Single-stranded polyanions ≥40 bases in length facilitate the formation of hamster scrapie prions in vitro, and polyanions co-localize with PrPSc aggregates in vivo [1], [2]. To test the hypothesis that intact polyanionic molecules might serve as a structural backbone essential for maintaining the infectious conformation(s) of PrPSc, we produced synthetic prions using a photocleavable, 100-base oligonucleotide (PC-oligo). In serial Protein Misfolding Cyclic Amplification (sPMCA) reactions using purified PrPC substrate, PC-oligo was incorporated into physical complexes with PrPSc molecules that were resistant to benzonase digestion. Exposure of these nuclease-resistant prion complexes to long wave ultraviolet light (315 nm) induced degradation of PC-oligo into 5 base fragments. Light-induced photolysis of incorporated PC-oligo did not alter the infectivity of in vitro-generated prions, as determined by bioassay in hamsters and brain homogenate sPMCA assays. Neuropathological analysis also revealed no significant differences in the neurotropism of prions containing intact versus degraded PC-oligo. These results show that polyanions >5 bases in length are not required for maintaining the infectious properties of in vitro-generated scrapie prions, and indicate that such properties are maintained either by short polyanion remnants, other co-purified cofactors, or by PrPSc molecules alone.
Vyšlo v časopise: In Situ Photodegradation of Incorporated Polyanion Does Not Alter Prion Infectivity. PLoS Pathog 7(2): e32767. doi:10.1371/journal.ppat.1002001 Kategorie: Research Article prolekare.web.journal.doi_sk: https://doi.org/10.1371/journal.ppat.1002001
1. GeogheganJCValdesPAOremNRDeleaultNRWilliamsonRA 2007 Selective incorporation of polyanionic molecules into hamster prions. J Biol Chem 282 36341 36353
2. SnowADWightTNNochlinDKoikeYKimataK 1990 Immunolocalization of heparan sulfate proteoglycans to the prion protein amyloid plaques of Gerstmann-Straussler syndrome, Creutzfeldt - Jakob disease and scrapie. Lab Invest 63 601 611
3. BaslerKOeschBScottMWestawayDWalchliM 1986 Scrapie and cellular PrP isoforms are encoded by the same chromosomal gene. Cell 46 417 428
4. CastillaJSaaPHetzCSotoC 2005 In vitro generation of infectious scrapie prions. Cell 121 195 206
5. DeleaultNRHarrisBTReesJRSupattaponeS 2007 Formation of native prions from minimal componenets in vitro. Proc Natl Acad Sci U S A 104 9741 9746
6. WangFWangXYuanCGMaJ 2010 Generating a Prion with Bacterially Expressed Recombinant Prion Protein. Science 327 1132 1135
7. BruceME 1993 Scrapie strain variation and mutation. Br Med Bull 49 822 838
8. CarlsonGA 1996 Prion strains. Curr Top Microbiol Immunol 207 35 47
9. BessenRAMarshRF 1994 Distinct PrP properties suggest the molecular basis of strain variation in transmissible mink encephalopathy. J Virol 68 7859 7868
10. TellingGCParchiPDeArmondSJCortelliPMontagnaP 1996 Evidence for the conformation of the pathologic isoform of the prion protein enciphering and propagating prion diversity. Science 274 2079 2082
11. DeleaultNRLucassenRWSupattaponeS 2003 RNA molecules stimulate prion protein conversion. Nature 425 717 720
12. DeleaultNRGeogheganJCNishinaKKascsakRWilliamsonRA 2005 Protease-resistant Prion Protein Amplification Reconstituted with Partially Purified Substrates and Synthetic Polyanions. J Biol Chem 280 26873 26879
13. WongCXiongLWHoriuchiMRaymondLWehrlyK 2001 Sulfated glycans and elevated temperature stimulate PrP(Sc)-dependent cell-free formation of protease-resistant prion protein. Embo J 20 377 386
14. ShakedGMMeinerZAvrahamITaraboulosAGabizonR 2001 Reconstitution of prion infectivity from solubilized protease-resistant PrP and nonprotein components of prion rods. J Biol Chem 276 14324 14328
15. Ben-ZakenOTzabanSTalYHoronchikLEskoJD 2003 Cellular heparan sulfate participates in the metabolism of prions. J Biol Chem 278 40041 40049
16. WarnerRGHundtCWeissSTurnbullJE 2002 Identification of the heparan sulfate binding sites in the cellular prion protein. J Biol Chem 277 18421 18430
17. CaugheyBBrownKRaymondGJKatzensteinGEThresherW 1994 Binding of the protease-sensitive form of PrP (prion protein) to sulfated glycosaminoglycan and congo red [corrected]. J Virol 68 2135 2141
18. AkowitzASklaviadisTManuelidisEEManuelidisL 1990 Nuclease-resistant polyadenylated RNAs of significant size are detected by PCR in highly purified Creutzfeldt-Jakob disease preparations. Microb Pathog 9 33 45
19. SafarJGKellingsKSerbanAGrothDCleaverJE 2005 Search for a prion-specific nucleic acid. J Virol 79 10796 10806
20. JeongBHKimNHJinJKChoiJKLeeYJ 2009 Reduction of prion infectivity and levels of scrapie prion protein by lithium aluminum hydride: implications for RNA in prion diseases. J Neuropathol Exp Neurol 68 870 879
21. AlperT 1985 Scrapie agent unlike viruses in size and susceptibility to inactivation by ionizing or ultraviolet radiation. Nature 317 750
22. Bellinger-KawaharaCCleaverJEDienerTOPrusinerSB 1987 Purified scrapie prions resist inactivation by UV irradiation. J Virol 61 159 166
23. GriffithJS 1967 Self-replication and scrapie. Nature 215 1043 1044
24. CohenFEPanKMHuangZBaldwinMFletterickRJ 1994 Structural clues to prion replication. Science 264 530 531
25. DeleaultNRKascsakRGeogheganJCSupattaponeS 2010 Species-dependent differences in cofactor utilization for formation of the protease-resistant prion protein in vitro. Biochemistry 49 3928 3934
26. GillACAgarwalSPinheiroTJGrahamJF 2010 Structural requirements for efficient prion protein conversion: Cofactors may promote a conversion-competent structure for PrP(C). Prion 4 235 242
Zvýšte si kvalifikáciu online z pohodlia domova
Nemáte účet? Registrujte sa
Zadajte e-mailovú adresu, s ktorou ste vytvárali účet. Budú Vám na ňu zasielané informácie k nastaveniu nového hesla.
