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Evidence of mitochondrial involvement in scrapie infection
J M Aiken1, J L Williamson, R F Marsh
1Department of Veterinary Science, University of Wisconsin-Madison 53706.
Abstract:
Two cDNA libraries were constructed from brain membrane and cytoskeletal preparations purified from scrapie-infected hamster brains. Four recombinants strongly preferential to the scrapie cytoskeletal preparation were identified by the differential hybridization of 7,000 recombinants. These clones were not, however, preferential to total nucleic acids extracted from scrapie-infected hamster brains. DNA sequence analysis revealed all four clones to have significant sequence similarities to the mouse mitochondrial genome. This correlation led us to consider a mitochondrial association with scrapie infectivity. Brain mitochondria were purified by sucrose gradient density centrifugation and found to contain high infectivity. Removal of mitochondrial outer membranes by osmotic shock or digitonin treatment resulted in no detectable loss of titer.
Insights
Scrapie infectivity in hamster brains is associated with mitochondria. Purified brain mitochondria showed high infectivity, suggesting a role in prion disease pathogenesis.
Area of Science:
- Neuroscience
- Molecular Biology
- Infectious Diseases
Background:
- Scrapie is a transmissible spongiform encephalopathy.
- The precise location and form of scrapie infectivity within the brain remain incompletely understood.
Purpose of the Study:
- To investigate the subcellular localization of scrapie infectivity.
- To identify potential molecular components associated with scrapie pathogenesis.
Main Methods:
- Construction of cDNA libraries from scrapie-infected hamster brain preparations (membrane and cytoskeletal).
- Differential hybridization screening of cDNA libraries.
- DNA sequence analysis of selected clones.
- Mitochondrial purification using sucrose gradient density centrifugation.
- Infectivity assays of purified fractions.
Main Results:
- Four cDNA clones preferentially hybridized to scrapie cytoskeletal preparations but not total brain nucleic acids.
- Sequence analysis revealed these clones are homologous to the mouse mitochondrial genome.
- Purified brain mitochondria exhibited high scrapie infectivity.
- Mitochondrial infectivity was retained after outer membrane removal.
Conclusions:
- Mitochondria are strongly implicated in the localization of scrapie infectivity within the brain.
- Mitochondrial components may play a direct role in scrapie pathogenesis or serve as a vehicle for the infectious agent.