Molecular dissection of ALS-associated toxicity of SOD1 in transgenic mice using an exon-fusion approach
Han-Xiang Deng, Hujun Jiang, Ronggen Fu, Hong Zhai, Yong Shi, Erdong Liu, Makito Hirano, Mauro C Dal Canto, Teepu Siddique, Han-Xiang Deng, Hujun Jiang, Ronggen Fu, Hong Zhai, Yong Shi, Erdong Liu, Makito Hirano, Mauro C Dal Canto, Teepu Siddique
Abstract
Mutations in Cu,Zn superoxide dismutase (SOD1) are associated with amyotrophic lateral sclerosis (ALS). Among more than 100 ALS-associated SOD1 mutations, premature termination codon (PTC) mutations exclusively occur in exon 5, the last exon of SOD1. The molecular basis of ALS-associated toxicity of the mutant SOD1 is not fully understood. Here, we show that nonsense-mediated mRNA decay (NMD) underlies clearance of mutant mRNA with a PTC in the non-terminal exons. To further define the crucial ALS-associated SOD1 fragments, we designed and tested an exon-fusion approach using an artificial transgene SOD1(T116X) that harbors a PTC in exon 4. We found that the SOD1(T116X) transgene with a fused exon could escape NMD in cellular models. We generated a transgenic mouse model that overexpresses SOD1(T116X). This mouse model developed ALS-like phenotype and pathology. Thus, our data have demonstrated that a 'mini-SOD1' of only 115 amino acids is sufficient to cause ALS. This is the smallest ALS-causing SOD1 molecule currently defined. This proof of principle result suggests that the exon-fusion approach may have potential not only to further define a shorter ALS-associated SOD1 fragment, thus providing a molecular target for designing rational therapy, but also to dissect toxicities of other proteins encoded by genes of multiple exons through a 'gain of function' mechanism.
Figures
References
- Rosen D.R., Siddique T., Patterson D., Figlewicz D.A., Sapp P., Hentati A., Donaldson D., Goto J., O'Regan J.P., Deng H.X., et al. Mutations in Cu/Zn superoxide dismutase gene are associated with familial amyotrophic lateral sclerosis. Nature. 1993;362:59–62.
- Deng H.X., Hentati A., Tainer J.A., Iqbal Z., Cayabyab A., Hung W.Y., Getzoff E.D., Hu P., Herzfeldt B., Roos R.P., et al. Amyotrophic lateral sclerosis and structural defects in Cu,Zn superoxide dismutase. Science. 1993;261:1047–1051.
- Gurney M.E., Pu H., Chiu A.Y., Dal Canto M.C., Polchow C.Y., Alexander D.D., Caliendo J., Hentati A., Kwon Y.W., Deng H.X., et al. Motor neuron degeneration in mice that express a human Cu,Zn superoxide dismutase mutation. Science. 1994;264:1772–1775.
- Reaume A.G., Elliott J.L., Hoffman E.K., Kowall N.W., Ferrante R.J., Siwek D.F., Wilcox H.M., Flood D.G., Beal M.F., Brown R.H., Jr, et al. Motor neurons in Cu/Zn superoxide dismutase-deficient mice develop normally but exhibit enhanced cell death after axonal injury. Nat. Genet. 1996;13:43–47.
- Deng H.X., Shi Y., Furukawa Y., Zhai H., Fu R., Liu E., Gorrie G.H., Khan M.S., Hung W.Y., Bigio E.H., et al. Conversion to the amyotrophic lateral sclerosis phenotype is associated with intermolecular linked insoluble aggregates of SOD1 in mitochondria. Proc. Natl Acad. Sci. USA. 2006;103:7142–7147.
- Furukawa Y., Fu R., Deng H.X., Siddique T., O'Halloran T.V. Disulfide cross-linked protein represents a significant fraction of ALS-associated Cu, Zn-superoxide dismutase aggregates in spinal cords of model mice. Proc. Natl Acad. Sci. USA. 2006;103:7148–7153.
- Kunst C.B. Complex genetics of amyotrophic lateral sclerosis. Am. J. Hum. Genet. 2004;75:933–947.
- Hardy J., Selkoe D.J. The amyloid hypothesis of Alzheimer's disease: progress and problems on the road to therapeutics. Science. 2002;297:353–356.
- Supattapone S., Bosque P., Muramoto T., Wille H., Aagaard C., Peretz D., Nguyen H.O., Heinrich C., Torchia M., Safar J., et al. Prion protein of 106 residues creates an artifical transmission barrier for prion replication in transgenic mice. Cell. 1999;96:869–878.
- Frenkel D., Balass M., Solomon B. N-terminal EFRH sequence of Alzheimer's beta-amyloid peptide represents the epitope of its anti-aggregating antibodies. J. Neuroimmunol. 1998;88:85–90.
- Bard F., Cannon C., Barbour R., Burke R.L., Games D., Grajeda H., Guido T., Hu K., Huang J., Johnson-Wood K., et al. Peripherally administered antibodies against amyloid beta-peptide enter the central nervous system and reduce pathology in a mouse model of Alzheimer disease. Nat. Med. 2000;6:916–919.
- Taylor J.P., Hardy J., Fischbeck K.H. Toxic proteins in neurodegenerative disease. Science. 2002;296:1991–1995.
- Jonsson P.A., Ernhill K., Andersen P.M., Bergemalm D., Brannstrom T., Gredal O., Nilsson P., Marklund S.L. Minute quantities of misfolded mutant superoxide dismutase-1 cause amyotrophic lateral sclerosis. Brain. 2004;127:73–88.
- Wang J., Xu G., Li H., Gonzales V., Fromholt D., Karch C., Copeland N.G., Jenkins N.A., Borchelt D.R. Somatodendritic accumulation of misfolded SOD1-L126Z in motor neurons mediates degeneration: alphaB-crystallin modulates aggregation. Hum. Mol. Genet. 2005;14:2335–2347.
- Watanabe Y., Yasui K., Nakano T., Doi K., Fukada Y., Kitayama M., Ishimoto M., Kurihara S., Kawashima M., Fukuda H., et al. Mouse motor neuron disease caused by truncated SOD1 with or without C-terminal modification. Brain Res. Mol. Brain Res. 2005;135:12–20.
- Maquat L.E. Nonsense-mediated mRNA decay: splicing, translation and mRNP dynamics. Nat. Rev. Mol. Cell Biol. 2004;5:89–99.
- Pereira F.J., do Ceu Silva M., Picanco I., Seixas M.T., Ferrao A., Faustino P., Romao L. Human alpha2-globin nonsense-mediated mRNA decay induced by a novel alpha-thalassaemia frameshift mutation at codon 22. Br. J. Haematol. 2006;133:98–102.
- Wong P.C., Pardo C.A., Borchelt D.R., Lee M.K., Copeland N.G., Jenkins N.A., Sisodia S.S., Cleveland D.W., Price D.L. An adverse property of a familial ALS-linked SOD1 mutation causes motor neuron disease characterized by vacuolar degeneration of mitochondria. Neuron. 1995;14:1105–1116.
- Bruijn L.I., Becher M.W., Lee M.K., Anderson K.L., Jenkins N.A., Copeland N.G., Sisodia S.S., Rothstein J.D., Borchelt D.R., Price D.L., et al. ALS-linked SOD1 mutant G85R mediates damage to astrocytes and promotes rapidly progressive disease with SOD1-containing inclusions. Neuron. 1997;18:327–338.
- Beckman J.S., Estevez A.G., Crow J.P., Barbeito L. Superoxide dismutase and the death of motoneurons in ALS. Trends Neurosci. 2001;24:S15–S20.
- Liochev S.I., Fridovich I. Mutant Cu,Zn superoxide dismutases and familial amyotrophic lateral sclerosis: evaluation of oxidative hypotheses. Free Radic. Biol. Med. 2003;34:1383–1389.
- Bruijn L.I., Miller T.M., Cleveland D.W. Unraveling the mechanisms involved in motor neuron degeneration in ALS. Annu. Rev. Neurosci. 2004;27:723–749.
- Manfredi G., Xu Z. Mitochondrial dysfunction and its role in motor neuron degeneration in ALS. Mitochondrion. 2005;5:77–87.
- Tiwari A., Hayward L.J. Mutant SOD1 instability: implications for toxicity in amyotrophic lateral sclerosis. Neurodegener. Dis. 2005;2:115–127.
- Rakhit R., Chakrabartty A. Structure, folding, and misfolding of Cu,Zn superoxide dismutase in amyotrophic lateral sclerosis. Biochim. Biophys. Acta. 2006;1762:1025–1037.
- Culotta V.C., Yang M., O'Halloran T.V. Activation of superoxide dismutases: putting the metal to the pedal. Biochim. Biophys. Acta. 2006;1763:747–758.
- Kato S., Hayashi H., Nakashima K., Nanba E., Kato M., Hirano A., Nakano I., Asayama K., Ohama E. Pathological characterization of astrocytic hyaline inclusions in familial amyotrophic lateral sclerosis. Am. J. Pathol. 1997;151:611–620.
- Yoshihara T., Ishii T., Iwata M., Nomoto M. Ultrastructural and histochemical study of the motor end plates of the intrinsic laryngeal muscles in amyotrophic lateral sclerosis. Ultrastruct. Pathol. 1998;22:121–126.
- Cha C.I., Kim J.M., Shin D.H., Kim Y.S., Kim J., Gurney M.E., Lee K.W. Reactive astrocytes express nitric oxide synthase in the spinal cord of transgenic mice expressing a human Cu/Zn SOD mutation. Neuroreport. 1998;9:1503–1506.
- Rafalowska J., Podlecka A. Does the pathological factor in amyotrophic lateral sclerosis (ALS) damage also astrocytes? Folia Neuropathol. 1998;36:87–93.
- Trotti D., Rolfs A., Danbolt N.C., Brown R.H., Jr, Hediger M.A. SOD1 mutants linked to amyotrophic lateral sclerosis selectively inactivate a glial glutamate transporter. Nat. Neurosci. 1999;2:427–433.
- Boillee S., Yamanaka K., Lobsiger C.S., Copeland N.G., Jenkins N.A., Kassiotis G., Kollias G., Cleveland D.W. Onset and progression in inherited ALS determined by motor neurons and microglia. Science. 2006;312:1389–1392.
- Beers D.R., Henkel J.S., Xiao Q., Zhao W., Wang J., Yen A.A., Siklos L., McKercher S.R., Appel S.H. Wild-type microglia extend survival in PU.1 knockout mice with familial amyotrophic lateral sclerosis. Proc. Natl Acad. Sci. USA. 2006;103:16021–16026.
- Ghadge G.D., Wang L., Sharma K., Monti A.L., Bindokas V., Stevens F.J., Roos R.P. Truncated wild-type SOD1 and FALS-linked mutant SOD1 cause neural cell death in the chick embryo spinal cord. Neurobiol. Dis. 2006;21:194–205.
- Fischer L.R., Culver D.G., Tennant P., Davis A.A., Wang M., Castellano-Sanchez A., Khan J., Polak M.A., Glass J.D. Amyotrophic lateral sclerosis is a distal axonopathy: evidence in mice and man. Exp. Neurol. 2004;185:232–240.
- Gould T.W., Buss R.R., Vinsant S., Prevette D., Sun W., Knudson C.M., Milligan C.E., Oppenheim R.W. Complete dissociation of motor neuron death from motor dysfunction by Bax deletion in a mouse model of ALS. J. Neurosci. 2006;26:8774–8786.
- Wang J., Xu G., Slunt H.H., Gonzales V., Coonfield M., Fromholt D., Copeland N.G., Jenkins N.A., Borchelt D.R. Coincident thresholds of mutant protein for paralytic disease and protein aggregation caused by restrictively expressed superoxide dismutase cDNA. Neurobiol. Dis. 2005;20:943–952.
Source: PubMed