Altered GABA distribution in hamster brain after infection by scrapie prions
Effect of Scrapie Infection on Neuronal Nitric-oxide Synthase
Nitric oxide: an atypical neuronal messenger: Bredt lab
Identification of a promoter region in the rat prion protein gene
Evidence for native dimers of prion protein
Different allelic effects of the codons 136 and 171 in scrapie
New variant prion protein in a Japanese family with GSS
GSS disease and the Indiana kindred.
Variant GSS with the mutation of codon P105L
A single hamster site blocks conversion to protease-resistant prion
Scanning for mutations in the human prion protein open reading frame
Stop codon 145: vascular prion amyloidosis with neurofibrillary tangles
Abnormal prion mRNA in muscle fibers in SIBM and HIBM
Identification of five allelic variants of the sheep prion gene
Marsupial prion gene has two conserved domains
Different allelic effects of prion codons 136 and 171 in scrapie.
Polymorphisms of prion and onset of scrapie in Japan
Human prion: two different 24 bp deletions in atypical Alzheimer's.
Presenile dementia in a Dutch family with a novel prion insertion.
Polymorphisms of the prion gene in Italian patients with CJD
Strain variation in transmissible mink encephalopathy
Creutzfeldt-Jakob syndrome with dominant ocular symptoms
TSE species barrier effect between ferret and mink.
Chromosomal localization of PRNP, IGGC, and IGKC in mink
NADH and RNP A1 homologies claimed for prion protein

Altered GABA distribution in hamster brain after infection by scrapie prions

Lu P; Sturman JA; Bolton DC Department of Developmental Biochemistry, New York State Office of Mental Retardation and Developmental Disabilities, Staten Island 10314, USA
Brain Res, 681: 1-2, 1995 May 29, 235-41
Antibodies specific for GABA, glutamate and taurine were used to study the distribution of these amino acid neurotransmitters during the progression ofscrapie in hamsters. Immunohistochemical distribution of glutamate and taurine were unaffected in scrapie hamsters compared with controls, but thedistribution of GABA was altered by 21 days after inoculation. We found both a greater number of neurons showing GABA-like immunoreactivity and moreintense staining in those neurons in scrapie-inoculated hamster brains, particularly in the hippocampus, inferior colliculus, frontal cortex andcerebellum. The overall concentrations of aspartate, GABA, glutamate and taurine, measured in seven different brain regions by PITC-amino acid analysis,were not significantly different between normal and scrapie-affected hamsters. The subtle alteration in GABA metabolism detected in this scrapie modelsuggests that PrPSc interacts directly with a component of the GABA system.

Effect of Scrapie Infection on Neuronal Nitric-oxide Synthase in Brain

Haim Ovadia , Hana Rosenmann , Elias Shezen , Michele Halimi , Ishai Ofran and Ruth Gabizon
JBC vol 271, number 28, Issue of July 12, 1996 pp. 16856-16861
(Received for publication, October 24,Ý1995, and in revised form, February 20,Ý1996)

Nitric-oxide synthase (NOS) is responsible for the synthesis of nitric oxide which serves as a neural messenger in the central nervous system. NOS activity was markedly inhibited in brains of mice and hamsters and neuroblastoma cells infected with scrapie (ScN2a). The decrease in activity was in accordance with decreased NADPH-diaphorase-positive cells and decreased staining of NOS-positive cells demonstrated by specific anti-NOS antibodies.

However, the specific nNOS mRNA in ScN2a was elevated when compared with normal neuroblastoma cells (N2a). Immunoblotting of fractions from these cell lines with an anti-nNOS monoclonal antibody revealed a band of nNOS from N2a and two bands with a lower molecular weight in ScN2a cells. Furthermore, NOS in ScN2a cells was insoluble in nondenaturing detergents. This insolubility is one of the landmark properties of PrPSc. It is, therefore, possible that nNOS in scrapie-infected cells and brains is aberrantly folded, resulting in an insoluble and inactive enzyme.

[Full text (PDF) Version]


Atypical Neuronal Messengers: Focus on Nitric Oxide

UCSF lab of David S. Bredt, M.D., Ph.D., colleague of S. Prusiner

Nitric oxide (NO) is now recognized as one of the major neurotransmittersin the brain. Unlike traditional transmitters which interact withhigh affinity receptors in a "lock-and-key" manner, the free radical NO achieves signaling specificity by diffusing directly into a target neuron and binding to intercellular enzyme targets. Our laboratory is using molecular and genetic approachest o define the roles of NO in the functioning and development of the nervous system.

In the adult brain nitric oxide is synthesized by a single gene product, nitric oxide synthase (NOS). We find this gene to be expressed in a discrete population of interneurons in the brain which are uniquely resistant to the neurodegeneration of stroke, Huntington's disease, and excitotoxicity. Utilizing subtractive cloning as well as biochemical approaches we are defining novelt argets for NO in the brain and attempting to identify those factors which confer resistance to NO containing neurons.

Another project is focused on understanding the role of nitric oxide in development. We find that NOS is transiently expressed in differentiating and regenerating neurons throughout the body.Pharmacologic blockade of NOS in the developing brain leads to an altered expression of specific neuronal markers, indicatingt hat NO functions as a neuronal differentiation factor. Utilizing molecular and transgenic animal models we are defining the elements that regulate NOS gene expression and working to understand roles for NO in neural development and regeneration.

To characterize cellular mechanisms for NO signaling in excitable tissues we have turned our attention to skeletal muscle as a model tissue. Here, we find that NOS is tethered to a glycoprotein complex containing dystrophin, the gene product mutated in muscular dystrophy. On-going studies in this area focus on understanding the rolefor NO in neuromuscular development and disease associated with dystrophin.

Selected Publications:

Bredt, D.S., Hwang, P.M., and Snyder, S.H. (1990). Localization of nitric oxide synthase indicating a neural role for nitric oxide.Nature 347:768-70.

Bredt, D.S., Hwang, P.M., Glatt, C.E., Lowenstein, C., Reed, R.R.,and Snyder-S-H. (1991). Cloned and expressed nitric oxide synthase structurally resembles cytochrome P-450 reductase. Nature 351:714-8.

Bredt, D.S. and Snyder, S.H. (1994). Transient nitric oxide synthase neurons in the embryonic cortical plate, sensory ganglia, andolfactory epithelium. Neuron 13:301-313.

Kobzik, L., Reid, M.B., Bredt, D.S., and Stamler, J.S. (1994).Nitric oxide in skeletal muscle. Nature 372:546-548.

Brenman, J.E., Chao, D.S., Xia, H., Aldape, K., and Bredt, D.S.(1995). Nitric oxide synthase complexed with dystrophin in skeletal muscle. Cell (in press).


A 60-kDa Prion Protein with Properties of Normal and Scrapie-associated Forms

Suzette A. Priola ,Ý Byron Caughey,Ý Kathy Wehrly ,Ý Bruce Chesebro
Volume 270, Number 7, Issue of February 17, 1995 pp. 3299-3305

Scrapie is a transmissible spongiform encephalopathy of sheep and other mammals in which disease appears to be caused by the accumulation of an abnormal form of a host protein, prion protein (PrP), in the brain and other tissues. The process by which the normal protease-sensitive form of PrP is converted into the abnormal protease-resistant form is unknown. Several hypotheses predict that oligomeric forms of either the normal or abnormal PrP may act as intermediates in the conversion process. We have now identified a 60-kDa PrP derived from hamster PrP expressed in murine neuroblastoma cells. Peptide mapping studies provided evidence that the 60-kDa PrP was composed solely of PrP and, based on its molecular mass, appeared to be a PrP dimer.

The 60-kDa PrP was not dissociated under several harsh denaturing conditions, which indicated that it was covalently linked. It was similar to the disease-associated form of PrP in that it formed large aggregates. However, it resembled the normal form of PrP in that it was sensitive to proteinase K and had a short metabolic half-life. The 60-kDa PrP, therefore, had characteristics of both the normal and disease-associated forms of PrP. Formation and aggregation of the 60-kDa hamster PrP occurs in uninfected mouse neuroblastoma cells, which suggests that hamster PrP has a predisposition to aggregate even in the absence of scrapie infectivity. Similar 60-kDa PrP bands were identified in scrapie-infected hamster brain but not in uninfected brain. Therefore, a 60-kDa molecule might participate in the scrapie-associated conversion of protease-sensitive PrP to protease-resistant PrP.


Identification of a promoter region in the rat prion protein gene

Saeki K; Matsumoto Y; Matsumoto Y; Onodera T Department of Molecular Immunology, Faculty of Agriculture, University, of Tokyo, Japan
Biochem Biophys Res Commun, 219: 1, 1996 Feb 6, 47-52
We have demonstrated the presence of a rat prion protein (RaPrP) gene promoter upstream of multiple initiation sites. A 0.1-kb fragment upstream ofthe 5'-untranslated region contains specific DNA motifs characteristic of promoter elements including an AP-1 binding site, an inverted CCAAT motif andthree inverted Sp-1 binding sites. This fragment directs transcription of a luciferase reporter gene in pheochromocytoma cells (PC12) and rat gliomacells (C6), suggesting that it contains the promoter for the RaPrP gene. To more precisely localize the transcription regulatory elements in this region, aseries of 5'-deletion mutants were generated. Deletion analysis showed that an inverted CCAAt and adjoining Sp-1 binding sequences may play animportant role in transcription of the RaPrP gene.

Vascular prion cerebral amyloidosis with tau-positive neurofibrillary tangles: the phenotype of stop codon 145

Ghetti B; Piccardo P; Spillantini MG; Ichimiya Y; Porro M; Perini F; Kitamoto T; Tateishi J; Seiler C; Frangione B; Bugiani; O; Giaccone G; Prelli F;Goedert M; Dlouhy SR; Tagliavini F Department of Pathology and Laboratory Medicine, Indiana University School of Medicine, Indianapolis 46202-5120, USA
Proc Natl Acad Sci U S A, 93: 2, 1996 Jan 23, 744-8
Deposition of PrP amyloid in cerebral vessels in conjunction with neurofibrillary lesions is the neuropathologic hallmark of the dementia associated witha stop mutation at codon 145 of PRNP, the gene encoding the prion protein (PrP). In this disorder, the vascular amyloid in tissue sections and theapproximately 7.5-kDa fragment extracted from amyloid are labeled by antibodies to epitopes located in the PrP sequence including amino acids 90-147.Amyloid-laden vessels are also labeled by antibodies against the C terminus, suggesting that PrP from the normal allele is involved in the pathologicprocess. Abundant neurofibrillary lesions are present in the cerebral gray matter. They are composed of paired helical filaments, are labeled withantibodies that recognize multiple phosphorylation sites in tau protein, and are similar to those observed in Alzheimer disease. A PrP cerebral amyloidangiopathy has not been reported in diseases caused by PRNP mutations or in human transmissible spongiform encephalopathies; we propose to name thisphenotype PrP cerebral amyloid angiopathy (PrP-CAA).

Scanning for mutations in the human prion protein open reading frame

Wiese U; Wulfert M; Prusiner SB; Riesner D Institut f¸r Physikalische Biologie and Biologisch-Medizinisches Forschungszentrum, Heinrich-Heine-Universit”t D¸sseldorf, Germany
Electrophoresis, 16: 10, 1995 Oct, 1851-60
Gerstmann-Str”ussler-Scheinker syndrome (GSS), fatal familial insomnia (FFI) and familial Creutzfeldt-Jakob disease (CJD) are caused by pointmutations or octarepeat insertions in the prion protein (PrP) gene. In the present work a method was established that is appropriate for a thoroughscreening for mutations in the PrP gene and is generally applicable to screenings of any given gene. Temperature gradient gel electrophoresis (TGGE) wasmodified at two critical steps by UV cross-linking of the DNA strands and by replacing the spatial gradient with a temporal one. The shift of a DNA band intemporal temperature gradient gel electrophoresis (tTGGE) due to a mutation can be calculated as a function of the position of the mutation in the sequence.Appropriate DNA fragments were selected for polymerase chain reaction (PCR) amplification and analysis by tTGGE on the basis that the predicted bandshifts amount to more than 10% of the migration distance for all possible mutations. The accuracy of the prediction was tested experimentally with tenknown mutations in the human PrP gene, and quantitative agreement between theory and experiment was achieved. Thus, this screening method is also asuitable means to verify the absence of mutations in a given gene segment.

A single hamster prion amino acid blocks conversion to protease-resistant prion

Priola SA; Chesebro B Laboratory of Persistent Viral Diseases, National Institute of Allergy and Infectious Diseases, Rocky Mountain Laboratories, Hamilton, Montana 59840,USA
J Virol, 69: 12, 1995 Dec, 7754-8
Neurodegeneration caused by the transmissible spongiform encephalopathies is associated with the conversion of a normal host protein, PrP-sen, into anabnormal aggregated protease-resistant form, PrP-res. In scrapie-infected mouse neuroblastoma cells, mouse PrP-sen is converted into PrP-res butrecombinant hamster PrP-sen expressed in these cells is not. In the present studies, recombinant hamster/mouse PrP-sen molecules were expressed inthese scrapie-infected cells to define specific PrP amino acid residues critical for the conversion to PrP-res. The results showed that homology to theregion of mouse PrP-sen from amino acid residues 112 to 138 was required for conversion of recombinant PrP-sen to PrP-res in scrapie-infectedmouse cells. Furthermore, a single hamster-specific PrP amino acid at residue 138 could inhibit the conversion of the recombinant PrP-sen intoPrP-res. The data are consistent with studies in humans which show that specific amino acid residue changes within PrP can influence diseasepathogenesis and transmission of transmissible spongiform encephalopathies across species barriers.

Variant GSS with the mutation of codon P105L

Kubo M; Nishimura T; Shikata E; Kokubun Y; Takasu T 
Department of Neurology, Nihon University School of Medicine. 
Rinsho Shinkeigaku 35: 873-7 (1995) 
Here we present a case of variant GSS disease with mutations in codons 105 and 129 in a prion protein. The patient was a 54-year-old male, who developed weakness in the lower limbs and spastic, wide-based gait at the age of 46 years. Subsequently he developed dementia and spastic quadriplegia at the age of 49. He had marked pseudobulbar palsy at the age of 50 and became bed-ridden in decorticated posture at teh age of 53. CT and MRI examinations revealed marked atrophy of the frontal and temporal lobes, but the occipital lobes and the cerebellum were spared. His sister had been reported by Amano, et al. in 1992 as a case of variant GSS syndrome, who had very similar clinical features, and had numerous prion protein positive plaques in her cerebral cortex at the time of autopsy. His sister was confirmed to have the same mutations in a prion protein as the present case in later genetic studies.

New variant prion protein in a Japanese family with Gerstmann-Str”ussler syndrome

Furukawa H; Kitamoto T; Tanaka Y; Tateishi J Department of Neurology, Faculty of Medicine, Kyushu University, Fukuoka, Japan
Brain Res Mol Brain Res, 30: 2, 1995 Jun, 385-8
We found novel variants in the open reading frame of the prion protein (PrP) gene in a family with Gerstmann-Str”ussler syndrome (GSS). Codon219Lys variant is a normal polymorphism which we found recently. Some GSS cases were identified with codon 102 mutation (proline to leucine) andcodon 219Lys polymorphism. While two families had a codon 102 mutation and codon 219Lys polymorphism in different alleles, 4 patients in one familyhad both in the same allele. The clinicopathological features of these 4 patients were clearly different from previously reported GSS patients with codon102 mutation. These cases should be reported as a new variant of GSS.

Different allelic effects of the codons 136 and 171 of the prion gene in sheep with scrapie

Clouscard C; Beaudry P; Elsen JM; Milan D; Dussaucy M; Bounneau C; Schelcher F; Chatelain J; Launay JM; Laplanche JL Centre INRA, Castanet-Tolosan, France
J Gen Virol, 76 ( Pt 8):1995 Aug, 2097-101
Scrapie is a transmissible degenerative disease of the central nervous system occurring naturally in sheep. It belongs to the group of prion diseases alsoaffecting man in which an abnormal isoform of the host-encoded prion protein (PrP) accumulating in the brain is responsible for neuronal death. Threemain polymorphisms have been described in the sheep PrP gene, at positions 136, 154 and 171. A strong association between susceptibility/resistance tonatural scrapie and a dimorphism at codon 136 of the ovine PrP gene has been reported in several breeds, including Romanov. This dimorphism, however,is not found in all scrapie-affected breeds. We have compared the PrP genotypes of Lacaune sheep obtained from enzootically affected flocks with those ofapparently healthy sheep. A third variant at codon 171 was also evidenced. The results were compared with those obtained in a single experimentalRomanov flock orally challenged with nematode parasites in which scrapie suddenly appeared and killed 80% of the sheep. We present evidence that, evenin different epizootological circumstances, the major genetic factor controlling the susceptibility/resistance to natural scrapie in sheep, is representedby codon 171 genotype of the PrP gene. We also suggest that a modification of the allelic effects of codon 136 can occur in heavily infected animals.

Abnormal accumulation of prion mRNA in muscle fibers in SIBM and HIBM

Sarkozi E; Askanas V; Engel WK 
Department of Neurology, University of Southern California School of Medicine, Los Angeles. 

Am J Pathol, 145: 6, 1994 Dec, 1280-4 
Sporadic inclusion-body myositis is the most common progressive muscle disease of older patients. The muscle biopsy demonstrates mononuclear cell inflammation and vacuolated muscle fibers containing paired helical filaments and 6 to 10-nm fibrils, both resembling those of Alzheimer brain, and Congo-red positivity. Hereditary inclusion-body myopathy designates patients cytopathologically similar but without inflammation. In both muscle diseases, prion, and several proteins characteristic of Alzheimer brain--eg, beta-amyloid protein and hyperphosphorylated tau (which normally are expressed mainly in neurons), and apolipoprotein E--are abnormally accumulated in vacuolated muscle fibers, by unknown mechanisms. We now demonstrate in both muscle diseases that prion mRNA is strongly expressed in the vacuolated muscle fibers, which suggests that their accumulated prion protein results, at least partly, from increased gene expression. This, to our knowledge, is the first demonstration of abnormally increased prion mRNA in human disease. Another novel finding is the increased prion mRNA in human muscle macrophages, and both increased prion protein and prion mRNA in regenerating muscle fibers. The latter indicates that prion may play a role in human muscle development.
Identification of five allelic variants of the sheep PrP gene and natural scrapie.
Belt PB; Muileman IH; Schreuder BE; Bos-de Ruijter J; Gielkens AL; Smits MA 
Department of Molecular Biology, DLO-Institute for Animal Science and Health (ID-DLO), Lelystad, The Netherlands. 

J Gen Virol, 76 ( Pt 3):1995 Mar, 509-17 
Scrapie is a fatal neurodegenerative disease of sheep that belongs to the group of prion diseases found in humans and animals. The host encoded prion protein (PrP) plays a central role in the disease process. In the PrP genes of man, mice and sheep, polymorphisms have been found that are associated with disease susceptibility and pathogenesis. We have used denaturing gradient gel electrophoresis (DGGE) to detect polymorphisms in the sheep PrP gene. In addition to the already described polymorphisms at codons 136, 154 and 171, we identified a hitherto unknown G --> T transition at codon 171. This transition is responsible for a glutamine to histidine substitution. An arginine to glutamine substitution at this position has been described previously. DGGE allowed us to identify five different combinations of these polymorphisms within the PrP gene representing five allelic variants, which were cloned and sequenced. Based on the triplet sequences present at codons 136, 154 and 171 these allelic variants were designated PrPVRQ, PrPARR, PrPARQ, PrPARH and PrPAHQ. To determine the association of these allelic variants with natural scrapie, we screened 34 scrapie affected and 91 healthy control sheep of the Texel breed for the presence of these allelic variants. In these two groups, the five variants gave rise to 13 different genotypes. The distribution of the allelic variants among both groups showed marked differences. The PrPVRQ variant was present with high frequency in scrapie affected sheep, whereas the PrPARR variant was almost exclusively present in the healthy group. Two other variants, PrPARQ and PrPARH, were found in both groups with equal frequencies. The data obtained suggest modulation of disease susceptibility in these Texel sheep by at least five different PrP allelic variants, with the PrPVRQ and PrPARR alleles acting in a dominant, but opposite fashion over the PrPARQ and PrPARH alleles. The frequency of the PrPAHQ variant was too low to draw any conclusions.

Gerstmann-Str”ussler-Scheinker disease and the Indiana kindred.

Ghetti B; Dlouhy SR; Giaccone G; Bugiani O; Frangione B; Farlow MR; Tagliavini F 
Department of Pathology and Laboratory Medicine, Indiana University, School of Medicine, Indianapolis 46202-5120, USA. 

Brain Pathol, 5: 1, 1995 Jan, 61-75 
Gerstmann-Str”ussler-Scheinker disease is an autosomal dominant disorder with a wide spectrum of clinical presentations including ataxia, spastic paraparesis, extrapyramidal signs, and dementia. The patients present with symptoms in the third to sixth decade of life and the mean duration of illness is five years. Mutations at codons 102, 105, 117, 145, 198 and 217 of the open reading frame of the prion protein gene have been associated with GSS disease. As a result of the mutations, a substitution at the corresponding residues of the prion protein occurs, or as in the case of the STOP mutation at codon 145, a truncated protein is produced. Neuropathologically, the common denominator is a cerebral prion protein amyloidosis; however, there is significant variability in the pattern of amyloid deposition in regions of the central nervous system among reported families. Amyloidosis coexists with severe spongiform degeneration in patients with the mutation at codon 102, and with neurofibrillary degeneration in the patients with mutation at codons 145, 198 and 217. The development of a transmissible spongiform encephalopathy in animals inoculated with brain tissue from affected subjects with mutation at codon 102 suggests that in some forms of genetically-determined Gerstmann-Str”ussler-Scheinker disease, and particularly those characterized by severe spongiosis, amyloidogenesis and production of an infectious "agent" occur concomitantly via mechanisms that are only partially understood.

Marsupial prion gene has two domains conserved in mammalian prion proteins.

Windl O; Dempster M; Estibeiro P; Lathe R 
Centre for Genome Research, University of Edinburgh, UK. 

Gene, 159: 2, 1995 Jul 4, 181-6 
The normal function of the pathogenicity-related protein, PrP (or prion protein), is unknown. To shed light on functionally important domains, we have characterized a candidate marsupial PrP gene. The deduced marsupial PrP has an overall identity of about 80% to eutherian PrP at the amino acid (aa) level. This similarity is not equally distributed and two regions (aa 118-142 and 177-223) are particularly highly conserved. In contrast, a repeat region in the N-terminal half of the marsupial PrP shows dipeptide inserts not described in other PrP. Another particular feature of the marsupial gene is the lack of a continuous ORF on the antisense strand, as is found in most eutherian PrP. We propose that antisense ORFs found in other species are artefactual. Comparison with all known PrP argues that the molecule characterised is the true marsupial PrP orthologue.

Different allelic effects of prion codons 136 and 171 in scrapie.

Clouscard C; Beaudry P; Elsen JM; Milan D; Dussaucy M; Bounneau C; Schelcher F; Chatelain J; Launay JM; Laplanche JL 
Centre INRA, Castanet-Tolosan, France. 

J Gen Virol, 76 ( Pt 8):1995 Aug, 2097-101 
Scrapie is a transmissible degenerative disease of the central nervous system occurring naturally in sheep. It belongs to the group of prion diseases also affecting man in which an abnormal isoform of the host-encoded prion protein (PrP) accumulating in the brain is responsible for neuronal death. Three main polymorphisms have been described in the sheep PrP gene, at positions 136, 154 and 171. A strong association between susceptibility/resistance to natural scrapie and a dimorphism at codon 136 of the ovine PrP gene has been reported in several breeds, including Romanov. This dimorphism, however, is not found in all scrapie-affected breeds. We have compared the PrP genotypes of Lacaune sheep obtained from enzootically affected flocks with those of apparently healthy sheep. A third variant at codon 171 was also evidenced. The results were compared with those obtained in a single experimental Romanov flock orally challenged with nematode parasites in which scrapie suddenly appeared and killed 80% of the sheep. We present evidence that, even in different epizootological circumstances, the major genetic factor controlling the susceptibility/resistance to natural scrapie in sheep, is represented by codon 171 genotype of the PrP gene. We also suggest that a modification of the allelic effects of codon 136 can occur in heavily infected animals.
Polymorphisms of prion and onset of scrapie in Japan

Polymorphisms of prion and onset of scrapie in Japan.

Ikeda T; Horiuchi M; Ishiguro N; Muramatsu Y; Kai-Uwe GD; Shinagawa M 
Department of Veterinary Public Health, Obihiro University of Agriculture and Veterinary Medicine, Hokkaido, Japan. 

J Gen Virol, 76 ( Pt 10):1995 Oct, 2577-81 
We investigated the relationships between amino acid polymorphisms of the prion protein (PrP), restriction fragment length polymorphisms (RFLP) of the PrP gene and the incidence of natural scrapie in Japan. Six variant alleles of the PrP gene were found in healthy sheep. Based on the substitutions at codons 112, 136, 154 and 171, these allelic variants were designated PrPMARQ, PrPTARQ, PrPMVRQ, PrPMAHQ, PrPMARR and PrPMARH. Each RFLP haplotype (e1h2, e1h2 or e3h1) consisted bo multiple alleles including PrPMARQ. Three of these variant alleles were found in scrapie-affected Suffolk sheep. PrPMARQ was associated with high disease incidence, PrPTARQ and PrPMARR were associated with low disease incidence. We found that one scrapie-affected Suffolk was homozygous for PrPMARR and four PrPSc-positive Suffolks carried PrPMVRQ. Both of two scrapie-affected Corriedales and two out of three scrapie-affected cross-breeds between Suffolk and Corriedale carried PrPMARH, suggesting that this allele associates with high incidence of scrapie in Corriedale and its cross-breeds.

Human prion: two different 24 bp deletions in an atypical Alzheimer's disease family.

Perry RT; Go RC; Harrell LE; Acton RT 
University of Alabama at Birmingham, USA. 

Am J Med Genet, 60: 1, 1995 Feb 27, 12-8 
Alzheimer's disease (AD) is a progressive, degenerative neurological disorder of the central nervous system. AD is the fourth leading cause of death in elderly persons 65 years or older in Western industrialized societies. The etiology of AD is unknown, but clinical, pathological, epidemiological, and molecular investigations suggest it is etiologically heterogeneous. Mutations in the amyloid protein are rare and segregate with the disease in a few early-onset familial AD (FAD) families. Similarities between AD and the unconventional viral (UCV) diseases, and between the amyloid and prion proteins, implicate the human prion protein gene (PRNP) as another candidate gene. Single strand conformation polymorphism (SSCP) analysis was used to screen for mutations at this locus in 82 AD patients from 54 families (30 FAD), vs. 39 age-matched controls. A 24-bp deletion around codon 68 that codes for one of five Gly-Pro rich octarepeats was identified in two affected sibs and one offspring of one late-onset FAD family. Two other affected sibs, three unaffected sibs, and three offspring from this family, in addition to one sporadic AD patient and three age-matched controls, were heterozygous for another octarepeat deletion located around codon 82. Two of the four affected sibs had features of PD, including one who was autopsy-verified AD and PD. Although these deletions were found infrequently in other AD patients and controls, they appear to be a rare polymorphism that is segregating in this FAD family. It does not appear that mutations at the PRNP locus are frequently associated with AD in this population.

Presenile dementia in a Dutch family with a novel insertion in the prion gene.

van Gool WA; Hensels GW; Hoogerwaard EM; Wiezer JH; Wesseling P; Bolhuis PA 
Department of Neurology, University of Amsterdam, The Netherlands. 

Brain, 118 ( Pt 6):1995 Dec, 1565-71 
The clinical features and disease course of six patients from a family with autosomal dominant inheritance of presenile dementia and a hypokinetic syndrome are described. In the past, these patients have carried diagnoses of Pick's disease, Huntington's disease, Parkinson-dementia, and one patient was described as suffering from a 'peculiar type of presenile dementia' in a case report. In the two cases examined, the most distinctive neuropathological features were extensive globular deposits of periodic acid-Schiff plus diastase (PAS)-positive material, having tinctural properties of amyloid only to a limited degree, in the cerebellum and cerebral cortex. These globules stained positively with antibodies against prion protein. Southern blot of MspI-digested genomic DNA showed an abnormal band of approximately 950 bp in all three patients from which material was available. Direct sequencing of the abnormal allele revealed an insert consisting of eight extra 24-nucleotide repeats in the patients, which was absent in a healthy first degree relative who was considered well beyond the age of onset of symptoms in this family. The nucleotide sequence of the abnormal insert of 192 bp was different from that of a previously described insert of equal length. Adding to previous descriptions of mutations in the prion protein gene, this report emphasizes the clinical, neuropathological and genetic heterogeneity of inherited prion disease.

Strain variation in transmissible mink encephalopathy

Bessen RA; Marsh RF 
Laboratory of Persistent Viral Diseases, Rocky Mountain Laboratories, National Institute of Allergy and Infectious Diseases, Hamilton, Montana 59840
 J Virol, 68: 12, 1994 Dec, 7859-68 
The molecular basis of strain variation in scrapie diseases is unknown. The only identified component of the agent is the posttranslationally modified host prion protein (PrPSc). The biochemical and physical properties of PrP from two strains of transmissible mink encephalopathy (TME), called hyper (HY) and drowsy (DY), were compared to investigate if PrP heterogeneity could account for strain diversity. The degradation rate of PrPTME digested with proteinase K was found to be strain specific and correlated with inactivation of the TME titer. Edman protein sequencing revealed that the major N-terminal end of HY PrPTME commenced at least 10 amino acid residues prior to that of DY PrPTME after digestion with proteinase K. Analysis of the brain distribution of PrPTME exhibited a strain-specific pattern and localization of PrPTME to the perikarya of specific neuron populations. Our findings are consistent with HY and DY PrPTME having distinct protein conformations and/or strain-specific ligand interactions that influence PrPTME properties. We propose that PrPTME conformation could play a role in targeting TME strains to different neuron populations in which strain-specific formation occurs. These data are consistent with the idea that PrPTME protein structure determines the molecular basis of strain variation

Creutzfeldt-Jakob syndrome with dominant ocular symptoms

Babincov· O; SĚblov· O; Selinger P 
OcnĚ oddelenĚ FNsP na Bulovce, Praha
 Cesk Oftalmol, 50: 6, 1994 Dec, 371-6 
A rare case of a 46-year-old man with Creutzfeldt-Jacob's disease, its causal agent being an unconventional virus (prion). Ocular symptoms were the first which made the patient see a doctor. The course of the disease with a fatal outcome caused great diagnostic difficulties among a number of specialists

Polymorphisms of the prion gene in Italian patients with CJD

Salvatore M; Genuardi M; Petraroli R; Masullo C; D'Alessandro M; Pocchiari M 
Laboratory of Virology, Istituto Superiore di Sanitý, Rome, Italy. 
Hum Genet, 94: 4, 1994 Oct, 375-9, erratum in May;95(5):605] 
Abstract Creutzfeldt-Jakob disease (CJD) is a transmissible neurodegenerative disorder characterized by the accumulation of the amyloid protein PrP in the CNS. Two coding polymorphisms of the PrP gene (PRNP) are a methionine (Met) to valine (Val) change at codon 129, and a deletion in the octapeptide coding region. In the United Kingdom, homozygosity at codon 129 appears to be associated with a predisposition to develop CJD. However, in Japan, where allelic frequencies and genotype distribution are significantly different, such an association has not been demonstrated. To determine whether such deletion(s) or codon 129 polymorphisms of PRNP predispose to the development of CJD in Italian patients, 31 sporadic CJD patients with no known PRNP mutations, and 186 unrelated control subjects were studied. Genotypic frequencies at codon 129 in these Italian CJD patients revealed a significant excess of methionine alleles, and a different genotype distribution in comparison with the normal Italian population. Deletions of a 24-bp segment located in the PrP octapeptide coding region were found in two control subjects, but in none of the sporadic CJD patients. These data suggest that Met homozygosity at codon 129 may contribute, with other environmental or endogenous factors, to CJD development.

TSE species barrier effect between ferret and mink.

Bartz JC; McKenzie DI; Bessen RA; Marsh RF; Aiken JM 
Department of Animal Health and Biomedical Sciences, University of Wisconsin, Madison 53706. 

J Gen Virol, 75 ( Pt 11):1994 Nov, 2947-53 
Experimental infection of transmissible mink encephalopathy (TME) in two closely related mustelids, black ferret (Mustela putorius furo) and mink (Mustela visa), revealed differences in their susceptibility to the TME agent. When challenged with the Stetsonville TME agent, a longer incubation period was observed in ferrets (28 to 38 months) than mink (4 months). Western blot analysis of ferret and mink prion proteins (PrP) demonstrated no detectable differences between the proteins. Northern blot analysis of ferret brain RNA indicated that PrP mRNA abundance is similar in infected and uninfected individuals. We amplified the PrP coding region from ferret DNA using the polymerase chain reaction and compared the deduced amino acid sequence of the ferret PrP gene with the mink PrP gene. This comparison revealed six silent base changes and two amino acid changes between mink and ferret: Phe-->Lys at codon 179 and Arg-->Gln at codon 224, respectively. These changes may indicate the region of PrP that is responsible for the species barrier effect between mink and ferret.

Chromosomal localization of PRNP, IGGC, ALDB, HOXB, GPT, and IGKC in mink

Khlebodarova TM; Malchenko SN; Matveeva NM; Pack SD; Sokolova OV; Alabiev BY; Belousov ES; Peremislov VV; Nayakshin AM; Brusgaard; K; et al 
Laboratory of Developmental Genetics, Academy of Russia, Siberian Department, Novosibirsk, Russia. 
Mamm Genome, 6: 10, 1995 Oct, 705-9 
Chromosomal localization of the genes for gamma- and kappa-immunoglobulins (IGGC and IGKC, respectively), aldolase B (ALDB), prion protein (PRNP), homeo box B (HOXB), and glutamate pyruvate transaminase (GPT) were determined with the use of mink-rodent hybrid cells. Analysis of segregation of the mink markers and chromosomes in these hybrid cells allowed us to assign the gene for HOXB to Chromosome (Chr) 8, IGGC to Chr 10, PRNP and IGKC to Chr 11, ALDB to Chr 12, and GPT to Chr 14 in mink. Furthermore, using a set of mink-mouse hybrid cells carrying fragments of mink Chr 8 of different sizes, we assigned the gene for HOXB to the pter-p26 region of the short arm of Chr 8. Comparative mapping of the genes of mink, human, and mouse, as well as other mammalian species, demonstrated that the mink genes HOXB, PRNP, ALDB, and IGGC are members of a conserved region shared by many mammalian species in common; the IGKC gene is a member of a conserved region common to carnivores and primates, not rodents; the GPT gene is a member of a syntenic gene group probably unique to the Mustelidae family or carnivores.

Prion function and dysfunction: a structure-based scenario.

Radulescu RT; Korth C 
Med Hypotheses 46: 225-8 (1996) 
PrP contains a putative nicotinamide adenine dinucleotide (NADH)-binding site. Moreover, the PrP octarepeats reveal homology to the nucleic acid-binding and strand-annealing octarepeats of mammalian heterogeneous ribonucleoprotein (RNP) A1. Therefore, PrP may have NADH-dependent oxidoreductase activity as well as A1-like functions such as nucleic acid annealing and splicing. Moreover, we propose that infectious prions are propagated through a dynamic molecular symbiosis between a ribozyme-like nucleic acid and a conformational isomer of the RNP-like prion protein. Thus, our model has important implications for the understanding and treatment of prion diseases.

[The authors probably err in reporting homology with hnRNP A1. Any sequence with high G content will necessarily align fairly well with the octapeptide repeat. This result was previously observed and homology rejected in the case of C. elegans DNA hybridizing to prion probe. Multiple protein isoforms for the hnRNP genes do arise by the insertion of short peptide sequences in the glycine-rich domain, in a manner reminiscent of octapeptide repeat insertions-- webmaster]

H.sapiens mRNA for hnRNP core protein A1.
SWISS-PROT:P09651
                    MSKSESPKEPEQLRKLFIGGLSFETTDESLRSHFEQWGTLTDCV
                     VMRDPNTKRSRGFGFVTYATVEEVDAAMNARPHKVDGRVVEPKRAVSREDSQRPGAHL
                     TVKKIFVGGIKEDTEEHHLRDYFEQYGKIEVIEIMTDRGSGKKRGFAFVTFDDHDSVD
                     KIVIQKYHTVNGHNCEVRKALSKQEMASASSSQRGRSGSGNFGGGRGGGFGGNDNFGR
                     GGNFSGRGGFGGSRGGGGYGGSGDGYNGFGNDGSNFGGGGSYNDFGNYNNQSSNFGPM
                     KGGNFGGRSSGPYGGGGQYFAKPRNQGGYGGSSSSSSYGSGRRF