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Home , Prion

I _r-.,-.,-.' '.'- -"'. . - ~ l,,: ". ft\'OV\-rdf '.ft:e I . 1..., The Prion

, Prions, once dismissedas an impossibility, have now gained I wide recognition as extraordinary agents that cause

: a number of infectious, genetic and spontaneousdisorders

I by Stanley B.Prusiner

'I Fifteen years ago I evokeda good the to becomeriddled with holes. .The last, often called , deal of skepticism when I pro- These ills, which can brew for years (or mad cow ,is the most worrisome. I.. posed that the infectious agents evenfor decadesin humans)are wide- GeraldA. H. Wells and John W. Wile- I, ' causingcertain degenerativedisorders spreadin . smith of the CentralVeterinary labora- I of the in ani- The most common form is( , I tory in Weybridge,England, identified I malsand. more rarely, in humansmight foundin sheepand . Afm~t~a:'an: I the conditionin 1986,after it began ! consist of and nothing else.At imals lose coordination and eventually striking cows in Great Britain. causing II the time, the notion washeretical. Dog- becomeso incapacitatedthat they can- them to becameuncoordinated and un- ! ma held that the conveyersof transmis- not stand. They also becomeirritable usuallyapprehensive. The sourceof the : sible diseasesrequired geneticmateri- and, in some cases,jeveloP.EI!~e emergingepidemic was soon traced to I al, composedof nucleicacid (DNAor itch that leadsthem to scrapeoff their a supplementthat indudedmeat I RNA),in order to establishan ~~~r hair (hencethe name"scrapie"):- and bone meal from dead .The I in a host. Evenviruses, among the sim- ~otherprton diseasesof anim~ ;1 plest microbes,rely on suchmaterial to by such names as transmissiblemink II direct synthesisof the proteinsneeded encephalopathy, for survivaland replication. of mule deer and , feline spongiform ~l_I~'11111111 . " Later,many scientistswere similarly encephalopathyand bovinespongiform r,. , dubious when my colleagues and I sug- ...:- gested that these "proteinaceous infec- 40000 >- -.,. ',~ I tious ~artides"-.or "prions," as I called ' ~ ' ~ "; -&fZ;~ the ?is.ease-~ausmgagents-could ~- w Q <"" !! ~. r." .".- ;. , derlie inhented, as well as commuru- (I) 35,000 Q A . ~. ' I cable,diseases. Such d~al b~vior was lli ~'~ ~ ,-*' .

then unknown to medical saence, And B 30,000 ~'~ we met resistanceagain when we con- s: 0 .~ i, ODS")cluded multiply that prions in (pronouncedan incredible"pree- way; 8~ 2 5,00 0 ~ - they convert normal protein molecules ~ ~ into dangerousones simply by induc- ~ 20,000 ~ ing the benign molecules to change ~ rr: theirToday, shape. however, a wealth of experi- ~ 15' 000 ~ mental and clinical data has made a ~ ::?. convincing case that we are correct on ~ 10,000 all three counts. Prionsare indeed re- ~ I I t. f;.t . I sponsiblefor trans---missibleand iDfierit- 8 5,000 ~ ~ : : Theye~ Qlsoraers caii (fIs-o of cause pro{em sporaffic conformartou.ffisease, ~ ~:g . . .

~ ~~ch neither.tran~missio~ ber:"'een ~ ~ ~ ~ ffi' 9?-rJ1' & ~ \,.' , mdiVIduals nor inhentance IS eVIdent. .?5 .?5 .?5 .?5 .'::J ,5?:J.'::J .'::J .'::J : ,",'. '.-,..~.. Moreover, there are hints that the prions SOURCE:John W Wilesmith . , ..:

causingthe diseasesexplored thus far . ~ may not be the only ones.Prions made CATn.EWERE INCINERATED to prevent of rather different may con- them" fro.m ~preading ~mad cow. dis- . tribute to other neurodegenerativedis- ease, This disorder,whi~ hasafflict~d eases that are quite prevalent in hu- ~°re. than 130,.000cattle m Grea~Bnt- . . . . am smcethe nud-1980s(graph),ls one !11ans,They nught even partiapate m of several fatal neurodegenerativedis- illnessesthat att~ckm~scles. easesof animals and humansthought The kno~ pnon diseases, all fat~, to be caused by prions-infectious pro- ~ .

ar~ sometimes referred to as spongl- teins. Studies are assessingwhether pli- 's: (orm encephalopatrues. They are so on disease can be ttansmined from cows Q.

; ..I named because they 1requently cause to people through the ingestion of beef" ~~..

CAN january 1995 ~ .~ I:" I . methods for processing sheep carcass- Creutzfeldt-Jakob disease,in contrast, . es had been changed in the late 1970s. occurs worldwide and usually becomes STANLEYB. PRUSINERis professorof Where once they would have eliminat- evident as . Most of the time neurologyand bio:chemistryat the Uni- ., ed the scrapie agent in the supplement, it appears sporadically, sttiking one per- versity of Califo~ Schoolof Medidne, . ... .' San Frandsco. He IS a member of the \ now they apparently did not. The Bnt- son m a million, typiCally around age 60. NationalAcademy of Sciencesthe Insti- '-' . ish government banned the use of ani- About 10 to 15 percent of cases are in- rote of Medidne and the 'American ;~ mal-derived feed supplements in 1988, herited, and a small number are, sadly, Academy of Arts and Sciences. He has ~ and the epidemic has probably peaked. iatrogenic-spread inadvertently by the won many awardsfor his researchinto Nevertheless, many people continue to attempt to treat some other medical pri~ns, m~st recently the Albert Lasker 1 wony that they will eventUally fall ill as a problem. Iatrogenic Creutzfeldt- Jakob Basic Me~lcal Research.A":"af~ and the result of having consumed tainted meat. disease has apparently been transmit- Pa~ Ehrlich .Aw~. This.IS his second The human prion diseases are more ted by corneal transplantation, implan- artIcle for SCIentificAmencan. obscure. has been seen only among tation of dura mater or electrodes in the the Forehighlanders of PapuaNew Guin- brain, use of contaminated surgical in- ea. They call it the "laughing death." struments, and iI\iection of growth hor- Rossella Medori of the University of B0-

o Vincent Zigas of the Australian Public mone derived from human pituitaries logna and Pierluigi Gambetti of Case Health Service and D. Carleton Gajdusek (before recombinant growth hormone Western Reserve University. of the U.S. Nati~titutes of Health became available). described it in~oting that many The two remaining human disorders In Search of the Cause highlanders became afflicted with a are Gerstmann-Striiussler-Scheinker dis- " strange, fatal disease marked by loss of ease (which is manifest as and I first became intrigued by the prion I coordinati~n (ataxia) and ?ft~. later other signs of dama~~ t~ the ce~eb~l- . diseases in 1972, when as a resident " by dementia. The affected mdiVlduals lum) and fatal familial msomma (m m neurology at the University of Cali- probably acquired kuru through ritual which dementia follows difficulty sleep- forma School of Medidne at San Fran- . : the Fore tribe reportedly ing). Both these conditions are usually dsco, I lost a patient to Creutzfeldt-Ja- '" honored the dead by eating their . inherited and typically appear in mid- kob disease. As I reviewed the scientific ,Ji The practice has since stopped, and . Fatal familial was discov- literature on that and related conditions . kuru has virtually disappeared ered only recently, by Elio Lugaresi and I learned that scrapie, Creutzfeldt-Jakob

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, SCIENTIFlCAMERICAN january 1995 49

I \ "~~;..t;;;;,~,;\,-~-- rl diseaseand kuru had all beenshown to I immediately began trying to solve duresthat denature(unfold) or degrade be transmissible'.--b in °ec . extracts0 this mystery when I set up a laboratory protein reducedinfectivity. I thus intro- <> seased brains into the brains 0 at U.C.S.F.in 1974.The first step had to duced the term "prion" to distinguish !,~ ~ animals..The were be a mechanicalone-purifying the in- this classof diseaseconveyer from vi- i - thought to be causedby a slow-acting fectious material in scrapie-infected ruses,, fungi and other known , , yet no one had managedto iso- brains so that its compositioncould be .Not long afterward,we de- late the culprit. analyzed.The task wasdaunting; many termined that scrapieprions Cj~d In the courseof reading,I cameacross investigatorshad tried and failed in the a single protein that we calle~~r I an astonishingreport in which Tikvah past. But with the optimism of youth, I "prion proteiIL" I Alper and her colleaguesat the Ham- forg~g_ah~ad(see"Prions;"by-Stanl~ Now the major questionbecame, mersmith Hospital in London suggest- ;-Prusiner;SCIENTIFIC AMERICAN, Octo- ~ere did the instructions specifying ed that the scrapie agent might lack ber 1984 . B-.".--1982my COlleal!Ues-4nd=F-111'esequence of amino acids in PrPre- I I ;wrucn usually can be de- progress,producing ex- side?Were they carriedby an undetect- grnaed by ultraVioletor ionizing ractia:- tracts of hamster brains consistingal- ed pieceof DNA that traveledwith PrP, tion. wilen me nucleicacid in extracts~ most exclusivelyof infectiousmaterial. or were they, perhaps,contained in a oficrapie-infected brains was presurn- We had, furthermore,subjected the ex- housed in the chromosomesof ably destroyedby thosetreatments, the tracts to a range of tests designedto cells? The key to this riddle was the I extracts retained their ability to trans- reveal the composition of the disease- identification in 1984 of some 15 arni- i mit scrapie. If the or~anism did lac~ causingcomponent. -no aCl~at one ena or me YrYprotein: i ; DNA and RNA.the finding would mean ~ group iaentified this short amino

I I tlmt~ not a virus or any other- AmazingDiscovery acid sequencein collaborationwith I ~wn type of-infectious agent,aJror Leroy E. Hood and his co-workers at II wmch contain genetic material-:-What, A Il our results pointed toward one the CaliforniaInstitute of Technology. Ii then, was it? Investigatorshad many j-\. startling conclusion:the infectious Knowledgeof the sequenceallowed I ideas-including,jokingly,linoleurn and agent in scrapie (and presumably in us and others to construct molecular i kryptonite-but no hard answers. me relatea ctiseases)did indeed lacK probes, or detectors,able to indicate I nuaci~~d consistedmainly, if noT whether mammalian cells carried the I ex-allsively,of prot$ Wededuced that PrP gene. With probes produced by

I i DNAand RNAwere aDsent because, like Hood's team, Bruno Oesch,working in .f Alper, we saw that proceduresknown the laboratoryof CharlesWeissmann at ~; to damagenucleic acid did not reduce the University of Zurich, showed that infectivity. And we knew protein was hamstercelIs do containa genefor PrP. an essentialcomponent because proce- At about the sametime, BruceChese-

I PRION DISEASES OF HUMANS (table), which may incubate for 30 years or more, can all cause progressive decline in cognition and motor function; hence, the dis- 'I tinctions among them are sometimes blurry. As the genetic underlying ~ familial fonDS of the diseasesare found. those disorders are likely to be identified

I ~ by their associated mutations alone. Choreographer George Balanchine (photo- ~ graph) died of sporadic Creutzfeldt-Jakob disease in 1983 at age 79.

! .']~i:f,,~i=- iTj~i~oir:.1l. ;'~~j7:!;jlj~.~~i;'~ :;:!~'=,J:li;1~.=~ r"l~oi~~,'jl;'~ill![O:~ .~[;.~.,;~_;Jc~lli~':!7i ;"i~,,1:i ~.~[~~'~l'j:~;;li '.!1~:!~,,~;.-:

Kuru Loss of coordination, often Infection (probably through Known only in highlands of Three months to one year followedby dementia cannibalism,which stopped PapuaNew Guinea;some by 1958) 2,600 cases have been identified since 1957

Creutzfeldt. Dementia, followed by loss Usually unknown (in "spora- Sporadic form: 1 person per Typically about one year; , .Jakob of coordination, although dic" disease) million worldwide range is one month to more I disease sometimesthe sequenceis than 10 years I reversed Sometimes (in 10 to 15 per- Inherited form: some 100 cent of cases) inheritance extended families have of a in the gene been identified coding for the prion protein (PrP)

Rarely, infection (as an Infectious form: about80 inadvertent consequence cases have been identified of a medical procedure)

Gerstmann. Loss of coordination, often Inheritance of a mutation Some 50 extended families Typically two to six years Straussler- followed by dementia in the PrP gene have been identified Scheinker disease

Fatal Trouble sleeping and dis- Inheritance of a mutation Nine extended families have Typically about one year familial turbanceof autonomic in the PrP gene been identified insomnia nervous system, followed by insomnia and dementia

50 SCIENTIFICAMERICAN january 1995

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boro of the Nlli RockyMountain Labo- ~ ratories made his own probes and es- ~ , tablished that mouse cells harbor the ~ gene as well. That work madeit possi- ~ V ble to isolate the geneand to establish ~ . that it resides not in prions but in the ~ of hamsters, mice, hu- mans and all other mammalsthat have been examined.What is more, most of I the time, theseanimals make PrPwith- out getting sick. One interpretation of such findings was that we had made a terrible mis- take: PrPhad nothing to do with prion diseases.Another possibility was that PrP could be produced in tWo forms, one that generateddisease and one that did not. We soon showedthe latter in- terpretation to be correct. The critical clue was the fact that the ~nd ~inf~.ct.edb~_~e~~~~g z breakdown by cellular enzymescalled ~ ~~~~~eins in .cells-are "de- ~ graae"Q~yed that if a nonnal. easily.nonthreatening I thereforesuspect- form ~ SOURCE:Fred E. Cohen of PrPexisted, it. too would be suscep~- PRIONPROTEIN (PrP) is usuallyharmless. In its benign state,its backbonetwists ble to degradation.~onal~.A. B~ ill into multiple helices(shown as spirals in the plausibleribbon modelat the left and my lab.oratorythen lden~ed this hy- as cylinders in the cartoonat the top right). PrPbecomes the infectious,"scrapie" pothetIcal -senSitIVeform. It fonn-a prion-when muchof the backbonesttetches out, fonning so-calledbeta thus becameclear that scrapie-causing strands(arrows in the hypotheticalstrUcture at the bottom right). Redsites on the PrPis a variant of a normal protein. We ribbon model of nonna! PrPhighlight positions at which substitutionof oneamino thereforecalled the normalprotein "cel- add for anotherprobably promotes folding into the scrapieforDL

""'" . lular PrP" and the infectious (protease- \., " resistant)term is now form used "scrapie \0 refer PrP." toThe the latter pro- half of the members of the affected amino add. In our dying patient, just tein molecules that constitute the pri- families.) It was this last pattern that one base pair (out of more than 750) ons causing all scrapielike diseases of drew our attention. Could it be that pri- had been exchanged for a different pair. animals and humans. ons were more unusual than we origi- The change, in turn, had altered the in- nally thought? Were they responsible formation carried by codon 102,caus- Prion Diseases Can Be Inherited for the appearance of both hereditary ing the amino add leucine to be substi- and transmissible illnesses? tuted for the amino add proline in the arly on we had hopedto use the PrP In 1988 Karen Hsiaoin my laborato- man's PrPprotein. Egeneto generatepure copiesof PrP. ry and I uncoveredsome of the earliest With the help of Tim j. Crowof North- Next,we would inject the protein mole- data showing that human prion dis- wick Park Hospital in Londonand jurg culesinto animals,secure in the knowl- easescan certainlybe inherited.We ac- Ott of Columbia University and their edge that no elusivevirus was clinging quired clones of a PrP geneobtained colleagues,we discoveredthe samemu- to them. If the injectionscaused scrapie from a man who had Gerstmann- tation in genesfrom a large numberof in the animals,we would have shown Straussler-Scheinkerdisease in his fam- patients with Gerstmann-Straussler- that protein moleculescould, as we had ily and was dying of it himself.Then we Scheinkerdisease, and we showedthat proposed, transmit disease.By 1986, comparedhis genewith PrPgenes ob- the high inddence in the affectedfami- however, we knew the plan would not tained from a healthy population and lies was statistically significant.In oth- work. For one thing, it provedvery dif- found a tiny abnormality known as a er words,we establishedgenetic linkage ficult to induce the gene to make the point mutation. betWeenthe mutation and the disease- high levels of PrPneeded for conduct- To graspthe nature of this mutation. a finding that strongly implies the mu- ing studies.For anotherthing, the pro- it helps to know somethingabout the tation is the cause.Over the past six tein that wasproduced was the normal, organizationof .Genes consist of years work by many investigatorshas cellularform. FortUnately,work on a dif- tWostrands of the DNAbuilding blocks uncovered18 mutationsin familieswith ferent problem led us to an alternative called nucleotides, which differ from inheritedprion diseases;for five of these approach for demonstratingthat pri- one anotherin the basesthey carry. The mutations,enough cases have now been ons could transmit scrapiewithout the baseson one strand combinewith the collectedto demonstrategenetic linkage. help of any accompanyingnucleic add. baseson the other strandto fonD base The discoveryof mutationsgave us a A. In many cases,the scrapielikeillness- pairs: the "rungs" on the familiar DNA way to eliminate the possibility that a ..-, es of humansseemed to occurwithout "ladder."In addition to holdingthe DNA nucleic add was traveling with prion \"./ having been spread from one host to laddertogether, these pairs spell out the proteins and directing their multiplica- another, and in some families theyap- sequence of amino adds that must be tion. ~~~~~~~~~~-f peared to be inherited (Today research- strung together to make a particular tered mice carrymg a mutated PrP gene. ers know that about 10 percent of hu- protein. Three base pairs together-a ¥ili'"e~r~s~~~ of' the~~~ n i man prion diseases are familial, felling unit called a codon-specify a single ese "transgenic amm~ls led b itself

I SCIENTIFICAMERICAN January 1995 51 /

brain extracts from the mutant would spread the infection to another host. Yet in the absenceof any evidence of a virus, this hypothesis looks to be untenable. In addition to showing that a protein can multiply and cause disease without help from nucleic adds, we have gained insight into how scrapie PrP propa- gates in cells. Many details remain to be worked out, but one aspect appears quite clear: the main difference betWeen normal PrP and scrapie PrP is confor- mational. Evidently, the scrapie protein propagates itself by contacting normal PrP molecules and somehow causing them to unfold and flip from their usu- al conformation to the scrapie shape. This change initiates a cascadein which newly converted molecules change the shape of other normal PrP molecules, and so on. These events apparently oc- - cur on a metMrane in the interior. "-~ed to think tfia"flhe Ji."faenc- es betWeen cellular and scrapie forms of PrP must be conformational after oth- er possibilities began to seem unlikely. For instance, it has long been known that the infectious form often has the same amino add sequence as the nor- mal type. Of course,molecules that start off being identical can later be chemi- cally modified in ways that alter their activity. But intensive investigations by Neil Stahl and Michael A. Baldwin in my laboratory have turned up no differ- ences of this kind.

One Protein, Two Shapes

H ow, exactly, do the structures of normal and scrapie forms of PrP differ? Studies by Keh-Ming Pan in our group indicate that the normal protein consists primarily of alpha helices, re- gions in which the protein backbone twists into a specific kind of spiral; the scrapie form, however, contains beta strands, regions in which the backbone is fully extended. Collections of these strands form beta sheets.Fred E.Cohen, . who directs another laboratory at U.C.S.F.,has used molecular modeling to try to predict the structure of the normal protein based on its amino add sequence. His calculations imply that the proteinprob~ty folds into a ~ ;)iict structure. having four helices in its ,. co e, -:: or P. po- sition that scrapie PrP can induce an al- pha-helical PrP molecule to switch to a beta-sheet form comes primarily from tWo important studies by investigators in my group. Maria Gasset learned that synthetic pep~lshort strings of rom- . ~oriesponding to three of th~

54 ,';:: D ";/ ";i" /...1;;JAAA j1fl ~~7'1 /A"////(ew'&-,, I ' """-'11'-- r ~~-.' 172 / 1/LV~--'r /~A:~~ I I . four putative alph~e!1cal reg!o~~ heimer's clumps consist of a different well. lain H. Pattison of the Agriculture Pr~can (oI(Umo beta sheets,-~k protein. The PrP laques are a usef ResearchCouncil in Compton, England, Nguyen has shown that in their beta- sign of non' ection but the seem initially called attention to this phe- ~°r:I:!~~~~~ti~~S~ at to be a m 'or irn airment. nomenon. Years ago he obtained prions ~ unpo:51:d Ul:ld-:5Ill:el struc~!! beli:. Ji1many people and ~s with p!:!?? from two separatesets of goats. One iso- \J ~~~~~~~~~J~= disease, the plaQue-s-aonot arise at ~. late made inoculated animals drowsy, w. Laugney o~o~y Mount~ Lab-1ven though we do not-yet know whereas the second made them hyper- .. f oratOnes and Peter T.Lansb~ of the much about how PrP scrapie harms active. Similarly, it is now evident that

~: ~ Massachusetts Institute of Technolo brain , we can foresee that an un- some prions cause disease quickly, ar PrP can be derstanding of the three-dimensional whereas others do so slowly. ,- 1.-(;'~1 c.?!!v~~a Wtoscrapie_Pt;P~ ~_t_:~_t_~be structure of the PrP protein will lead to .6'?f/"" oy ~ the two proleinS-together. therapies. If, for example, the four-he- ~~ ~ ystery of "Strains" -- PrP mOlecules arising from mutated !ix-bundle model of PrP is correct, drug ~~ g7nes probabl~ do not adopt the scra- developers might be a~le to design a A Ian G. Dickinson, Hugh Fraser and pie conformation as soon as they are compound that would bmd to a central j-\. Moira E. Bruce of the Institute for s}'Ilthesized. Otherwise, people carry- pocket that could be formed by the four Animal Health in Edinburgh, who have - ing mutant genes would become sick in helices. So bound, the drug would sta- examined the differential effects of var- - -- . early childhood. We suspect that muta- bilize these helices and prevent their ied isolates in mice, are among those "'/.' tions in the PrP gene render the result- conversion into beta sheets. who note that only pathogens contain- ing proteins susceptible to flipping from Another idea for therapy is inspired ing nucleic acids are known to occur in .Jl ~ - an alpha-helical to a beta-sheet shape. by research in which Weissmann and multiple strains. Hence,they and others , ~...Presumably, it takes time until one of his colleagues applied gene-targeting assert, the existence of prion "strains" r ~"'1 the molecules spontaneously flips over technology to create mice that lacked indicates the prion hypothesis must be ~ ,~~ and still more time for scrapie PrP to ac- the PrP gene and so could not make PrP. incorrect; must be at the root ..(;t,.i- -tf, ~- - cumulate and damage the brain enough By knocking out a gene and noting the of scrapie and its relatives. Yet because 6[ It. ~::::1.~'" to cause symptoms. consequences of its loss, one can often efforts to find viral nucleic acids have t! Fred Cohen and I think we might be deduce the usual functions of the gene's been unrewarding, the explanation for able to explain why the various muta- protein product. In this case, however, the differences must lie elsewhere. ~ tions that_~ve been ~ote_d in PrP genes t!!~ ~als- missin~ ~P *~played n~ One possibility is that prions can could facilitate folding mto the beta- det~tableabnormalities. If it turns out adopt multiple conformations. Folded sheet form. Many of the human muta- tfii"iPrP is truly messentIiil, then physi- in one way, a~ mi-gni convert nor- tions give rise to the substitution of one cians might one day consider delivering mal PrP to the scrapie form highly effi- for another within the four so-called antisense or antigene therapies ciently, giving rise to short incubation "' putative helices or at their borders. In- to the brains of patients with prion dis- times. Folded another way, it might 1 sertion of inco_crectaminoacids at those eases.Such therapies aim to block genes ork less efficiently. Similarly, one "con- \-, .P-osJtionsmight destabilize a helix, thus from giving rise to unwanted proteins former" might be attracted to neuronal i!!~s~g the likelih~~t:the affect:. and could potentially shut down pro- populations in one part of the brain, ~~ei&hbOrs..will.refold.into duction of cellular PrP [see "The New a beta-sheet conformation. coiiV-ersely, GeneticMedidnes," by JackS. Cohen and Hermann Schatzel in my laboratory Michael E. Hogan; SCIENm1CAMERICAN. finds that th~harn1less differe!!ces ~ December 1994). They would thereby tin~s!!!!!g the PrP ~ene Qf h!!!¥~s block PrP from propagating itself. fr,-9I?Jl1°~e.oLaPfS-and.monkeYs affect It is worth noting that the knockout _,,'mino acids lyjng-9utside of thp pro-_mice provided a welcomed opportunity llQS.ed..helicaLdomains=F-11ere"the-di to challenge the prion hypothesis. If verg~!!t amino ac!~p_rQj;!gbjy-WDuld- the animals became ill after inoculation i ll9Lnrofrnmrll¥ inf1uence-1he-~ with prions, their sickness would have o~_helicalr~j. indicated that prions could multiply even in the absence of a preexisting Treatment Ideas Emerge pool of PrP molecules. As I expected, inoculation with prions did not produce N o one knows exactly how propaga- scrapie, and no evidence of prion repli- tion of scrapie PrP damages cells. cation could be detected. In cell cultures, the conversion of nor- The enigma of how scrapie PrP multi- mal m-~~~apieTo:im:Q~~: plies and causes disease is not the only -sme-rie~ns. after which scra ie PrP puzzle starting to be solved. Another

\' accumJl~_t~s_~~UY1i!r.~!£~ long-standing question-the mystery of

~~~~~ the brain, filled how prions consisting of a single kind lysosomes could conceivably b~stan_d of protein can vary markedly in their cl~ge c~s. As the diseasedcells died, effects-is beginning to be answered as creating holes in the brain, their prions would be released to attack other cells. ... We do kn~w wi~ certainty thatcleav- HOLESIN BRAIN 1lSSUE(white spots) ~ .., age of scra le PrP is w t ro~P are a frequent feature of prion diseases. ~ \.-/ ~ua~~,!:~!. ~ulat~ as Dla~ues They give the brain a spongelikeap- ~ . ~ hr"in~ of S~~p I:!a!i~s. Tho~e pearance. This micrograph shows the ~ aggregates resemble plaques seen in cerebral cortex of a patient suffering ~ Alzheimer's disease, although the Alz- from Creutzfeldt-Jakob disease. ~ SCIENTIFICAMERICAN january 1995 55

-..CC- r:it' " - I ,. . '1 I A Persuasive Experiment j: "" S everal studies have shown that prions composedonly someof which appearsto adopt the scrapieconformation. ~ of PrPare able to convey infection from one animal to EventuallY,al1the mice displayedsym.pt,oms of br~in ~am. another. In one such experiment, the author and his col- age and died (b). Then the workers Injectedbrain tissue " leaguescreated mice carrying many copiesof a mutant PrP from the diseasedanimals into genetically altered mice - gene (0); these animals made high levels of mutant PrP, making low levels of the same mutant PrPprotein. (Such i ~ y E- o ~ ---~ MUTANT PrP /TISSUEBRAIN

ffi DISEA ~ BRAIN i - ~ ~ t ~ ~

". ~" '. 0 ". 5 - ~ - .. a MOUSE MAKING HIGH LEVELS b MUTANTMOUSE - ~ t ~ OF MUTANT PrP FALLS ILL AND DIES f ~ IS INITIALLY HEALTHY ~ 0 I .t-~

~ ~--tA-.-~-. - . ,( whereasanother might be attracted to ments, we realized that the barrIer re- ease.Their deathsmay havenothing to I~ elsewhere,thus produdng dif- ~es in theamin~add sequ~ do with the bovine epidemic,but the I; ferent symptoms.Considering that PrP PrP:themore the seQuenceof a scrapie situation bears _watching.I! may turn j can fold in at leasttwo v.:ays,it would PrP mole~e resem ~ e. . s~- Q~! tha_~~-!-~.s_m -~~P I~ not be surprising to find It can collapse qiii~1iS1iOSt":ifie~~.!!k~YjUs.~ mol-ea:ii~-~2E-!!P~_rJ'j!!1L~.q_th- I~ into other structures as well. ~~~srwm a<;:g~.p~ll.<.lisease-- e[.5J?I:_preaking_.th~~peaes_banier._1f I ..., Since the mid-1980s we have also In one oTffiO5e"Other expenmentS'.-ror ~tJ~_W~--cA~~_~q.i!_£2~-Prp_99~.ely sought insight into a phenomenon example,we examinedtransgenic mice r,e~.e.l!1;~l~~..human PrPin the critical r_~-

ceptknown refersas the to spedesthe factbarrier. that somethingThis con- incarrying additionthe toSyrian their hamsterown mousePrP gene.gene ,g!~ns!_~~~~::~~~~'~~:g~~~,~~2_b~~.~8h£!:~-~~,."..

.. makesit difficult for prions madeby Thosemice make normal forms of both pJ~_cQ~parisonof ~~.'<;9,mI!!~te__amino j: one spedesto causedisease in animals hamsterand ~ouse P,rP.When we ~oc- a9g..§:~~~!!~~ sugg~::,t. ... i of another species.The causeof this ulated the anImalsWIth mousepnons, Webegan to consId~ possIbility ~ difficulty is of considerableinterest to- they made more mouse prions. When that some parts of the PrP molecule I day becauseof the epidemic of mad we inoculated them with hamster pri- might be particularly important to the ; cow diseasein Britain.We and others ons,they madehamster prions. From spedesbarrier after a studyrelated to ! have been trying to find out whether this behavior, we learned that prions this blockadetook an odd turn. My col-

I the speciesbarrier is strong enoughto preferentially interact with cellular PrP league Glenn C. Telling had created ; prevent the spread of prion disease of homologous,or like, composition. transgenicmice carrying a h}'hrid PrP . from cows to humans. The attraction of scrapiePrP for cel- gene that consisted of human codes i lular PrP having the same sequence flanked on either side by mousecodes: i Breakingthe Barrier probablyexplains why scrapiemanaged this genegave rise to a hybrid protein. , to spreadto cowsin Englandfrom food Then he introduced brain tissue from T he barrier was discoveredby Patti- consisting of sheep tissue: sheepand patients who had died of Creutzfeldt- son,who in the 1960sfound it hard bovine PrP differ only at seven posi- Jakobdisease or Gerstmann-Straussler- to transmit scrapiebetween sheep and tions. In contrast, the sequencediffer- Scheinkerdisease into the transgenic : rodents. To determinethe causeof the encebetween human and bovinePrP is animals.Oddly enough.the animalsbe- I trouble, my colleagueMichael R. Scott large: the molecules diverge at more cameill much more frequentlyand fast- ; and I later generatedtransgenic mice than 30 positions.Because the variance er than did mice carryinga full human ! expressing the PrPgene of the Syrian is great, the likelihood of transmission PrP gene,which divergesfrom mouse hamster-that is, making the hamster from cows to peoplewould seemto be PrPat 28 positions. This outcomeim- PrP protein. The mouse gene differs low. Consistentwith this assessmentare plied that similarity in the centralregion from that of the hamstergene at 16 co- epidemiological studies by W. Bryan of the PrPmolecule may be more criti- dons out of 254. Normalmice inoculat- Matthews,a professor emeritus at the cal than it is in the other segments. ed with hamster prions rarely acquire University of Oxford. Matthewsfound The result also lent support to earli- scrapie,but the transgenicmice became no link betweenscrapie in sheepand er indications-uncovered by Shu-llan ill within about two months. the occurrenceof Creutzfeldt-Jakob dis- Yangin DeAnnond'slaboratory and AI- We thus concludedthat we had bro- easein sheep-farmingcountries. bert Taraboulosin my group-that mol- ken the speciesbarrier by inserting the On the other hand, two farmers who eculesmade by the host can influence hamster genesinto the mice.Moreover, had "mad cows"in their herdshave re- the behavior of scrapiePrP. We specu- , on the basis of this and other experi- cently died of Creutzfeldt-Jakobdis- late that in the hybrid-genestudy, a :; :. 56 SCIENm1CAMERICAN january 1995 J c". I " . . the inherited prion diseaseshave been mice were chosen as recipientsbecause scrapie PrP is most attracted to PrP modeled in ttansgenic mice canying moleculeshaving the samecomposition.) Uninoculated mice did not becomeill mutant PrPgenes. These murine repre- (indicatingthat makinglow levelsof the aberrantprotein was safe),but many of sentations of the human prion afflic- ~ the treatedones did (c). Moreover,brain tissuetransferred from the diseasedre- tions should not only extend under- " .' ' cipients to their healthy counterparts caused illness once again (d). If the aber- standing of how prions cause brain de- rant protein were unableto transmit infection, none of the inoculatedanimals generation, they should also create would havesickened. opportunities to evaluatetherapies for thesedevastating maladies.

Striking Similarities

O ngoing research may also help de- termine whether prions consisting of other proteins playa part in more common neurodegenerativeconditions, induding Alzheimer's disease, Parkin- son's disease and amyotrophic lateral sderosis. There are some marked simi- larities in all these disorders. As is true of the known prion diseases, the more C MOUSE MAKING LOW LEVELS d IDENTICAL MOUSE BECOMES ILL widespreadills .mostly ~~ spo~~di- OF MUTANT PrP BECOMES ILL AFTER RECEIVINGINOCULATION callybut sometimes"nm m families. AFTER INOCULATION FROM FIRST RECIPIENT All are also usually diseasesof middle to later life and are markedby similar :neurons degenerate, protein deposits can accumulateas plaques, mouseprotein, possiblya "" begin when the wear and tear of living and glial cells(which support and nour- normally involved in folding nascent led to a mutation of the PrPgene in at ish nerve cells) grow larger in reaction protein chains, recognizedone of the least one cell in the body. Eventually, to damage to neurons. Strikingly, in two mouse-derivedregions of the hy- the mutated protein might switch to noneof thesedisorders do white blood brid PrPprotein. This chaperonebound the scrapieform and gradually propa- cells-those everpresent warriors of the to that region and helpedto refold the gate itself, until the buildup of scrapie immune system-infiltrate the brain. lf hybrid .molecule into the s~rapiecon- PrPcrossed the thresh?ld to overt dis- a ~ wereinvolved in theseillnesses, ., formation. The chaperonedid not pro- ease.The mouse studies suggest that white cellswould be expectedto appear. vide similar help in mice making a to- at somepoint in the of the one in Recentfindings in encourage tally human PrP protein. presumably a million individuals who acquire spa- speculation that prions unrelated in because the human protein lacked a radic Creutzfeldt-Jakobdisease, cellu- amino acid sequenceto the PrPprotein binding site for the mousefactor. lar PrP may spontaneouslyconvert to could exist. ReedB. Wickner of the Nlli the scrapieform. The experimentsalso reports that a protein called Ure2p The list May Grow raisethe possibilitythat peoplewho be- might sometimeschange its conforma- comeafflicted with sporadicCreutzfeldt- tion. therebyaffecting its activity in the A n unforeseen story has recently Jakob disease overproduce PrP, but we cell. In one shape, the protein is active; j-\. emerged from studies of trans- do not yet know if, in fact, they do. in the other, it is silent. genic mice making unusually high All the known prion diseasesin hu- The collectedstudies described here amounts of normal PrPproteins. DeAr- mans havenow been modeledin mice. arguepersuasively that the prion is an mond, David Westawayin our group With our most recent work we havein- entirely new dass of infectious patha- and GeorgeA. Carlsonof the McLaugh- advertentlydeveloped an animalmodel genand that prion diseasesresult from lin Laboratoryin GreatFalls, Mont., be- for sporadicprion disease.Mice inocu- aberrations of protein conformation. came perplexed when they noted that lated with brain extracts from scrapie- Whetherchanges in protein shapeare some older transgenicmice developed infected animals and from humansaf- responsiblefor common neurodegen- an illness characterized by rigidity and flicted with Creutzfeldt-Jakob disease erative diseases, such as Alzheimer's, diminished grooming. Mlen we pursued have long provided a model for the in- remains unknown, but it is a possibility the cause,we found that making exces- fectious forms of prion disorders.And that should not be ignored. sive amounts of PrPcan eventually lead to neurodegenerationand, surprising- ly, to destruction of both musclesand FURlliERREADING widenperipheral the spectrumnerves. Theseof prion discoveries diseases SCRAP~ DISEASE IN SHEEP. ~erbert B. Par. Edited by C. Guilleminault et at. Raven . ry. Edited by D. R. OppenheImer. Academ- Press, 1994. an? are.prompting a search for h~ ic Press,1983. MOLECULARBIOLOGY OF PRION DISEASES. pnon diseases that affect the penpher- MOLECUlARBIOLOGY OF PRIONDISEASES. Spedai issue of PhilosophicalTransactions al nervous system and musdes. S. B. Prusinerin Sdence,Vol. 252, pages of the Royal Societyof London.Series B, Investigations of animals that over- 1515-1522;June 14, 1991. VoL343, No. 1306; March 29.1994. produce PrP have yielded another ben- PRIONDISEASES OF HUMANS AND ANIMALS. STRUCIURALCLUES TO PRIONREPUCA- efit as well. They offer a due as to how Editedby S.B. Prusiner, J. Collinge, J. Pow- nON.F. E. Cohen, K.-M. Pan. Z. Huang,M. ~ the sporadic form of Creutzfeldt-Jakob ell and B. Anderton. Ellis HolWood, 1992. Bald~, R. J. Fletterick an~ S. B.:Prusine! disease might arise. For a time I sus- FATAL FAMIIlAL INSOMNIA:lNHERn"ED PRI- in Sdence,VoL 264, pages :>30-:>31;April t d th t di di . h ON DISEASES, SlEEP, AND l1IE . 22.1994. pec e a spora c sease rnrg t SCIENI"IFICAMERICAN january 1995 57