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A FRET Sensor for Non-Invasive Imaging of Amyloid Formation in Vivo

Authors

  • Dr. Gabriele S. Kaminski Schierle,

    1. Department of Chemical Engineering and Biotechnology, University of Cambridge, Pembroke Street, Cambridge CB2 3RA (U.K.), Fax: (+44) 1223 334796
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    • These authors contributed equally to the work.

  • Dr. Carlos W. Bertoncini,

    1. Department of Chemistry, University of Cambridge, Lensfield Road, Cambridge, CB2 1EW (U.K.), Fax: (+44) 1223 336362
    2. Laboratory of Molecular Biophysics, Institute for Research in Biomedicine, Baldiri Reixac 10-12, 08028, Barcelona (Spain)
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    • These authors contributed equally to the work.

  • Fiona T. S. Chan,

    1. Department of Chemical Engineering and Biotechnology, University of Cambridge, Pembroke Street, Cambridge CB2 3RA (U.K.), Fax: (+44) 1223 334796
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  • Annemieke T. van der Goot,

    1. Department of Genetics, University Medical Centre Groningen and University of Groningen, 9700 RB Groningen (The Netherlands)
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  • Dr. Stefanie Schwedler,

    1. Physikalische Chemie I, Fakultät für Chemie, Universität Bielefeld, Universitätsstr. 25, 33615, Bielefeld (Germany)
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  • Dr. Jeremy Skepper,

    1. Department of Physiology, Development and Neuroscience, University of Cambridge, Downing Street, Cambridge CB2 3DY (U.K.)
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  • Dr. Simon Schlachter,

    1. Department of Chemical Engineering and Biotechnology, University of Cambridge, Pembroke Street, Cambridge CB2 3RA (U.K.), Fax: (+44) 1223 334796
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  • Dr. Tjakko van Ham,

    1. Department of Genetics, University Medical Centre Groningen and University of Groningen, 9700 RB Groningen (The Netherlands)
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  • Dr. Alessandro Esposito,

    1. Department of Chemical Engineering and Biotechnology, University of Cambridge, Pembroke Street, Cambridge CB2 3RA (U.K.), Fax: (+44) 1223 334796
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  • Dr. Janet R. Kumita,

    1. Department of Chemistry, University of Cambridge, Lensfield Road, Cambridge, CB2 1EW (U.K.), Fax: (+44) 1223 336362
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  • Dr. Ellen A. A. Nollen,

    1. Department of Genetics, University Medical Centre Groningen and University of Groningen, 9700 RB Groningen (The Netherlands)
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  • Prof. Christopher M. Dobson,

    Corresponding author
    1. Department of Chemistry, University of Cambridge, Lensfield Road, Cambridge, CB2 1EW (U.K.), Fax: (+44) 1223 336362
    • Department of Chemistry, University of Cambridge, Lensfield Road, Cambridge, CB2 1EW (U.K.), Fax: (+44) 1223 336362
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  • Dr. Clemens F. Kaminski

    Corresponding author
    1. Department of Chemical Engineering and Biotechnology, University of Cambridge, Pembroke Street, Cambridge CB2 3RA (U.K.), Fax: (+44) 1223 334796
    2. Friedrich-Alexander University of Erlangen Nürnberg, 91052 Erlangen (Germany)
    • Department of Chemical Engineering and Biotechnology, University of Cambridge, Pembroke Street, Cambridge CB2 3RA (U.K.), Fax: (+44) 1223 334796
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Abstract

Misfolding and aggregation of amyloidogenic polypeptides lie at the root of many neurodegenerative diseases. Whilst protein aggregation can be readily studied in vitro by established biophysical techniques, direct observation of the nature and kinetics of aggregation processes taking place in vivo is much more challenging. We describe here, however, a Förster resonance energy transfer sensor that permits the aggregation kinetics of amyloidogenic proteins to be quantified in living systems by exploiting our observation that amyloid assemblies can act as energy acceptors for variants of fluorescent proteins. The observed lifetime reduction can be attributed to fluorescence energy transfer to intrinsic energy states associated with the growing amyloid species. Indeed, for α-synuclein, a protein whose aggregation is linked to Parkinson′s disease, we have used this sensor to follow the kinetics of the self-association reactions taking place in vitro and in vivo and to reveal the nature of the ensuing aggregated species. Experiments were conducted in vitro, in cells in culture and in living Caenorhabditis elegans. For the latter the readout correlates directly with the appearance of a toxic phenotype. The ability to measure the appearance and development of pathogenic amyloid species in a living animal and the ability to relate such data to similar processes observed in vitro provides a powerful new tool in the study of the pathology of the family of misfolding disorders. Our study confirms the importance of the molecular environment in which aggregation reactions take place, highlighting similarities as well as differences between the processes occurring in vitro and in vivo, and their significance for defining the molecular physiology of the diseases with which they are associated.

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