December 2002
Volume 43, Issue 13
ARVO Annual Meeting Abstract  |   December 2002
Mutations in Gamma-Crystallins Lead to Amyloid Fibre Formation
Author Affiliations & Notes
  • RA Quinlan
    University of Durham Durham United Kingdom
  • A Sandilands
    University of Dundee Dundee United Kingdom
  • H Long
    University of Durham Durham United Kingdom
  • V Pigaga
    University of Durham Durham United Kingdom
  • A Prescott
    University of Dundee Dundee United Kingdom
  • G Vrensen
    Dept of Morphology Netherlands Ophthalmic Research Institute Amsterdam Netherlands
  • J Löster
    GSF-National Research Center Neuherberg Germany
  • J Graw
    GSF-National Research Center Neuherberg Germany
  • C MacPhee
    University of Cambridge Cambridge University United Kingdom
  • C Dobson
    University of Cambridge Cambridge United Kingdom
  • Footnotes
    Commercial Relationships   R.A. Quinlan, None; A. Sandilands, None; H. Long, None; V. Pigaga, None; A. Prescott, None; G. Vrensen, None; J. Löster, None; J. Graw, None; C. MacPhee, None; C. Dobson, None. Grant Identification: Wellcome Trust, BBSRC, Darwin Foundation
Investigative Ophthalmology & Visual Science December 2002, Vol.43, 1922. doi:
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    • Get Citation

      RA Quinlan, A Sandilands, H Long, V Pigaga, A Prescott, G Vrensen, J Löster, J Graw, C MacPhee, C Dobson; Mutations in Gamma-Crystallins Lead to Amyloid Fibre Formation . Invest. Ophthalmol. Vis. Sci. 2002;43(13):1922.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract: : Purpose:To determine if gamma-crystallin mutants can form amyloid fibrils.Methods:Lenses were collected from the Crygbnop mouse. This mouse contains a truncated Crygb. The lenses were processed for electron microscopy. Crygbnop was cloned by RT-PCR, sequenced and subcloned into mammalian (pcDNA3.1, pEGFP) and bacterial expression vectors. A GFP-fusion construct was also generated. Contructs were transfected into PtK2 cells. Crygbnop was expressed in E. coli and the protein purified by ion exchange chromatography for in vitro studies.Results:Transfection of Crygbnop into cells induced the formation of protein inclusions. These were both cytoplasmic and intranuclear. Electron microscopy revealed a filamentous substructure to the inclusions. These were similar in morphology to the intranuclear inclusions in E17.5 Crygbnop mouse lenses. In vitro studies showed that Crygbnop formed 6-15nm filaments with amyloid fibril-like characteristics. The filaments were further characterized in terms of Congo Red staining, thioflavin T binding and x-ray diffraction.Conclusion:The data show the value of careful analysis of established maouse mutations. These data suggest a dramatic change in our perception of cataract etiologies and future treatment strategies.

Keywords: 378 crystallins • 527 protein structure/function • 528 proteins encoded by disease genes 

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