脊髓小腦運動失調症(spinocerebellar ataxia；SCA)，為一群異質性的退化性神經疾病，大部分起因於三核重複擴增，患者在小腦、腦幹、脊髓及周邊神經系統等出現漸進式的退化。臨床上SCA不易分類，在多種相關的致病基因被分離後，分子檢測的建立及致病機轉的研究，有助於我們對這群疾病的了解。SCA8和致病基因3'端非轉譯區的CTG三核擴增相關，除家族性及偶發性的運動失調患者外，在阿茲海默氏症(AD)、精神疾病症、帕金森氏症(PD)及其他類型之SCA患者中，甚至於極少數的正常人中，亦可見到異常的CTG三核擴增(68 ~ 800個)。本研究首先建立台灣正常人及上述退化性神經疾病患者，其SCA8基因CTG重複之遺傳資料庫，結果發現有5個擴增的allele，分別來自1個AD患者、3個PD患者與1個SCA3患者。擴增之allele經定序分析顯示包含63 ~ 93個三核重複。另一方面也構築含有不同CTG重複(0, 23, 88, 157個)的SCA8 cDNA於表現載體上，轉移至人類胚胎腎細胞(HEK-293)或老鼠胚胎癌細胞(P19)，來研究SCA8可能之致病機轉。結果發現包含0個CTG重複之SCA8基因表現最好，當CTG重複23-157次時，SCA8基因的表現量下降，但下降程度不隨CTG擴增而有明顯差異，且RT-PCR分析顯示SCA8 mRNA在細胞核、質的分布情形與穩定性均不受CTG重複次數影響。和第一型肌強直萎縮症(myotonic dystrophy type 1；DM1)一樣，擴增之SCA8 RNA (CTG重複88或157個)在核內會形成RNA foci。而在SCA8和TBP cDNA或(CAG)36之共轉實驗中，也發現SCA8對TBP RNA或(CAG)36 RNA有類似antisense RNA之作用。上述結果可幫助我們初步了解SCA8表現時其RNA可能造成之效應。

Alwazzan, M., Newman, E., Hamshere, M. G., and Brook, J. D. (1999). Myotonic dystrophy is associated with a reduced level of RNA from the DMWD allele adjacent to the expanded repeat. Hum Mol Genet 8, 1491-1497.

Amack, J. D., Paguio, A. P., and Mahadevan, M. S. (1999). Cis and trans effects of the myotonic dystrophy (DM) mutation in a cell culture model. Hum Mol Genet 8, 1975-1984.

Amack, J. D., and Mahadevan, M. S. (2001). The myotonic dystrophy expanded CUG repeat tract is necessary but not sufficient to disrupt C2C12 myoblast differentiation. Hum Mol Genet 10, 1879-1887.

Brook, J. D., McCurrach, M. E., Harley, H. G., Buckler, A. J., Church, D., Aburatani, H., Hunter, K., Stanton, V. P., Thirion, J. P., Hudson, T., and et al. (1992). Molecular basis of myotonic dystrophy: expansion of a trinucleotide (CTG) repeat at the 3' end of a transcript encoding a protein kinase family member. Cell 68, 799-808.

Boucher, C. A., King, S. K., Carey, N., Krahe, R., Winchester, C. L., Rahman, S., Creavin, T., Meghji, P., Bailey, M. E., Chartier, F. L., and et al. (1995). A novel homeodomain-encoding gene is associated with a large CpG island interrupted by the myotonic dystrophy unstable (CTG)n repeat. Hum Mol Genet 4, 1919-1925.

Benzow, K. A., and Koob, M. D. (2002). The KLHL1-antisense transcript (KLHL1AS) is evolutionarily conserved. Mamm Genome 13, 134-141.

Bryer, A., Krause, A., Bill, P., Davids, V., Bryant, D., Butler, J., Heckmann, J., Ramesar, R., and Greenberg, J. (2003). The hereditary adult-onset ataxias in South Africa. J Neurol Sci 216, 47-54.

Brusco, A., Gellera, C., Cagnoli, C., Saluto, A., Castucci, A., Michielotto, C., Fetoni, V., Mariotti, C., Migone, N., Di Donato, S., and Taroni, F. (2004). Molecular genetics of hereditary spinocerebellar ataxia: mutation analysis of spinocerebellar ataxia genes and CAG/CTG repeat expansion detection in 225 Italian families. Arch Neurol 61, 727-733.

Bowman, A. B., Yoo, S. Y., Dantuma, N. P., and Zoghbi, H. Y. (2005). Neuronal dysfunction in a polyglutamine disease model occurs in the absence of ubiquitin-proteasome system impairment and inversely correlates with the degree of nuclear inclusion formation. Hum Mol Genet 14, 679-691.

Cellini, E., Nacmias, B., Forleo, P., Piacentini, S., Guarnieri, B. M., Serio, A., Calabro, A., Renzi, D., and Sorbi, S. (2001). Genetic and clinical analysis of spinocerebellar ataxia type 8 repeat expansion in Italy. Arch Neurol 58, 1856-1859.

Davis, B. M., McCurrach, M. E., Taneja, K. L., Singer, R. H., and Housman, D. E. (1997). Expansion of a CUG trinucleotide repeat in the 3' untranslated region of myotonic dystrophy protein kinase transcripts results in nuclear retention of transcripts. Proc Natl Acad Sci U S A 94, 7388-7393.

Day, J. W., Schut, L. J., Moseley, M. L., Durand, A. C., and Ranum, L. P. (2000). Spinocerebellar ataxia type 8: clinical features in a large family. Neurology 55, 649-657.

Dorsman, J. C., Pepers, B., Langenberg, D., Kerkdijk, H., Ijszenga, M., den Dunnen, J. T., Roos, R. A., and van Ommen, G. J. (2002). Strong aggregation and increased toxicity of polyleucine over polyglutamine stretches in mammalian cells. Hum Mol Genet 11, 1487-1496.

Fu, Y. H., Pizzuti, A., Fenwick, R. G., Jr., King, J., Rajnarayan, S., Dunne, P. W., Dubel, J., Nasser, G. A., Ashizawa, T., de Jong, P., and et al. (1992). An unstable triplet repeat in a gene related to myotonic muscular dystrophy. Science 255, 1256-1258.

Furling, D., Doucet, G., Langlois, M. A., Timchenko, L., Belanger, E., Cossette, L., and Puymirat, J. (2003). Viral vector producing antisense RNA restores myotonic dystrophy myoblast functions. Gene Ther 10, 795-802.

Harris, S., Moncrieff, C., and Johnson, K. (1996). Myotonic dystrophy: will the real gene please step forward! Hum Mol Genet 5 Spec No, 1417-1423.

Hashem, V. I., Pytlos, M. J., Klysik, E. A., Tsuji, K., Khajavi, M., Ashizawa, T., and Sinden, R. R. (2004). Chemotherapeutic deletion of CTG repeats in lymphoblast cells from DM1 patients. Nucleic Acids Res 32, 6334-6346.

Ikeda, Y., Shizuka-Ikeda, M., Watanabe, M., Schmitt, M., Okamoto, K., and Shoji, M. (2000a). Asymptomatic CTG expansion at the SCA8 locus is associated with cerebellar atrophy on MRI. J Neurol Sci 182, 76-79.

Ikeda, Y., Shizuka, M., Watanabe, M., Okamoto, K., and Shoji, M. (2000b). Molecular and clinical analyses of spinocerebellar ataxia type 8 in Japan. Neurology 54, 950-955.

Izumi, Y., Maruyama, H., Oda, M., Morino, H., Okada, T., Ito, H., Sasaki, I., Tanaka, H., Komure, O., Udaka, F., et al. (2003). SCA8 repeat expansion: large CTA/CTG repeat alleles are more common in ataxic patients, including those with SCA6. Am J Hum Genet 72, 704-709.

Jansen, G., Mahadevan, M., Amemiya, C., Wormskamp, N., Segers, B., Hendriks, W., O'Hoy, K., Baird, S., Sabourin, L., Lennon, G., and et al. (1992). Characterization of the myotonic dystrophy region predicts multiple protein isoform-encoding mRNAs. Nat Genet 1, 261-266.

Jansen, G., Bachner, D., Coerwinkel, M., Wormskamp, N., Hameister, H., and Wieringa, B. (1995). Structural organization and developmental expression pattern of the mouse WD-repeat gene DMR-N9 immediately upstream of the myotonic dystrophy locus. Hum Mol Genet 4, 843-852.

Juvonen, V., Hietala, M., Paivarinta, M., Rantamaki, M., Hakamies, L., Kaakkola, S., Vierimaa, O., Penttinen, M., and Savontaus, M. L. (2000). Clinical and genetic findings in Finnish ataxia patients with the spinocerebellar ataxia 8 repeat expansion. Ann Neurol 48, 354-361.

Juvonen, V., Kairisto, V., Hietala, M., and Savontaus, M. L. (2002). Calculating predictive values for the large repeat alleles at the SCA8 locus in patients with ataxia. J Med Genet 39, 935-936.

Klesert, T. R., Otten, A. D., Bird, T. D., and Tapscott, S. J. (1997). Trinucleotide repeat expansion at the myotonic dystrophy locus reduces expression of DMAHP. Nat Genet 16, 402-406.

Koch, K. S., and Leffert, H. L. (1998). Giant hairpins formed by CUG repeats in myotonic dystrophy messenger RNAs might sterically block RNA export through nuclear pores. J Theor Biol 192, 505-514.

Koob, M. D., Moseley, M. L., Schut, L. J., Benzow, K. A., Bird, T. D., Day, J. W., and Ranum, L. P. (1999). An untranslated CTG expansion causes a novel form of spinocerebellar ataxia (SCA8). Nat Genet 21, 379-384.
Korade-Mirnics, Z., Tarleton, J., Servidei, S., Casey, R. R., Gennarelli, M., Pegoraro, E., Angelini, C., and Hoffman, E. P. (1999). Myotonic dystrophy: tissue-specific effect of somatic CTG expansions on allele-specific DMAHP/SIX5 expression. Hum Mol Genet 8, 1017-1023.

Klesert, T. R., Cho, D. H., Clark, J. I., Maylie, J., Adelman, J., Snider, L., Yuen, E. C., Soriano, P., and Tapscott, S. J. (2000). Mice deficient in Six5 develop cataracts: implications for myotonic dystrophy. Nat Genet 25, 105-109.

Langlois, M. A., Lee, N. S., Rossi, J. J., and Puymirat, J. (2003). Hammerhead ribozyme-mediated destruction of nuclear foci in myotonic dystrophy myoblasts. Mol Ther 7, 670-680.

Mahadevan, M., Tsilfidis, C., Sabourin, L., Shutler, G., Amemiya, C., Jansen, G., Neville, C., Narang, M., Barcelo, J., O'Hoy, K., and et al. (1992). Myotonic dystrophy mutation: an unstable CTG repeat in the 3' untranslated region of the gene. Science 255, 1253-1255.

Moseley, M. L., Benzow, K. A., Schut, L. J., Bird, T. D., Gomez, C. M., Barkhaus, P. E., Blindauer, K. A., Labuda, M., Pandolfo, M., Koob, M. D., and Ranum, L. P. (1998). Incidence of dominant spinocerebellar and Friedreich triplet repeats among 361 ataxia families. Neurology 51, 1666-1671.

Michalowski, S., Miller, J. W., Urbinati, C. R., Paliouras, M., Swanson, M. S., and Griffith, J. (1999). Visualization of double-stranded RNAs from the myotonic dystrophy protein kinase gene and interactions with CUG-binding protein. Nucleic Acids Res 27, 3534-3542.

Miller, J. W., Urbinati, C. R., Teng-Umnuay, P., Stenberg, M. G., Byrne, B. J., Thornton, C. A., and Swanson, M. S. (2000). Recruitment of human muscleblind proteins to (CUG)(n) expansions associated with myotonic dystrophy. Embo J 19, 4439-4448.

McCampbell, A., Taylor, J. P., Taye, A. A., Robitschek, J., Li, M., Walcott, J., Merry, D., Chai, Y., Paulson, H., Sobue, G., and Fischbeck, K. H. (2000). CREB-binding protein sequestration by expanded polyglutamine. Hum Mol Genet 9, 2197-2202.

Moseley, M. L., Schut, L. J., Bird, T. D., Koob, M. D., Day, J. W., and Ranum, L. P. (2000). SCA8 CTG repeat: en masse contractions in sperm and intergenerational sequence changes may play a role in reduced penetrance. Hum Mol Genet 9, 2125-2130.

Maruyama, H., Izumi, Y., Morino, H., Oda, M., Toji, H., Nakamura, S., and Kawakami, H. (2002). Difference in disease-free survival curve and regional distribution according to subtype of spinocerebellar ataxia: a study of 1,286 Japanese patients. Am J Med Genet 114, 578-583.

Michalik, A., and Van Broeckhoven, C. (2003). Pathogenesis of polyglutamine disorders: aggregation revisited. Hum Mol Genet 12 Spec No 2, R173-186.

McLeod, C. J., O'Keefe, L. V., and Richards, R. I. (2005). The pathogenic agent in Drosophila models of 'polyglutamine' diseases. Hum Mol Genet 14, 1041-1048.

Nemes, J. P., Benzow, K. A., Moseley, M. L., Ranum, L. P., and Koob, M. D. (2000). The SCA8 transcript is an antisense RNA to a brain-specific transcript encoding a novel actin-binding protein (KLHL1). Hum Mol Genet 9, 1543-1551.

Otten, A. D., and Tapscott, S. J. (1995). Triplet repeat expansion in myotonic dystrophy alters the adjacent chromatin structure. Proc Natl Acad Sci U S A 92, 5465-5469.

Perez, M. K., Paulson, H. L., Pendse, S. J., Saionz, S. J., Bonini, N. M., and Pittman, R. N. (1998). Recruitment and the role of nuclear localization in polyglutamine-mediated aggregation. J Cell Biol 143, 1457-1470.

Pato, C. N., Macedo, A., Ambrosio, A., Vincent, J. B., Bauer, A., Schindler, K., Xu, J., Coelho, I., Dourado, A., Valente, J., et al. (2000). Detection of expansion regions in Portuguese bipolar families. Am J Med Genet 96, 854-857.

Robinson, D. N., and Cooley, L. (1997). Drosophila kelch is an oligomeric ring canal actin organizer. J Cell Biol 138, 799-810.

Ravikumar, B., Vacher, C., Berger, Z., Davies, J. E., Luo, S., Oroz, L. G., Scaravilli, F., Easton, D. F., Duden, R., O'Kane, C. J., and Rubinsztein, D. C. (2004). Inhibition of mTOR induces autophagy and reduces toxicity of polyglutamine expansions in fly and mouse models of Huntington disease. Nat Genet 36, 585-595.

Shaw, D. J., McCurrach, M., Rundle, S. A., Harley, H. G., Crow, S. R., Sohn, R., Thirion, J. P., Hamshere, M. G., Buckler, A. J., Harper, P. S., and et al. (1993). Genomic organization and transcriptional units at the myotonic dystrophy locus. Genomics 18, 673-679.

Schols, L., Amoiridis, G., Buttner, T., Przuntek, H., Epplen, J. T., and Riess, O. (1997). Autosomal dominant cerebellar ataxia: phenotypic differences in genetically defined subtypes? Ann Neurol 42, 924-932.

Strong, P. N., and Brewster, B. S. (1997). Myotonic dystrophy: molecular and cellular consequences of expanded DNA repeats are elusive. J Inherit Metab Dis 20, 159-170.

Sasagawa, N., Takahashi, N., Suzuki, K., and Ishiura, S. (1999). An expanded CTG trinucleotide repeat causes trans RNA interference: a new hypothesis for the pathogenesis of myotonic dystrophy. Biochem Biophys Res Commun 264, 76-80.

Saleem, Q., Choudhry, S., Mukerji, M., Bashyam, L., Padma, M. V., Chakravarthy, A., Maheshwari, M. C., Jain, S., and Brahmachari, S. K. (2000). Molecular analysis of autosomal dominant hereditary ataxias in the Indian population: high frequency of SCA2 and evidence for a common founder mutation. Hum Genet 106, 179-187.

Sarkar, P. S., Appukuttan, B., Han, J., Ito, Y., Ai, C., Tsai, W., Chai, Y., Stout, J. T., and Reddy, S. (2000). Heterozygous loss of Six5 in mice is sufficient to cause ocular cataracts. Nat Genet 25, 110-114.

Schols, L., Szymanski, S., Peters, S., Przuntek, H., Epplen, J. T., Hardt, C., and Riess, O. (2000). Genetic background of apparently idiopathic sporadic cerebellar ataxia. Hum Genet 107, 132-137.

Stevanin, G., Herman, A., Durr, A., Jodice, C., Frontali, M., Agid, Y., and Brice, A. (2000). Are (CTG)n expansions at the SCA8 locus rare polymorphisms? Nat Genet 24, 213; author reply 215.

Silveira, I., Alonso, I., Guimaraes, L., Mendonca, P., Santos, C., Maciel, P., Fidalgo De Matos, J. M., Costa, M., Barbot, C., Tuna, A., et al. (2000). High germinal instability of the (CTG)n at the SCA8 locus of both expanded and normal alleles. Am J Hum Genet 66, 830-840.

Sobrido, M. J., Cholfin, J. A., Perlman, S., Pulst, S. M., and Geschwind, D. H. (2001). SCA8 repeat expansions in ataxia: a controversial association. Neurology 57, 1310-1312.
Soong, B. W., Lu, Y. C., Choo, K. B., and Lee, H. Y. (2001). Frequency analysis of autosomal dominant cerebellar ataxias in Taiwanese patients and clinical and molecular characterization of spinocerebellar ataxia type 6. Arch Neurol 58, 1105-1109.

Silveira, I., Miranda, C., Guimaraes, L., Moreira, M. C., Alonso, I., Mendonca, P., Ferro, A., Pinto-Basto, J., Coelho, J., Ferreirinha, F., et al. (2002). Trinucleotide repeats in 202 families with ataxia: a small expanded (CAG)n allele at the SCA17 locus. Arch Neurol 59, 623-629.

Schols, L., Bauer, P., Schmidt, T., Schulte, T., and Riess, O. (2004). Autosomal dominant cerebellar ataxias: clinical features, genetics, and pathogenesis. Lancet Neurol 3, 291-304.

Thornton, C. A., Wymer, J. P., Simmons, Z., McClain, C., and Moxley, R. T., 3rd (1997). Expansion of the myotonic dystrophy CTG repeat reduces expression of the flanking DMAHP gene. Nat Genet 16, 407-409.

Tian, B., White, R. J., Xia, T., Welle, S., Turner, D. H., Mathews, M. B., and Thornton, C. A. (2000). Expanded CUG repeat RNAs form hairpins that activate the double-stranded RNA-dependent protein kinase PKR. Rna 6, 79-87.

Tang, B., Liu, C., Shen, L., Dai, H., Pan, Q., Jing, L., Ouyang, S., and Xia, J. (2000). Frequency of SCA1, SCA2, SCA3/MJD, SCA6, SCA7, and DRPLA CAG trinucleotide repeat expansion in patients with hereditary spinocerebellar ataxia from Chinese kindreds. Arch Neurol 57, 540-544.

Takahashi, N., Sasagawa, N., Usuki, F., Kino, Y., Kawahara, H., Sorimachi, H., Maeda, T., Suzuki, K., and Ishiura, S. (2001). Coexpression of the CUG-binding protein reduces DM protein kinase expression in COS cells. J Biochem (Tokyo) 130, 581-587.

Timchenko, N. A., Cai, Z. J., Welm, A. L., Reddy, S., Ashizawa, T., and Timchenko, L. T. (2001). RNA CUG repeats sequester CUGBP1 and alter protein levels and activity of CUGBP1. J Biol Chem 276, 7820-7826.

Topisirovic, I., Dragasevic, N., Savic, D., Ristic, A., Keckarevic, M., Keckarevic, D., Culjkovic, B., Petrovic, I., Romac, S., and Kostic, V. S. (2002). Genetic and clinical analysis of spinocerebellar ataxia type 8 repeat expansion in Yugoslavia. Clin Genet 62, 321-324.

Tazon, B., Badenas, C., Jimenez, L., Munoz, E., and Mila, M. (2002). SCA8 in the Spanish population including one homozygous patient. Clin Genet 62, 404-409.

Vincent, J. B., Neves-Pereira, M. L., Paterson, A. D., Yamamoto, E., Parikh, S. V., Macciardi, F., Gurling, H. M., Potkin, S. G., Pato, C. N., Macedo, A., et al. (2000a). An unstable trinucleotide-repeat region on chromosome 13 implicated in spinocerebellar ataxia: a common expansion locus. Am J Hum Genet 66, 819-829.

Vincent, J. B., Yuan, Q. P., Schalling, M., Adolfsson, R., Azevedo, M. H., Macedo, A., Bauer, A., DallaTorre, C., Medeiros, H. M., Pato, M. T., et al. (2000b). Long repeat tracts at SCA8 in major psychosis. Am J Med Genet 96, 873-876.

Wang, Y. H., Amirhaeri, S., Kang, S., Wells, R. D., and Griffith, J. D. (1994). Preferential nucleosome assembly at DNA triplet repeats from the myotonic dystrophy gene. Science 265, 669-671.

Wieringa, B. (1994). Myotonic dystrophy reviewed: back to the future? Hum Mol Genet 3, 1-7.

Wang, Y. H., and Griffith, J. (1995). Expanded CTG triplet blocks from the myotonic dystrophy gene create the strongest known natural nucleosome positioning elements. Genomics 25, 570-573.

Worth, P. F., Houlden, H., Giunti, P., Davis, M. B., and Wood, N. W. (2000). Large, expanded repeats in SCA8 are not confined to patients with cerebellar ataxia. Nat Genet 24, 214-215.

Yamada, M., Sato, T., Shimohata, T., Hayashi, S., Igarashi, S., Tsuji, S., and Takahashi, H. (2001). Interaction between neuronal intranuclear inclusions and promyelocytic leukemia protein nuclear and coiled bodies in CAG repeat diseases. Am J Pathol 159, 1785-1795.