25nm virus-like particles - mad cow cause?
Cells infected with scrapie and Creutzfeldt-Jakob disease agents produce intracellular 25-nm virus-like particles
Published online before print January 31, 2007
Proc. Natl. Acad. Sci. USA, 10.1073/pnas.0610999104
Neuroscience
Cells infected with scrapie and Creutzfeldt-Jakob disease agents produce intracellular 25-nm virus-like particles
Laura Manuelidis *, Zhoa-Xue Yu, Nuria Banquero, and Brian Mullins
Yale Medical School, 333 Cedar Street, New Haven, CT 06510
Communicated by Sheldon Penman, Massachusetts Institute of Technology, Cambridge, MA, December 11, 2006 (received for review October 10, 2006)
We had repeatedly found 25-nm-diameter virus-like particles in highly infectious brain fractions with little prion protein (PrP), and therefore we searched for similar virus-like particles in situ in infected cell lines with high titers. Neuroblastoma cells infected with the 22L strain of scrapie as well as hypothalamic GT cells infected with the FU Creutzfeldt-Jakob disease agent, but not parallel mock controls, displayed dense 25-nm virus-like particles in orthogonal arrays. These particles had no relation to abnormal PrP amyloid in situ, nor were they labeled by PrP antibodies that faithfully recognized rough endoplasmic reticulum membranes and amyloid fibrils, the predicted sites of normal and pathological intracellular PrP. Additionally, phorbol ester stimulated the production of abnormal PrP gel bands by >5-fold in infected N2a + 22L cells, yet this did not increase either the number of virus-like arrays or the infectious titer of these cells. Thus, the 25-nm infection-associated particles could not be prions. Synaptic differentiation and neurodegeneration, as well as retroviruses that populate the rough endoplasmic reticulum of neuroblastoma cells, were not required for particle production. The 25-nm particle arrays in cultured cells strongly resembled those first described in 1968 in synaptic regions of scrapie-infected brain and subsequently identified in many natural and experimental TSEs. The high infectivity of comparable, isolated virus-like particles that show no intrinsic PrP by antibody labeling, combined with their loss of infectivity when nucleic acid-protein complexes are disrupted, make it likely that these 25-nm particles are the causal TSE virions that induce late-stage PrP brain pathology.
Author contributions: L.M. designed research; L.M., Z.-X.Y., N.B., and B.M. performed research; L.M., Z.-X.Y., and N.B. analyzed data; and L.M. wrote the paper.
The authors declare no conflict of interest.
*To whom correspondence should be addressed.
Laura Manuelidis, E-mail: laura.manuelidis@yale.edu
pnas.org
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