Below is the uncorrected machine-read text of this chapter, intended to provide our own search engines and external engines with highly rich, chapter-representative searchable text of each book. Because it is UNCORRECTED material, please consider the following text as a useful but insufficient proxy for the authoritative book pages.
Basic Biomedical Research _~ on Transmissible Spongiform Encephalopathies Research focused directly on the development of antemortem tests for the diagnosis of transmissible spongiform encephalopathies (TSEs) may prove fruitful, and certain promising approaches will be discussed in the next chapter. However, the committee strongly believes that the fastest way to develop rapid, noninvasive, early-stage diagnostic tools for TSEs is through basic biomedical research that can fill gaps in the fundamental knowledge of prions and their disease-causing properties. The European experience provides evidence that applied research alone is insufficient. The European Union has spent approximately 30 million euros ($30.7 million) over the past 10 years in an attempt to develop satis- factory postmortem diagnostic tests for BSE, yet the Western blot test, which has been in use for three decades, is still the most commonly employed method (personal communication, A. Aguzzi, University Hospital of Zurich, July15, 20021. The history of medicine also provides numerous examples of basic dis- coveries that were essential precursors to the development of diagnostics. For instance, before inexpensive and rapid diagnostics were available for non-A, non-B hepatitis, the hepatitis C virus had to be identified and cloned. Likewise, before an effective blood test became available for screening for HIV, the virus first had to be identified and isolated. Many brilliant and dedicated scientists have been working for more than 20 years to solve the mysteries related to disease-causing priors. Col- lectively they have made great progress, yet many fundamental questions remain. To answer these questions, scientists must solve the structure of 60
BASIC BIOMEDICAL RESEARCH ON TSEs 6 PrPSc and relate that structure to prion strain differences (see Box 3-11; determine endogenous and exogenous mechanisms of prion replication (see Box 3-21; clarify the pathways and pathogenic mechanisms used by prions (see Box 3-31; and elucidate the physiological function of prpC (see Box 3- 41. Each of these priority areas for basic research on prions is discussed, in turn, below. STRUCTURAL FEATURES OF PRIONS Current models of prion conformation and tertiary structure are nei- ther complete nor conclusive. Defining the critical structural differences between infectious and noninfectious forms of the prion protein could pro- vide the basis for TSE diagnostics and elucidate the correlations between PrP structures and strains. The committee believes that research to these ends should be high priority for support by the National Prion Research Program (NPRP). Defining the structural differences between PrP isoforms might enable scientists to synthesize a PrPSc-specific antibody probe or aptamer, opening the door to the development of a TSE diagnostic tool. Antibody probes are increasingly being used to detect infectious agents in tissue, but their appli- cation to prion detection is limited because no independently validated antibody binds exclusively to PrPSc without prior digestion of PrPC. Of note, two different groups of investigators have reported separate antibodies spe- cific to PrPSc (Korth et al., 1997; Paramithiotis et al., 20031. Many of the tests for detection of PrPSc use antibody probes, as de- scribed in Chapter 4. Most tests, however, must use proteinase K to distin-
62 ADVANCING PRION SCIENCE guish prpC from PrPSc. Proteinase K digestion reduces the already miniscule amount of material usually available for prion detection. The committee looks forward to the development of new test methods that exclude pro- teinase K. Defining the prion structure could also reveal structure-based pheno- typic differences among prion strains. This information would be impor- tant because it is thought that prpC binds to PrPSc before it is converted into PrPSc (Caughey, 20021. If the structure of the prion strain differs too greatly from the host's PrPC, binding may occur, but conversion will not (Bessen et al., 19951. Defining the structures of prpC and PrPSc at the sites where they interact during binding and conformational change would support the de- velopment of molecules to block those interactions. Since prions are largely insoluble, it is particularly difficult to study their structures with standard proteomic tools. A newer technique called solid-state nuclear magnetic resonance (NMR) overcomes the solubility problem and is in the early stages of application to research on the three- dimensional structure of prions (Laws et al., 2001; Wemmer, 20021. In addition, electron crystallography is being used to probe the two-dimen- sional structures of PrPSc crystals (Wille, 20021. Nobel Laureate Kurt Wuthrich produced three-dimensional models of prpC using liquid-phase NMR experiments (Zahn et al., 20001. Nevertheless, the research commu- nity must do much more to obtain a comprehensive understanding of the structural differences between infectious and noninfectious PrP. MOLECULAR MECHANISMS OF PRION REPLICATION It is believed that both the conversion of cellular PrP to PrPSc and the accumulation of prions require the assistance of one or more molecules
BASIC BIOMEDICAL RESEARCH ON TSEs 63 (Caughey, 2001~. These ancillary or chaperoning factors could serve as sur- rogate markers for prion detection and as drug targets for TSE therapeutics and prophylaxes. Therefore, it is critical that the NPRP fund research de- signed to identify the molecules that facilitate PrPSc formation and accumu- . . . anon In salvo. Experiments have demonstrated that the chaperone proteins GroEL and hsplO4 can stimulate the cell-free conversion reaction, as can sulfated gly- cans (Won" et al., 2001) and partial denaturants (DebBurman et al., 19971. Other chaperone proteins thought to modulate PrP conversion include hsp73 (Tatzelt et al., 1995), members of the hsp60 class (DebBurman et al., 1997; Edenhofer et al., 1996; Stocke! and HartI, 2001), and BiP (Tin et al., 2000~. There are at least a half-dozen apparently natural PrP ligands or conversion modulators: copper (II) (Hornshaw et al., 1995), the laminin receptor (Weiss and Randour, 2002), laminin (Graner et al., 2000), stress- induced protein 1 (Zanata et al., 2002), and nucleic acids (Cordeiro et al., 2001; Gabus et al., 2001; Nandi and Leclerc, 19991. The molecule or molecules associated with prion conversion may be easier to detect than prions themselves. For instance, there may be a known antibody that binds specifically to a chaperone protein involved with PrP formation or accumulation. Ancillary or chaperoning factors could poten- tially amplify PrPSc, helping to overcome current limits to prion detection. A related goal of NPRP should be to fund research aimed at isolating the multiprotein complexes that contain priors. Such studies might identify new cofactors that are important in the formation and stabilization of PrPSc and infectivity. For instance, molecules such as sulfated glycosaminogly- cans appear to be associated with PrPSc deposits in viva (Snow et al., 1990) and may play a role in their formation in viva, as can be the case with a variety of other amyloid protein deposits. Further understanding of the iden- tities and roles of PrPSc-associated molecules might suggest new therapeutic and diagnostic approaches. MECHANISMS OF TSE PATHOGENESIS If better diagnostics are to be developed, much more understanding of the nature and dynamics of prion infection must be gained. Research on the pathogenesis of TSEs holds the keys to such understanding. Researchers, clinicians, and public health officials must know which tissues are infec- tious and when, what mechanisms are involved when the infectious agent enters and disseminates in the body and then invades the brain, what causes cellular toxicity (priors, prions plus another molecular species, or a totally different molecular entity), what the mechanisms are by which the toxic events lead to cellular dysfunction and clinical symptoms, how the infec- tious agent spreads from host to host, and what features of the host deter-
64 ADVANCING PRION SCIENCE mine susceptibility to infection. Understanding of these matters will lead to improved characterization of diagnostic targets, better diagnostic strate- gies, greater target discrimination, and improved diagnostic sensitivity and ... . spec~c~ty. The pathogenesis of TSE agents may vary among strains and hosts, and the same host may be infected by more than one strain. For example, we cannot assume that known TSE strains are pure or uniform. In mouse as- says, a given strain (phenotype) has characteristic histopathological fea- tures and incubation periods. Yet in a recent study, transgenic mice express- ing human PrP and inoculated with the BSE agent manifested not only the expected vCJD phenotype, but also, surprisingly, the sCJD phenotype (Asante et al., 20021. The researchers suggest, in general, that TSE infec- tions may involve multiple strains resulting in variable host responses. More specifically, they postulate that some of the sCTD occurring in the United Kingdom and elsewhere may be due to BSE exposure. A better understand- ing of prion strains and subtypes and their differential pathogenesis, there-
BASIC BIOMEDICAL RESEARCH ON TSEs 65 fore, is crucial. In addition, the conditions under which a TSE agent may exist in an asymptomatic carrier state within the host should be explored. The general factors affecting host resistance are known and include genetic, environmental, and agent-specific characteristics. Their interrela- tionships are poorly understood, however. A recent experiment by lean Manson and colleagues demonstrates this point. Manson's group showed that mice expressing PrP with the LeulO1 mutation, corresponding to hu- man GSS, did not spontaneously develop clinical or subclinical prion dis- ease (Manson et al., 19991. However, their experiment also showed that this disease-associated mutation differentially increased susceptibility to in- fection with human GSS-derived infectivity and simultaneously decreased susceptibility to several mouse scrapie strains. Thus, the study demonstrated that mutant PrP expression was an important genetic susceptibility factor, although it was unable to generate spontaneous infectivity in viva alone. Continued research in this area will lead to a firmer understanding of how genetic factors interact with the TSE agent and environmental factors. All pathogenetic studies must be interpreted in their specific contexts, a requirement that makes the development of diagnostics more difficult. At the same time, what investigators learn about the pathogenesis of one prion disease will yield information relevant to the understanding of other prion diseases. Studies raising questions about presumed pathogenic mechanics are important in stimulating new theories and new scientific inquiry. For ex- ample, a new team of investigators studying transmissible mink encephal- opathy (TME) in hamsters showed that the agent proceeded along cranial nerves in the tongue directly to the brain, bypassing the intestinal route (Bartz et al., 20031. This finding suggests that prions could enter the body through oral lesions on the tongue, then migrate to the central nervous system through cranial nerves. Bartz and colleagues also showed that the TME agent, if injected into the brain of an uninfected hamster, would travel down to the tongue in a retrograde fashion. This finding generated some concern that BSE-infected cattle or CWD-infected cervids may harbor the infectious agent in their tongues. However, similar studies must be done to determine whether the agents of BSE and CWD appear in the tongues of infected bovines and cervids, since the aforementioned work involved a different agent (that of TME) as well as a different host animal (hamster). The possibility that cranial nerves may serve as fast tracks for the TSE agent to enter the central nervous system was recently heightened by the discovery of PrPSC in the olfactory tracts of nine patients with sCJD, sug- gesting that the olfactory pathway might serve as a portal of entry for the natural transmission of sCTD (Zanusso et al., 20031. This could have impli- cations for iatrogenic transmission of the disease. Understanding TSE pathogenesis at the cellular level essential for the
66 ADVANCING PRION SCIENCE development of effective diagnostic tools and therapeutic agents requires knowledge of the mechanisms of toxicity. TSE researchers do not know precisely how and where cellular toxicity occurs. Although most experts believe toxicity occurs at or near the cell membrane, new research in this area is challenging current theories. For example, studies by Ma and col- leagues seem to indicate that cellular toxicity may occur in the cytoso! due to an aggregation of misfolded proteins that accumulates by retrograde transport through the endoplasmic reticulum (Ma and Linguist, 2002; Ma et al., 20021. Although these studies need validation, and have even been disputed (Drisaldi et al., 2003), they open up new research possibilities. The study of TSEs should not be limited to mammalian species. Much can be learned from the study of prions found in other, nonmammalian organisms. For example, the prions found in fungi have been studied exten- sively, including two prions of the yeast Saccharomyces cerevisiae. In 1994, those two priors, fURE3] and EPSI+], were discovered to be infectious forms of their normal proteins, Ure2P and Sup35p, respectively (Wickner, 19941. The first prior-inducing domain was defined, and the protease resistance of the Ure2P in fURE3] prion strains gave the first hint of the mechanism involved (Masison and Wickner, 19951. Soon after, it was discovered that, as in mammals, multiple strains of yeast prions can exist (Derkatch et al., 19961. Ter-Avanesyan's group was the first to show that the EPSI+] prion is a self-propagating aggregation of Sup35p in viva and in vitro (Paushkin et al., 19971. Later that year, King and colleagues showed that the prion domain of Sup35p could form amyloid in vitro (King et al., 19971. Then Glover and colleaguesshowed that the full length of Sup35p could form filaments in vitro having characteristics of amyloid that were stimulated by extracts of EPSI+] cells, but not the extracts of Epsi-] cells (Grover et al., 19971. Also in 1997, a prion was found in the filamentous fungus Podospora anserine (Coustou et al., 19971. Another prion, EPIN+], of the Rnqlp protein of S. cerevisiae was reported in 2001 (Derkatch et al., 20011. Work with these prions has been highly rewarding. The experiments with S. cerevisiae described above provided the first evidence that chaper- one proteins are involved in prion propagation (Chernoff et al., 19951. Ad- ditionally, the Mksl protein was shown to be necessary for generation of the fURE3] protein (Edskes and Wickner, 20001. Also, the Ras-cyclic AMP pathway was found to negatively regulate generation of the fURE3] prion protein (Edskes and Wickner, 20001. The presence of one prion in a cell can promote the generation of an- other (Derkatch et al., 20011. Amyloid of the HET-s protein formed in vitro was shown to be infectious for fungal colonies, whereas nonspecific aggre- gates or the soluble form of the protein had no effect (Maddelein et al., 20021. Efforts to replicate this process in mammals have not yet been suc-
BASIC BIOMEDICAL RESEARCH ON TSEs 67 cessful. In addition, investigators have shown that artificial prions can be constructed by using a prion domain of one protein and a reporter domain of another (Li and Linguist, 2000~. Another landmark discovery from a study of S. cerevisiae that used tHet-s] was that prions can be advantageous to the prion host (Coustou et al., 19971. In view of the close parallels between yeast and mammalian priors, scientists could potentially use yeast to screen for prior-curing agents that might aid in the development of a TSE treatment. For example, yeast prions can be cured by growth on low concentrations of guanidine, an inhibitor of the chaperone HsplO4 (Bach et al., 2003;Jung et al., 2002; Tuite et al., 19811. Also, fragments of yeast prion proteins cure the respective prion (Edskes et al., 19991. Drosophila and Caenorhabditis elegans have also proved to be superb models for the study of a variety of cellular and molecular processes (Hariharan and Haber, 2003) and should be exploited to study prion dis- eases. These organisms have recently been used to mode! several human neurodegenerative conditions, including Parkinson's disease, tauopathies, and polyglutamine disorders. These mode! systems may clarify the basis for neurodegeneration in prion diseases, as they have in other neurodegenerative processes. PErYSIOLOGICAL FUNCTIO N OF prpC Because the primary structure of PrPSc is virtually identical to that of normal PrP, understanding as much as possible about prpC would be very
68 ADVANCING PRION SCIENCE helpful in the development of TSE diagnostic tests. A successful test must discriminate between these two closely related molecules. Moreover, under- standing the normal role of prpC may reveal associated molecules and path- ways that are appropriate detection targets. It remains unclear whether the basis for nerve cell dysfunction and death in prion disease is related to the toxicity of PrPSc, to the loss of function of prpC as a result of its conversion to PrPSc and its aggregation during a prion infection, or to other factors. Recommendation 3.1: Fund basic research to elucidate (1) the struc- tural features of priors, (2) the molecular mechanisms of prion rep- lication, (3) the mechanisms of pathogenesis of transmissible spongiform encephalopathies, and (4) the physiological function of PrPC. [Priority 11i REFERENCES Asante EA, Linehan JM, Desbruslais M, Joiner S. Gowland I, Wood AL, Welch J. Hill AF, Lloyd SE, Wadsworth JD, Collinge J. 2002. BSE prions propagate as either variant CJD- like or sporadic CJD-like prion strains in transgenic mice expressing human prion pro- tein. EMBO Journal 21(23):6358-6366. Bach S. Talarek N. Andrieu T. Vierfond JM, Mettey Y. Galons H. Dormont D, Meijer L, Cullin C, Blondel M. 2003. Isolation of drugs active against mammalian prions using a yeast-based screening assay. Nature Biotechnology 21(9):1075-1081. Bartz JC, Kincaid AK, Bessen RA. 2003. Rapid prion neuroinvasion following tongue infec- tion. Journal of Virology 77(1):583-591. Bessen RA, Kocisko DA, Raymond GJ, Nandan S. Lansbury PT, Caughey B. 1995. Non- genetic propagation of strain-specific properties of scrapie prion protein. Nature 375(6533):698-700. Caughey B. 2001. Prion protein interconversions. Philosophical Transactions of the Royal Society of London. Series B.: Biological Sciences 356(1406):197-200; Discussion, 200- 202. Caughey B. 2002. The PrP Conversion Process. Presentation to the IOM Committee on Trans- missible Spongiform Encephalopathies: Assessment of Relevant Science, Meeting II. Washington, DC. Chernoff YO, Lindquist SL, Ono B. Inge-Vechtomov SG, Liebman SW. 1995. Role of the chaperone protein HsplO4 in propagation of the yeast prior-like factor [psi+]. Science 268(5212):880-884. Cordeiro Y. Machado F. Juliano L, Juliano MA, Brentani RR, Foguel D, Silva JL. 2001. DNA converts cellular prion protein into the beta-sheet conformation and inhibits prion pep- tide aggregation. Journal of Biological Chemistry 276(52) :49400-49409. Coustou V, Deleu C, Saupe S. Begueret J. 1997. The protein product of the het-s heterokaryon incompatibility gene of the fungus Podospora anserina behaves as a prion analog. Pro- ceedings of the National Academy of Sciences of the United States of America 94(18):9773-9778. iThe committee denotes each recommendation as priority level 1, 2, or 3 based on the criteria and process described in the Introduction.
BASIC BIOMEDICAL RESEARCH ON TSEs 69 DebBurman SK, Raymond GJ, Caughey B. Lindquist S. 1997. Chaperone-supervised conver- sion of prion protein to its pro/ease-resistant form. Proceedings of the National Academy of Sciences ofthe United States of America 94(25):13938-13943. Derkatch IL, Bradley ME, Hong JY, Liebman SW. 2001. Prions affect the appearance of other priors: the story of [PIN+]. Cell 106(2):171-182. Derkatch IL, Chernoff YO, Kushnirov W. Inge-Vechtomov SG, Liebman SW. 1996. Genesis and variability of [PSI] prion factors in Saccharomyces cerevisiae. Genetics 144(4):1375- 1386. Drisaldi B. Stewart RS, Adles C, Stewart LR, Quaglio E, Biasini E, Fioriti L, Chiesa R. Harris DA. 2003. Mutant PrP is delayed in its exit from the endoplasmic reticulum, but neither wild-type nor mutant PrP undergoes retrotranslocation prior to proteasomal degrada- tion. Journal of Biological Chemistry 278(24):21732-21743. Edenhofer F. Rieger R. Famulok M, Wendler W. Weiss S. Winnacker EL. 1996. Prion protein PrPc interacts with molecular chaperones of the Hsp60 family. Journal of Virology 70(7):4724-4728. Edskes HK, Gray VT, Wickner RB. 1999. The [URE3] prion is an aggregated form of Ure2p that can be cured by overexpression of Ure2p fragments. Proceedings of the National Academy of Sciences of the United States of America 96(4):1498-1503. Edskes HK, Wickner RB. 2000. A protein required for prion generation: [URE3] induction requires the Ras-regulated Mksl protein. Proceedings of the National Academy of Sci- ences of the United States of America 97(12):6625-6629. Gabus C, Derrington E, Leblanc P. Chnaiderman J. Dormont D, Swietnicki W. Morillas M, Surewicz WK, Marc D, Nandi P. Darlix JL. 2001. The prion protein has RNA binding and chaperoning properties characteristic of nucleocapsid protein NCP7 of HIV-1. Jour- nal of Biological Chemistry 276(22):19301-19309. Glover JR, Kowal AS, Schirmer EC, Patino MM, Liu JJ, Lindquist S. 1997. Self-seeded fibers formed by Sup35, the protein determinant of [PSI+], a heritable prior-like factor of S. cerevisiae. Cell 89(5):811-819. Graner E, Mercadante AF, Zanata SM, Forlenza OV, Cabral AL, Veiga SS, Juliano MA, Roesler R. Walz R. Minetti A, Izquierdo I, Martins VR, Brentani RR. 2000. Cellular prion protein binds laminin and mediates neuritogenesis. Molecular Brain Research 76(1):85-92. Hariharan IK, Haber DA. 2003. Yeast, flies, worms, and fish in the study of human disease. New England Journal of Medicine 348(24):2457-2463. Hornshaw MP, McDermott JR, Candy JM. 1995. Copper binding to the N-terminal tandem repeat regions of mammalian and avian prion protein. Biochemical and Biophysical Re- search Communications 207(2):621-629. Jin T. Gu Y. Zanusso G. Sy M, Kumar A, Cohen M, Gambetti P. Singh N. 2000. The chaper- one protein BiP binds to a mutant prion protein and mediates its degradation by the proteasome. Journal of Biological Chemistry 275(49) :38699-38704. Jung G. Jones G. Masison DC. 2002. Amino acid residue 184 of yeast HsplO4 chaperone is critical for prior-curing by guanidine, prion propagation, and thermotolerance. Proceed- ings of the NationalAcademy of Sciences of the United States of America 99(15):9936- 9941. King CY, Tittmann P. Gross H. Gebert R. Aebi M, Wuthrich K. 1997. Prion-inducing domain 2-114 of yeast Sup35 protein transforms in vitro into amyloid-like filaments. Proceedings ofthe NationalAcademy of Sciences ofthe United States of America 94(13):6618-6622. Korth C, Stierli B. Streit P. Moser M, Schaller O. Fischer R. Schulz-Schaeffer W. Kretzschmar H. Raeber A, Braun U. Ehrensperger F. Hornemann S. Glockshuber R. Riek R. Billeter M, Wuthrich K, Oesch B. 1997. Prion (PrPSc)-specific epitope defined by a monoclonal antibody. Nature. 390(6655):74-77.
70 ADVANCING PRION SCIENCE Laws DD, Bitter HE, Liu K, Ball HE, Kaneko K, Wille H. Cohen FE, Prusiner SB, Pines A, Wemmer DE. 2001. Solid-state NMR studies of the secondary structure of a mutant prion protein fragment of 55 residues that induces neurodegeneration. Proceedings of the NationalAcademy of Sciences of the United States of America 98(20):11686-11690. Li L, Lindquist S. 2000. Creating a protein-based element of inheritance. Science 287(5453):661-664. Ma J. Lindquist S. 2002. Conversion of PrP to a self-perpetuating PrPSc-like conformation in the cytosol. Science 298(5599):1785-1788. Ma J. Wollmann R. Lindquist S. 2002. Neurotoxicity and neurodegeneration when PrP accu- mulates in the cytosol. Science 298(5599):1781-1785. Maddelein ML, Dos Reis S. Duvezin-Caubet S. Coulary-Salin B. Saupe SJ. 2002. Amyloid aggregates of the HET-s prion protein are infectious. Proceedings of the National Acad- emy of Sciences of the United States of America 99(11):7402-7407. Manson JC, Jamieson E, Baybutt H. Tuzi NL, Barron R. McConnell I, Somerville R. Ironside J. Will R. Sy MS, Melton DW, Hope J. Bostock C. 1999. A single amino acid alteration (lOlL) introduced into murine PrP dramatically alters incubation time of transmissible spongiform encephalopathy. European Molecular Biology Organization Journal 18(23):6855-6864. Masison DC, Wickner RB. 1995. Prion-inducing domain of yeast Ure2p and protease resis- tance of Ure2p in prior-containing cells. Science 270(5233):93-95. Nandi PK, Leclerc E. 1999. Polymerization of murine recombinant prion protein in nucleic acid solution. Archives of Virology 144(9):1751-1763. Paramithiotis E., Pinard M, Lawton T. LaBoissiere S. Leathers VL, Zou W. Estey LA, Lamontagne J. Lehto MT, Kondejewski LH, Francoeur GP, Papadopoulos M, Haghighat A, Spatz SJ, Head M, Will R. Ironside J. O'Rourke K, Tonelli Q. Ledebur HC, Chakrabartty A, Cashman NR. 2003. A prion protein epitope selective for the pathologi- cally misfolded conformation. Nature Medicine. 9(7):893-899. Paushkin SV, Kushnirov VV, Smirnov VN, Ter-Avanesyan MD. 1997. In vitro propagation of the prior-like state of yeast Sup35 protein. Science 277(5324):381-383. Snow AD, Wight TN, Nochlin D, Koike Y. Kimata K, DeArmond SJ, Prusiner SB. 1990. Immunolocalization of heparan sulfate proteoglycans to the prion protein amyloid plaques of Gerstmann-Straussler syndrome, Creutzieldt-Jakob disease and scrapie. Labo- ratory Investigation 63(5):601-611. Stockel J. Hartl FU. 2001. Chaperonin-mediated de novo generation of prion protein aggre- gates. Journal of Molecular Biology 313(4):861-872. Tatzelt J. Zuo J. Voellmy R. Scott M, Hartl U. Prusiner SB, Welch WJ. 1995. Scrapie prions selectively modify the stress response in neuroblastoma cells. Proceedings of the National Academy of Sciences of the United States of America 92(7):2944-2948. Tuite MF, Mundy CR, Cox BS. 1981. Agents that cause a high frequency of genetic change from [psi+] to [psi-] in Saccharomyces cerevisiae. Genetics 98(4):691-711. Weiss RH, Randour CJ. 2002. Attenuation of matrix protein secretion by antisense oligodeoxynucleotides to the cyclin kinase inhibitor p21(Wafl/Cipl). Atherosclerosis 161(1):105-112. Wemmer D. 2002. Solid State NMR and Protein Structure Analysis. Presentation to the IOM Committee on Transmissible Spongiform Encephalopathies: Assessment of Relevant Sci- ence, Meeting II. Washington, DC: National Academy Press. Wickner RB. 1994. [URE3] as an altered URE2 protein: Evidence for a prion analog in Sac- charomyces cerevisiae. Science 264(5158):566-569.
BASIC BIOMEDICAL RESEARCH ON TSEs 71 Wille H. 2002. Electron Crystallographic Analyses on 2D Crystals of the Scrapie Prion Pro- tein. Presentation to the IOM Committee on Transmissible Spongiform Encephalopa- thies: Assessment of Relevant Science, Meeting II. Washington, DC: National Academy Press. Wong C, Xiong LW, Horiuchi M, Raymond L, Wehrly K, Chesebro B. Caughey B. 2001. Sulfated glycans and elevated temperature stimulate PrP(Sc)-dependent cell-free forma- tion of pro/ease-resistant prion protein. EMBO Journal 20(3):377-386. Zahn R. Liu A, Luhrs T. Rick R. von Schroetter C, Lopez Garcia F. Billeter M, Calzolai L, Wider G. Wuthrich K. 2000. NMR solution structure of the human prion protein. Pro- ceedings ofthe NationalAcademy of Sciences ofthe United States of America 97(1):145- 150. Zanata SM, Lopes MH, Mercadante AF, Hajj GN, Chiarini LB, Nomizo R. Freitas AR, Cabral AL, Lee KS, Juliano MA, de Oliveira E, Jachieri SG, Burlingame A, Huang L, Linden R. Brentani RR, Martins VR. 2002. Stress-inducible protein 1 is a cell surface ligand for cellular prion that triggers neuroprotection. EMBO Journal 21(13):3307-3316. Zanusso G. Ferrari S. Cardone F. Zampieri P. Gelati M, Fiorini M, Farinazzo A, Gardiman M, Cavallaro T. Bentivoglio M, Righetti PG, Pocchiari M, Rizzuto N. Monaco S. 2003. Detection of pathologic prion protein in the olfactory epithelium in sporadic Creutzieldt- Jakob disease. New England Journal of Medicine 348(8):711-719.