We previously reported that a (CTG)n expansion causes spinocerebellar ataxia type 8 (SCA8), a slowly progressive ataxia with reduced penetrance. We now report a transgenic mouse model in which the full-length human SCA8 mutation is transcribed using its endogenous promoter. (CTG)116 expansion, but not (CTG)11 control lines, develop a progressive neurological phenotype with in vivo imaging showing reduced cerebellar-cortical inhibition. 1C2-positive intranuclear inclusions in cerebellar Purkinje and brainstem neurons in SCA8 expansion mice and human SCA8 autopsy tissue result from translation of a polyglutamine protein, encoded on a previously unidentified antiparallel transcript (ataxin 8, ATXN8 ) spanning the repeat in the CAG direction. The neurological phenotype in SCA8 BAC expansion but not BAC control lines demonstrates the pathogenicity of the (CTG-CAG)n expansion. Moreover, the expression of noncoding (CUG)n expansion transcripts (ataxin 8 opposite strand, ATXN8OS ) and the discovery of intranuclear polyglutamine inclusions suggests SCA8 pathogenesis involves toxic gain-of-function mechanisms at both the protein and RNA levels.
Subscribe to Journal
Get full journal access for 1 year
only $17.42 per issue
All prices are NET prices.
VAT will be added later in the checkout.
Rent or Buy article
Get time limited or full article access on ReadCube.
All prices are NET prices.
Zoghbi, H.Y. & Orr, H.T. Glutamine repeats and neurodegeneration. Annu. Rev. Neurosci. 23, 217–247 (2000).
Ranum, L.P.W. & Day, J.W. Pathogenic RNA repeats: an expanding role in genetic disease. Trends Genet. 20, 506–512 (2004).
Koob, M.D. et al. Rapid cloning of expanded trinucleotide repeat sequences from genomic DNA. Nat. Genet. 18, 72–75 (1998).
Koob, M.D. et al. An untranslated CTG expansion causes a novel form of spinocerebellar ataxia (SCA8). Nat. Genet. 21, 379–384 (1999).
Juvonen, V. et al. Clinical and genetic findings in Finnish ataxia patients with the spinocerebellar ataxia 8 repeat expansion. Ann. Neurol. 48, 354–361 (2000).
Schols, L. et al. Do CTG expansions at the SCA8 locus cause ataxia? Genetic background of apparently idiopathic sporadic cerebellar ataxia. Ann. Neurol. 54, 110–115 (2003).
Ikeda, Y. et al. Asymptomatic CTG expansion at the SCA8 locus is associated with cerebellar atrophy on MRI. J. Neurol. Sci. 182, 76–79 (2000).
Silveira, I. et al. High germinal instability of the (CTG)n at the SCA8 locus of both expanded and normal alleles. Am. J. Hum. Genet. 66, 830–840 (2000).
Brusco, A. et al. Analysis of SCA8 and SCA12 loci in 134 Italian ataxic patients negative for SCA1–3, 6 and 7 CAG expansions. J. Neurol. 249, 923–929 (2002).
Day, J.W., Schut, L.J., Moseley, M.L., Durand, A.C. & Ranum, L.P.W. Spinocerebellar ataxia type 8: clinical features in a large family. Neurology 55, 649–657 (2000).
Ikeda, Y., Shizuka, M., Watanabe, M., Okamoto, K. & Shoji, M. Molecular and clinical analyses of spinocerebellar ataxia type 8 in Japan. Neurology 54, 950–955 (2000).
Ikeda, Y. et al. Spinocerebellar ataxia type 8: molecular genetic comparisons and haplotype analysis of 37 families with ataxia. Am. J. Hum. Genet. 75, 3–16 (2004).
Worth, P.F., Houlden, H., Giunti, P., Davis, M.B. & Wood, N.W. Large, expanded repeats in SCA8 are not confined to patients with cerebellar ataxia. Nat. Genet. 24, 214–215 (2000).
Stevanin, G. et al. Are (CTG)n expansions at the SCA8 locus rare polymorphisms? Nat. Genet. 24, 213 (2000).
Vincent, J.B. et al. An unstable trinucleotide-repeat region on chromosome 13 implicated in spinocerebellar ataxia: a common expansion locus. Am. J. Hum. Genet. 66, 819–829 (2000).
Sobrido, M.J., Cholfin, J.A., Perlman, S., Pulst, S.M. & Geschwind, D.H. SCA8 repeat expansions in ataxia: a controversial association. Neurology 57, 1310–1312 (2001).
Izumi, Y. et al. 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 (2003).
Sulek, A., Hoffman-Zacharska, D., Zdzienicka, E. & Zaremba, J. SCA8 repeat expansion coexists with SCA1–not only with SCA6. Am. J. Hum. Genet. 73, 972–974 (2003).
Yang, X.W., Model, P. & Heintz, N. Homologous recombination based modification in Escherichia coli and germline transformation in transgenic mice of a bacterial artificial chromosome. Nat. Biotechnol. 15, 859–865 (1997).
Shakkottai, V.G. et al. Enhanced neuronal excitability in the absence of neurodegeneration induces cerebellar ataxia. J. Clin. Invest. 113, 582–590 (2004).
Dorsman, J.C. et al. Strong aggregation and increased toxicity of polyleucine over polyglutamine stretches in mammalian cells. Hum. Mol. Genet. 11, 1487–1496 (2002).
Trottier, Y. et al. Polyglutamine expansion as a pathological epitope in Huntington's disease and four dominant ataxias. Nature 378, 403–406 (1995).
Burright, E.N. et al. SCA1 transgenic mice: a model for neurodegeneration caused by an expanded CAG trinucleotide repeat. Cell 82, 937–948 (1995).
Mittmann, W., Koch, U. & Hausser, M. Feed-forward inhibition shapes the spike output of cerebellar Purkinje cells. J. Physiol. (Lond.) 563, 369–378 (2005).
Hausser, M. & Clark, B.A. Tonic synaptic inhibition modulates neuronal output pattern and spatiotemporal synaptic integration. Neuron 19, 665–678 (1997).
Suter, K.J. & Jaeger, D. Reliable control of spike rate and spike timing by rapid input transients in cerebellar stellate cells. Neuroscience 124, 305–317 (2004).
Llano, I. & Gerschenfeld, H.M. Inhibitory synaptic currents in stellate cells of rat cerebellar slices. J. Physiol. (Lond.) 468, 177–200 (1993).
Moseley, M.L. et al. 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 (2000).
Martins, S. et al. Haplotype diversity and somatic instability in normal and expanded SCA8 alleles. Am. J. Med. Genet. B Neuropsychiatr. Genet. 139, 109–114 (2005).
Zoghbi, H.Y. & Orr, H.T. Polyglutamine diseases: protein cleavage and aggregation. Curr. Opin. Neurobiol. 9, 566–570 (1999).
Orr, H.T. Beyond the Qs in the polyglutamine diseases. Genes Dev. 15, 925–932 (2001).
Orr, H.T. The ins and outs of a polyglutamine neurodegenerative disease: spinocerebellar ataxia type 1 (SCA1). Neurobiol. Dis. 7, 129–134 (2000).
Marsh, J.L. et al. Expanded polyglutamine peptides alone are intrinsically cytotoxic and cause neurodegeneration in Drosophila . Hum. Mol. Genet. 9, 13–25 (2000).
Philips, A.V., Timchenko, L.T. & Cooper, T.A. Disruption of splicing regulated by a CUG-binding protein in myotonic dystrophy. Science 280, 737–741 (1998).
Mankodi, A. et al. Myotonic dystrophy in transgenic mice expressing an expanded CUG repeat. Science 289, 1769–1773 (2000).
Liquori, C. et al. Myotonic dystrophy type 2 caused by a CCTG expansion in intron 1 of ZNF9. Science 293, 864–867 (2001).
Kanadia, R.N. et al. A muscleblind knockout model for myotonic dystrophy. Science 302, 1978–1980 (2003).
Ranum, L.P.W. & Day, J.W. Myotonic dystrophy: RNA pathogenesis comes into focus. Am. J. Hum. Genet. 74, 793–804 (2004).
Nemes, J.P., Benzow, K.A., Moseley, M.L., Ranum, L.P.W. & Koob, M.D. 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 (2000).
Benzow, K.A. & Koob, M.D. The KLHL1-antisense transcript (KLHL1AS) is evolutionarily conserved. Mamm. Genome 13, 134–141 (2002).
Cho, D.H. et al. Antisense transcription and heterochromatin at the DM1 CTG repeats are constrained by CTCF. Mol. Cell 20, 483–489 (2005).
Margolis, R.L. et al. Huntington's Disease-like 2 (HDL2) in North America and Japan. Ann. Neurol. 56, 670–674 (2004).
Katayama, S. et al. Antisense transcription in the mammalian transcriptome. Science 309, 1564–1566 (2005).
Gao, W. et al. Optical imaging of long-term depression in the mouse cerebellar cortex in vivo . J. Neurosci. 23, 1859–1866 (2003).
Dunbar, R.L. et al. Imaging parallel fiber and climbing fiber responses and their short-term interactions in the mouse cerebellar cortex in vivo . Neuroscience 126, 213–227 (2004).
Reinert, K.C., Dunbar, R.L., Gao, W., Chen, G. & Ebner, T.J. Flavoprotein autofluorescence imaging of neuronal activation in the cerebellar cortex in vivo . J. Neurophysiol. 92, 199–211 (2004).
Ebner, T.J., Chen, G., Gao, W. & Reinert, K. Optical imaging of cerebellar functional architectures: parallel fiber beams, parasagittal bands and spreading acidification. Prog. Brain Res. 148, 125–138 (2005).
Carter, A.G. & Regehr, W.G. Quantal events shape cerebellar interneuron firing. Nat. Neurosci. 5, 1309–1318 (2002).
Hanson, C.L., Chen, G. & Ebner, T.J. Role of climbing fibers in determining the spatial patterns of activation in the cerebellar cortex to peripheral stimulation: an optical imaging study. Neuroscience 96, 317–331 (2000).
Nakamura, K. et al. SCA17, a novel autosomal dominant cerebellar ataxia caused by an expanded polyglutamine in TATA-binding protein. Hum. Mol. Genet. 10, 1441–1448 (2001).
We thank SCA8 family members for their participation; M.S. Swanson for critically reviewing our manuscript; H.T. Orr, M.T. Su, H.M. Hsieh-Li, G.W. Lee-Chen for helpful discussions and D. Norton, A. Rose and A. Koeppen for providing control autopsy tissues. Grant support from the National Ataxia Foundation and the US National Institutes of Health (RO1 NS40389) is gratefully acknowledged.
The authors declare no competing financial interests.
Immunohistochemistry of TATA binding protein. (PDF 72 kb)
Immunohistochemistry with polyglutamine-specific MW5 antibody. (PDF 98 kb)
Ataxin 8 (ATXN8) ORF and flanking sequence. (PDF 25 kb)
Genomic organization of SCA8 region and Klhl1 expression in BAC transgenic mice. (PDF 70 kb)
Primer sequences. (PDF 10 kb)
SCA8 BAC-Exp2 mouse. A high-copy SCA8 BAC-expansion animal (BAC-Exp2 at 20 weeks) displays severe motor deficits including stiffness and posturing with hindlimbs fully extended and unable to relax. (MOV 2276 kb)
SCA8 BAC-Exp5 mouse. A single-copy SCA8 BAC-expansion animal (BAC-Exp5 at 19 months) displays milder motor deficits including abnormal gait, low stance and hindlimb dragging. (MOV 691 kb)
SCA8 BAC-Control mouse. A single-copy SCA8 BAC-control animal (BAC-Ctl1 at 18 months) shows no motor phenotype. (MOV 1528 kb)
About this article
Cite this article
Moseley, M., Zu, T., Ikeda, Y. et al. Bidirectional expression of CUG and CAG expansion transcripts and intranuclear polyglutamine inclusions in spinocerebellar ataxia type 8. Nat Genet 38, 758–769 (2006). https://doi.org/10.1038/ng1827
Antisense Transcription across Nucleotide Repeat Expansions in Neurodegenerative and Neuromuscular Diseases: Progress and Mysteries
Frontiers in Cell and Developmental Biology (2020)
Proceedings of the National Academy of Sciences (2020)
Journal of Biological Chemistry (2020)
Current Opinion in Neurology (2020)