Nature Methods
- 4, 345 - 351 (2007)
Published online: 11 March 2007; | doi:10.1038/nmeth1026
Fluorescence imaging of amyloid formation in living cells by a functional, tetracysteine-tagged  -synucleinMaría J Roberti1, 2, Carlos W Bertoncini1, Reinhard Klement1, Elizabeth A Jares-Erijman2 & Thomas M Jovin11
Department of Molecular Biology, Max Planck Institute for Biophysical Chemistry, 37077 Göttingen, Germany. 2
Departamento de Química Orgánica, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, 1428 Buenos Aires, Argentina.
Correspondence should be addressed to Elizabeth A Jares-Erijman eli@qo.fcen.uba.ar or Thomas M Jovin tjovin@gwdg.de  -synuclein is a major component of intraneuronal protein aggregates constituting a distinctive feature of Parkinson disease. To date, fluorescence imaging of dynamic processes leading to such amyloid deposits in living cells has not been feasible. To address this need, we generated a recombinant -synuclein (-synuclein-C4) bearing a tetracysteine target for fluorogenic biarsenical compounds. The biophysical, biochemical and aggregation properties of -synuclein-C4 matched those of the wild-type protein in vitro and in living cells. We observed aggregation of -synuclein-C4 transfected or microinjected into cells, particularly under oxidative stress conditions. Fluorescence resonance energy transfer (FRET) between FlAsH and ReAsH confirmed the close association of fibrillized -synuclein-C4 molecules. -synuclein-C4 offers the means for directly probing amyloid formation and interactions of -synuclein with other proteins in living cells, the response to cellular stress and screening drugs for Parkinson disease.
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