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Dissociation of infectivity from seeding ability in prions with alternate docking mechanism.

ABSTRACT: Previous studies identified two mammalian prion protein (PrP) polybasic domains that bind the disease-associated conformer PrP(Sc), suggesting that these domains of cellular prion protein (PrP(C)) serve as docking sites for PrP(Sc) during prion propagation. To examine the role of polybasic domains in the context of full-length PrP(C), we used prion proteins lacking one or both polybasic domains expressed from Chinese hamster ovary (CHO) cells as substrates in serial protein misfolding cyclic amplification (sPMCA) reactions. After ?5 rounds of sPMCA, PrP(Sc) molecules lacking the central polybasic domain (?C) were formed. Surprisingly, in contrast to wild-type prions, ?C-PrP(Sc) prions could bind to and induce quantitative conversion of all the polybasic domain mutant substrates into PrP(Sc) molecules. Remarkably, ?C-PrP(Sc) and other polybasic domain PrP(Sc) molecules displayed diminished or absent biological infectivity relative to wild-type PrP(Sc), despite their ability to seed sPMCA reactions of normal mouse brain homogenate. Thus, ?C-PrP(Sc) prions interact with PrP(C) molecules through a novel interaction mechanism, yielding an expanded substrate range and highly efficient PrP(Sc) propagation. Furthermore, polybasic domain deficient PrP(Sc) molecules provide the first example of dissociation between normal brain homogenate sPMCA seeding ability from biological prion infectivity. These results suggest that the propagation of PrP(Sc) molecules may not depend on a single stereotypic mechanism, but that normal PrP(C)/PrP(Sc) interaction through polybasic domains may be required to generate prion infectivity.

SUBMITTER: Miller MB

PROVIDER: S-EPMC3136465 | biostudies-literature | 2011 Jul

REPOSITORIES: biostudies-literature

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Dissociation of infectivity from seeding ability in prions with alternate docking mechanism.

Miller Michael B MB Geoghegan James C JC Supattapone Surachai S

PLoS pathogens 20110714 7

Previous studies identified two mammalian prion protein (PrP) polybasic domains that bind the disease-associated conformer PrP(Sc), suggesting that these domains of cellular prion protein (PrP(C)) serve as docking sites for PrP(Sc) during prion propagation. To examine the role of polybasic domains in the context of full-length PrP(C), we used prion proteins lacking one or both polybasic domains expressed from Chinese hamster ovary (CHO) cells as substrates in serial protein misfolding cyclic amp ...[more]

PMID: 21779169

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Project description:The global outbreak of bovine spongiform encephalopathy (BSE) has been attributed to the recycling of contaminated meat and bone meals (MBMs) as feed supplements. The use of MBMs has been prohibited in many countries; however, the development of a method for inactivating BSE prions could enable the efficient and safe use of these products as an organic resource. Subcritical water (SCW), which is water heated under pressure to maintain a liquid state at temperatures below the critical temperature (374°C), exhibits strong hydrolytic activity against organic compounds. In this study, we examined the residual in vitro seeding activity of protease-resistant prion protein (PrPSc) and the infectivity of BSE prions after SCW treatments. Spinal cord homogenates prepared from BSE-infected cows were treated with SCW at 230-280°C for 5-7.5 min and used to intracerebrally inoculate transgenic mice overexpressing bovine prion protein. Serial protein misfolding cyclic amplification (sPMCA) analysis detected no PrPSc in the SCW-treated homogenates, and the mice treated with these samples survived for more than 700 days without any signs of disease. However, sPMCA analyses detected PrPSc accumulation in the brains of all inoculated mice. Furthermore, secondary passage mice, which inoculated with brain homogenates derived from a western blotting (WB)-positive primary passage mouse, died after an average of 240 days, similar to mice inoculated with untreated BSE-infected spinal cord homogenates. The PrPSc accumulation and vacuolation typically observed in the brains of BSE-infected mice were confirmed in these secondary passage mice, suggesting that the BSE prions maintained their infectivity after SCW treatment. One late-onset case, as well as asymptomatic but sPMCA-positive cases, were also recognized in secondary passage mice inoculated with brain homogenates from WB-negative but sPMCA-positive primary passage mice. These results indicated that SCW-mediated hydrolysis was insufficient to eliminate the infectivity of BSE prions under the conditions tested.

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Dissociation of infectivity from seeding ability in prions with alternate docking mechanism.

Publications

Dissociation of infectivity from seeding ability in prions with alternate docking mechanism.

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