr, M.D. (1967). Parkinsonism: onset, progression and mortality. Neurology 17, 427-442.
Huang, Y., Halliday, G.M., Vandebona, H., Mellick, G.D., Mastaglia, F., Stevens, J., Kwok, J., Garlepp, M., Silburn, P.A., Horne, M.K., et al. (2007). Prevalence and clinical features of common LRRK2 mutations in Australians with Parkinson's disease. Mov Disord 22, 982-989.
Johnston, C., Jiang, W., Chu, T., and Levine, B. (2001). Identification of genes involved in the host response to neurovirulent alphavirus infection. J Virol 75, 10431-10445.
Joutel, A., Corpechot, C., Ducros, A., Vahedi, K., Chabriat, H., Mouton, P., Alamowitch, S., Domenga, V., Cecillion, M., Marechal, E., et al. (1996). Notch3 mutations in CADASIL, a hereditary adult-onset condition causing stroke and dementia. Nature 383, 707-710.
Kitada, T., Asakawa, S., Hattori, N., Matsumine, H., Yamamura, Y., Minoshima, S., Yokochi, M., Mizuno, Y., and Shimizu, N. (1998). Mutations in the parkin gene cause autosomal recessive juvenile parkinsonism. Nature 392, 605-608.
Le Ber, I., van der Zee, J., Hannequin, D., Gijselinck, I., Campion, D., Puel, M., Laquerriere, A., De Pooter, T., Camuzat, A., Van den Broeck, M., et al. (2007). Progranulin null mutations in both sporadic and familial frontotemporal dementia. Hum Mutat 28, 846-855.
Leroy, E., Boyer, R., Auburger, G., Leube, B., Ulm, G., Mezey, E., Harta, G., Brownstein, M.J., Jonnalagada, S., Chernova, T., Dehejia, A., Lavedan, C., Gasser, T., Steinbach, P.J., Wilkinson, K.D., Polymeropoulos, M.H. (1998). The ubiquitin pathway in Parkinson's disease. Nature 395, 451-452.
Lesage, S., Durr, A., Tazir, M., Lohmann, E., Leutenegger, A.L., Janin, S., Pollak, P., and Brice, A. (2006). LRRK2 G2019S as a cause of Parkinson's disease in North African Arabs. N Engl J Med 354, 422-423.
Li, C., Ting, Z., Qin, X., Ying, W., Li, B., Guo Qiang, L., Jian Fang, M., Jing, Z., Jian Qing, D., and Sheng Di, C. (2007a). The prevalence of LRRK2 Gly2385Arg variant in Chinese Han population with Parkinson's disease. Mov Disord 22, 2439-2443.
Li, X., Tan, Y.C., Poulose, S., Olanow, C.W., Huang, X.Y., and Yue, Z. (2007b). Leucine-rich repeat kinase 2 (LRRK2)/PARK8 possesses GTPase activity that is altered in familial Parkinson's disease R1441C/G mutants. J Neurochem 103, 238-247.
Lin, C.H., Tzen, K.Y., Yu, C.Y., Tai, C.H., Farrer, M.J., and Wu, R.M. (2008). LRRK2 mutation in familial Parkinson's disease in a Taiwanese population: clinical, PET, and functional studies. J Biomed Sci.
Mackenzie, I.R., Baker, M., Pickering-Brown, S., Hsiung, G.Y., Lindholm, C., Dwosh, E., Gass, J., Cannon, A., Rademakers, R., Hutton, M., et al. (2006). The neuropathology of frontotemporal lobar degeneration caused by mutations in the progranulin gene. Brain 129, 3081-3090.
Mandelkow, E.M., Biernat, J., Drewes, G., Gustke, N., Trinczek, B., and Mandelkow, E. (1995). Tau domains, phosphorylation, and interactions with microtubules. Neurobiol Aging 16, 355-362, discussion 362-353.
Marjama-Lyons, J.M., and Koller, W.C. (2001). Parkinson's disease. Update in diagnosis and symptom management. Geriatrics 56, 24-25, 29-30, 33-25.
Mata, I.F., Kachergus, J.M., Taylor, J.P., Lincoln, S., Aasly, J., Lynch, T., Hulihan, M.M., Cobb, S.A., Wu, R.M., Lu, C.S., et al. (2005). Lrrk2 pathogenic substitutions in Parkinson's disease. Neurogenetics 6, 171-177.
Mata, I.F., Wedemeyer, W.J., Farrer, M.J., Taylor, J.P., and Gallo, K.A. (2006). LRRK2 in Parkinson's disease: protein domains and functional insights. Trends Neurosci 29, 286-293.
Melrose, H., Lincoln, S., Tyndall, G., Dickson, D., and Farrer, M. (2006). Anatomical localization of leucine-rich repeat kinase 2 in mouse brain. Neuroscience 139, 791-794.
Nyholt, D.R. (2004). A simple correction for multiple testing for single-nucleotide polymorphisms in linkage disequilibrium with each other. Am J Hum Genet 74, 765-769.
Ohmi, K., Greenberg, D.S., Rajavel, K.S., Ryazantsev, S., Li, H.H., and Neufeld, E.F. (2003). Activated microglia in cortex of mouse models of mucopolysaccharidoses I and IIIB. Proc Natl Acad Sci USA 100, 1902-1907.
Ozelius, L.J., Senthil, G., Saunders-Pullman, R., Ohmann, E., Deligtisch, A., Tagliati, M., Hunt, A.L., Klein, C., Henick, B., Hailpern, S.M., et al. (2006). LRRK2 G2019S as a cause of Parkinson's disease in Ashkenazi Jews. N Engl J Med 354, 424-425.
Paisan-Ruiz, C., Jain, S., Evans, E.W., Gilks, W.P., Simon, J., van der Brug, M., Lopez de Munain, A., Aparicio, S., Gil, A.M., Khan, N., et al. (2004). Cloning of the gene containing mutations that cause PARK8-linked Parkinson's disease. Neuron 44, 595-600.
Paisan-Ruiz, C., Lang, A.E., Kawarai, T., Sato, C., Salehi-Rad, S., Fisman, G.K., Al-Khairallah, T., St George-Hyslop, P., Singleton, A., and Rogaeva, E. (2005). LRRK2 gene in Parkinson disease: mutation analysis and case control association study. Neurology 65, 696-700.
Polymeropoulos, M.H., Lavedan, C., Leroy, E., Ide, S.E., Dehejia, A., Dutra, A., Pike, B., Root, H., Rubenstein, J., Boyer, R., et al. (1997). Mutation in the alpha-synuclein gene identified in families with Parkinson's disease. Science 276, 2045-2047.
Ross, O.A., Wu, Y.R., Lee, M.C., Funayama, M., Chen, M.L., Soto, A.I., Mata, I.F., Lee-Chen, G.J., Chen, C.M., Tang, M., et al. (2008). Analysis of Lrrk2 R1628P as a risk factor for Parkinson's disease. Ann Neurol.
Ruas, M., Brookes, S., McDonald, N.Q., and Peters, G. (1999). Functional evaluation of tumour-specific variants of p16INK4a/CDKN2A: correlation with protein structure information. Oncogene 18, 5423-5434.
Russo, A.A., Tong, L., Lee, J.O., Jeffrey, P.D., and Pavletich, N.P. (1998). Structural basis for inhibition of the cyclin-dependent kinase Cdk6 by the tumour suppressor p16INK4a. Nature 395, 237-243.
Schapira, A.H. (2006). Etiology of Parkinson's disease. Neurology 66, S10-23.
Schultheis, P.J., Hagen, T.T., O'Toole, K.K., Tachibana, A., Burke, C.R., McGill, D.L., Okunade, G.W., and Shull, G.E. (2004). Characterization of the P5 subfamily of P-type transport ATPases in mice. Biochem Biophys Res Commun 323, 731-738.
Spanaki, C., Latsoudis, H., and Plaitakis, A. (2006). LRRK2 mutations on Crete: R1441H associated with PD evolving to PSP. Neurology 67, 1518-1519.
Spillantini, M.G., Schmidt, M.L., Lee, V.M., Trojanowski, J.Q., Jakes, R., and Goedert, M. (1997). Alpha-synuclein in Lewy bodies. Nature 388, 839-840.
Stephens, M., Smith, N.J., Donnelly, P. (2001). A new statistical method for haplotype reconstruction from population data. Am J Hum Genet 68, 978-989.
Tan, E.K., Shen, H., Tan, L.C., Farrer, M., Yew, K., Chua, E., Jamora, R.D., Puvan, K., Puong, K.Y., Zhao, Y., et al. (2005). The G2019S LRRK2 mutation is uncommon in an Asian cohort of Parkinson's disease patients. Neurosci Lett 384, 327-329.
Tan, E.K., Zhao, Y., Skipper, L., Tan, M.G., Di Fonzo, A., Sun, L., Fook-Chong, S., Tang, S., Chua, E., Yuen, Y., et al. (2007). The LRRK2 Gly2385Arg variant is associated with Parkinson's disease: genetic and functional evidence. Hum Genet 120, 857-863.
Tan, E.K., Lim, H.Q., Yuen, Y., and Zhao, Y. (2008). Pathogenicity of LRRK2 P755L variant in Parkinson's disease. Mov Disord 23, 734-736.
Taylor, J.P., Mata, I.F., and Farrer, M.J. (2006). LRRK2: a common pathway for parkinsonism, pathogenesis and prevention? Trends Mol Med 12, 76-82.
Thery, C., and Mallat, M. (1993). Influence of interleukin-1 and tumor necrosis factor alpha on the growth of microglial cells in primary cultures of mouse cerebral cortex: involvement of colony-stimulating factor 1. Neurosci Lett 150, 195-199.
Weiss, B. (2008). ROCO kinase activity is controlled by internal GTPase function. Sci Signal 1, pe27.
West, A.B., Moore, D.J., Biskup, S., Bugayenko, A., Smith, W.W., Ross, C.A., Dawson, V.L., and Dawson, T.M. (2005). Parkinson's disease-associated mutations in leucine-rich repeat kinase 2 augment kinase activity. Proc Natl Acad Sci U S A 102, 16842-16847.
Wood-Kaczmar, A., Gandhi, S., and Wood, N.W. (2006). Understanding the molecular causes of Parkinson's disease. Trends Mol Med 12, 521-528.
Wu, T., Zeng, Y., Ding, X., Li, X., Li, W., Dong, H., Chen, S., Zhang, X., Ma, G., Yao, J., et al. (2006). A novel P755L mutation in LRRK2 gene associated with Parkinson's disease. Neuroreport 17, 1859-1862.
Zabetian, C.P., Samii, A., Mosley, A.D., Roberts, J.W., Leis, B.C., Yearout, D., Raskind, W.H., and Griffith, A. (2005). A clinic-based study of the LRRK2 gene in Parkinson disease yields new mutations. Neurology 65, 741-744.
Zhu, J., Nathan, C., Jin, W., Sim, D., Ashcroft, G.S., Wahl, S.M., Lacomis, L., Erdjument-Bromage, H., Tempst, P., Wright, C.D., et al. (2002). Conversion of proepithelin to epithelins: roles of SLPI and elastase in host defense and wound repair. Cell 111, 867-878.
Zimprich, A., Biskup, S., Leitner, P., Lichtner, P., Farrer, M., Lincoln, S., Kachergus, J., Hulihan, M., Uitti, R.J., Calne, D.B., et al. (2004). Mutations in LRRK2 cause autosomal-dominant parkinsonism with pleomorphic pathology. Neuron 44, 601-607.