Mutations In Prion Protein Gene Research Articles

Fatal familial insomnia with an unusual prion protein deposition pattern: an autopsy report with an experimental transmission study

We recently performed a post-mortem examination on a Japanese patient who had a prion protein gene mutation responsible for fatal familial insomnia (FFI). The patient initially developed cerebellar ataxia, but finally demonstrated insomnia, hyperkinetic delirium, autonomic signs and myoclonus in the late stage of the illness. Histological examination revealed marked neuronal loss in the thalamus and inferior olivary nucleus; however, prion protein (PrP) deposition was not proved in these lesions by immunohistochemistry. Instead, PrP deposition and spongiform change were both conspicuous within the cerebral cortex, whereas particular PrP deposition was also observed within the cerebellar cortex. The abnormal protease-resistant PrP (PrP(res)) molecules in the cerebral cortex of this case revealed PrP(res) type 2 pattern and were compatible with those of FFI cases, but the transmission study demonstrated that a pathogen in this case was different from that in a case with classical FFI. By inoculation with homogenate made from the cerebral cortex, the disease was transmitted to mice, and neuropathological features that were distinguishable from those previously reported were noted. These findings indicate the possibility that a discrete pathogen was involved in the disease in this case. We suggest that not only the genotype of the PrP gene and some other as yet unknown genetic factors, but also the variation in pathogen strains might be responsible for the varying clinical and pathological features of this disease.

P3-382 Insertional mutation in the prion protein gene presenting with Schizophrenia

P3-382 Insertional mutation in the prion protein gene presenting with Schizophrenia

High capacity and low cost detection of prion protein gene variant alleles by denaturing HPLC

Mutations in the human prion protein gene ( PRNP) are responsible for hereditary diseases called transmissible spongiform encephalopathies (TSE) and a polymorphic site at codon 129 determines sensitivity to infectious forms of these maladies. More recently, codon 129 has been related to cognition performance in the elderly, in Alzheimer disease (AD) and in Down syndrome. Furthermore, a rare polymorphism at codon 171 was described in 23% of patients with mesial temporal lobe epilepsy related to hippocampal sclerosis (MTLE-HS), the most common form of surgically remediable epileptic syndrome. Thus, a method that permits fast and efficient screening of PRNP mutations and polymorphisms in patients, in high risk populations, and in family members is desirable. In the present study, we established the conditions for analysis of the PRNP open reading frame using denaturing high-performance liquid chromatography (DHPLC), whereby unpurified PCR products were subjected to denaturing and reannealing steps leading to heteroduplex formation. We described specific profiles for the PRNP polymorphisms at codons 129 (M/V), 117 (A/A silent), 219 (E/K), 171 (N/S), and the octarepeat deletion using amplified DNA from 562 samples. The chromatograms for TSE-associated mutations at codons 102 (P/L), 183 (T/A), and 210 (V/I) were also determined. Specificity of the DHPLC profile for each PRNP variant allele was confirmed in 100% of the samples by direct and cloned DNA sequencing in addition to endonuclease digestion when applicable. Therefore, the present study shows that DHPLC is a rapid, highly accurate and efficient technique for the detection of PRNP genetic variants.

5 Structural Studies of Prion Proteins

Although many aspects of prion disease biology are unorthodox, perhaps the most fundamental paradox is posed by the coexistence of inherited, sporadic, and infectious forms of these diseases. Sensible molecular mechanisms for prion propagation must explain all three forms of prion diseases in a manner that is compatible with the formidable and ever-increasing array of experimental data derived from histopathologic, biochemical, biophysical, human genetic, and transgenetic studies. In this chapter, we explore the phenomenologic constraints on models of prion replication with a specific emphasis on biophysical studies of prion protein structures. We examine how an inherited disease can also present as a sporadic or infectious illness in the context of the structural data on PrPs that are currently available. Since the first version of this chapter was written, much more has been learned about prion biology. Many ideas have been reinforced and structural models have been revised. Exciting new advances toward the treatment of prion disease have resulted. THEORY OF PRION DISEASES The inherited prion diseases include Gerstmann-Straussler-Scheinker disease (GSS), familial Creutzfeldt-Jakob disease (fCJD), and fatal familial insomnia (FFI). These patients present with characteristic clinical and neuropathologic findings as early as their third or fourth decade of life, and their family histories are compatible with an autosomal dominant pattern of inheritance (Chapter 14). Molecular genetic studies argue that these diseases are caused by mutations in the prion protein (PrP) gene based on high LOD scores for 5 of the 30 known mutations (Hsiao et al. 1989; Dlouhy et al. 1992;...

Differential Expression of Metallothioneins in Human Prion Diseases

We herein report an immunohistochemical and a Western blot analysis on metal/free radical chelating proteins, metallothioneins (MTs; MT-I/II and MT-III), in the brains of human prion disease patients with or without prion protein gene mutation and polymorphism. Irrespective of the isoforms of MTs, the immunoreaction was detected in the cytoplasm and processes of the astrocytes in the cerebral cortex and white matter in normal controls and prion disease brains. Although the immunoreactivities for MTs in Creutzfeldt-Jakob disease (CJD) brains varied from case to case, they were generally dependent upon the disease duration. In CJD patients with a relatively long disease course, the immunoreaction for both MT-I/II and MT-III in the astrocytes was significantly reduced, and this finding was not modified by the genotypes of the patients. On the other hand, in patients with Gerstmann-Sträussler-Scheinker syndrome, MT-I/II immunoreactivity in the astrocytes was exclusively reduced, while the immunoreaction for MT-III was relatively well preserved. Especially the astrocytes in the vicinities of the kuru plaques exhibited a weak or no immunoreaction even for MTs but a strong immunoreaction for glial fibrillary acidic protein. A quantitative Western blot analysis also revealed that MT-I/II protein accumulated in CJD brain with a short disease duration, whereas MT-III in CJD brain with a long disease duration was statistically significantly reduced in comparison to the normal brains. These findings suggest that the protein expression of MTs in the astrocytes is thus regulated differentially among human prion diseases and modified locally by such abnormal prion protein depositions as kuru plaques.

Transmissible Spongiform Encephalopathies in Humans

Creutzfeldt-Jakob disease (CJD), the first transmissible spongiform encephalopathy (TSE) to be described in humans, occurs in a sporadic, familial, or iatrogenic form. Other TSEs in humans, shown to be associated with specific prion protein gene mutations, have been reported in different parts of the world. These TSEs compose a heterogeneous group of familial diseases that traditionally have been classified as familial CJD, Gerstmann-Sträussler-Scheinker syndrome, or fatal familial insomnia. In 1996, a newly recognized variant form of CJD among young patients (median age, 28 years) with unusual clinical features and a unique neuropathologic profile was reported in the United Kingdom. In the absence of known CJD risk factors or prion protein gene abnormalities, the UK government concluded that the clustering of these cases may represent transmission to humans of the agent causing bovine spongiform encephalopathy. Additional epidemiologic and recent laboratory data strongly support the UK government's conclusion.

Prion Protein Aggregation Reverted by Low Temperature in Transfected Cells Carrying a Prion Protein Gene Mutation

Prion diseases are characterized by the conversion of the normal cellular prion protein (PrPC), a glycoprotein that is anchored to the cell membrane by a glycosylphosphatidylinositol moiety, into an isoform that is protease-resistant (PrPres) and pathogenic. In inherited prion diseases, mutations in the prion protein (PrPM) engender the conversion of PrPM into PrPres. We developed a cell model of Gerstmann-Sträussler-Scheinker disease, a neurodegenerative condition characterized by PrPM-containing amyloid deposits and neuronal loss, by expressing the Gerstmann-Sträussler-Scheinker haplotype Q217R-129V in human neuroblastoma cells. By comparison to PrPC, this genotype results in the following alterations of PrPM: 1) expression of an aberrant form lacking the glycosylphosphatidylinositol anchor, 2) increased aggregation and protease resistance, and 3) impaired transport to the cell surface. Most of these alterations are temperature-sensitive, indicating that they are due to misfolding of PrPM.

Scanning for mutations in the human prion protein open reading frame by temporal temperature gradient gel electrophoresis.

Gerstmann-Sträussler-Scheinker syndrome (GSS), fatal familial insomnia (FFI) and familial Creutzfeldt-Jakob disease (CJD) are caused by point mutations or octarepeat insertions in the prion protein (PrP) gene. In the present work a method was established that is appropriate for a thorough screening for mutations in the PrP gene and is generally applicable to screenings of any given gene. Temperature gradient gel electrophoresis (TGGE) was modified 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 in temporal 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 band shifts amount to more than 10% of the migration distance for all possible mutations. The accuracy of the prediction was tested experimentally with ten known mutations in the human PrP gene, and quantitative agreement between theory and experiment was achieved. Thus, this screening method is also a suitable means to verify the absence of mutations in a given gene segment.

Codon 178 mutation of the human prion protein gene in a German family (Backer family): sequencing data from 72-year-old celloidin-embedded brain tissue

Familial Creutzfeldt-Jakob disease was first described in a family from northern Germany in the 1920s (Backer family). PCR amplification of DNA extracted from brain tissue embedded in celloidin 72 years ago shows a GAC to AAC substitution at codon 178 of the prion protein gene. This mutation is associated with fatal familial insomnia and familial Creutzfeldt-Jakob disease in a number of families of diverse ethnic background.

Codon 178 mutation of the human prion protein gene in a German family (Backer family): sequencing data from 72-year-old celloidin-embedded brain tissue

Codon 178 mutation of the human prion protein gene in a German family (Backer family): sequencing data from 72-year-old celloidin-embedded brain tissue

The apolipoprotein E alleles as major susceptibility factors for Creutzfeldt-Jakob disease

SummaryCreutzfeldt-Jakob disease (CJD) is a rapid progressive mental and neurological disorder characterised by dementia and is both infectious and genetic. Pathogenic mutations and a predisposing polymorphism have been described in the prion protein gene and an abnormal prion product accumulates in the brain of affected patients. Apolipoprotein E (APOE), a protein of lipid metabolism, has been detected in some prion protein deposits. This ApoE exists as three common isoforms, coded by specific allele (∈2, ∈3, ∈4). The presence of at least one ∈4 allele was described as a major risk factor for Alzheimer's disease, another neurodegenerative disorder. From a series of 61 patients with CJD we found that ∈4 allele of the APOE gene was a risk factor for the disease (p < 0·01). This association was observed in both definite and probable cases, and for patients with and without prion protein gene mutations. Moreover, in affected subjects, ∈2 allele of the APOE gene delayed occurrence of death (p<0·01) independently of other known mutations influencing the phenotype of the disease.These effects on neurodegenerative disease associated with APOE alleles suggest a strong involvement of the APOE locus in brain metabolism.

Human prion diseases

The prion diseases, sometimes referred to as the "transmissible spongiform encephalopathies," include kuru, Creutzfeldt-Jakob disease, and Gerstmann-Sträussler-Scheinker disease of humans as well as scrapie and bovine spongiform encephalopathy of animals. For many years, the prion diseases were thought to be caused by viruses despite intriguing evidence to the contrary. The unique characteristic common to all of these disorders, whether sporadic, dominantly inherited, or acquired by infection, is that they involve the aberrant metabolism of the prion protein (PrP). In many cases, the cellular prion protein is converted into the scrapie isoform by a posttranslational process that involves a conformational change. Often, the human prion diseases are transmissible to experimental animals and all of the inherited prion diseases segregate with PrP gene mutations.

The neuropathological phenotype in transgenic mice expressing different prion protein constructs.

Neuropathologic examination of transgenic (Tg) mice which express different prion protein (PrP) constructs is essential because spongiform (vacuolar) degeneration of neurons, the distribution of PrPSc and whether PrP amyloid plaques form are the phenotypes of prion diseases. In Tg models of experimental scrapie, it was found that all of the parameters that define prion isolates ('strains') can be manipulated by changing the structure of PrP. In those studies, further evidence that PrPSc causes scrapie neuropathology and determines scrapie incubation time was obtained. In addition, the distribution of PrPSc in the brain was unique for each prion isolate. The implications of these findings are first, that prion isolates target different neuron populations for synthesis of nascent pathogenic PrPSc and, secondly, that prion isolate diversity is determined by neurons. In Tg mice which express mutated PrP mimicking human prion protein gene mutations linked to familial prion diseases, the neuropathological changes have been faithfully reproduced. A new age-related, neuromuscular disorder has also been identified in uninfected Tg mice which overexpress wild-type PrPc. All of the findings with different PrP constructs plus the absence of scrapie pathology in PrP null mice are the strongest argument that the prion protein is the main etiologic and pathogenic factor of prion disorders.

Detecting prion protein gene mutations by denaturing gradient gel electrophoresis.

Mutations of the prion protein (PrP) gene are present in patients with Gerstmann-Sträussler-Scheinker syndrome (GSS), familial Creutzfeldt-Jakob disease (CJD), and fatal familial insomnia (FFI). We developed a denaturing gradient gel electrophoresis (DGGE) strategy that readily identifies point mutations in the PrP coding sequence. By comparison with appropriate controls, haplotypes often may be deduced. This method permits samples from many patients with GSS, CJD, as well as patients with unusual degenerative neurologic disorders, to be screened rapidly, sensitively, and inexpensively for the presence of known and novel PrP mutations. We illustrate the sensitivity of this approach by reporting 2 novel polymorphisms in the PrP coding sequence.

Cerebral amyloid in human prion disease.

The clinical and neuropathological features of 21 patients with prion disease were reviewed with special reference to the morphology and immunoreactivity of cerebral amyloid. Six cases had a mutation at codon 102 of the prion protein (PrP) gene and in these the characteristic pathology was the formation of multicentric amyloid plaques which were stained with PrP antibody, whereas spongiform changes were absent in one and minimal in two. In one case, with a 216 base-pair insertion in the PrP gene, there was no spongiform encephalopathy (SE) but cerebellar amyloid was a prominent feature of the pathology. One case with a PrP gene mutation at codon 200 had severe SE but no amyloid. Two iatrogenic and 11 sporadic cases had SE and some form of amyloid was identified in all but three. Amyloid angiopathy and senile neuritic plaques, which stained with antibody to beta-protein, were present in familial as well as in sporadic cases, including some who were rather young to be regarded as having Alzheimer's disease. Cerebellar amyloid and degeneration of granule and Purkinje cells were particularly common findings in sporadic as well as in genetically determined cases. This study serves to emphasize the association between prion disease and amyloid deposition in the brain. PrP is a component of some amyloid plaques in a high proportion of cases with inherited prion disease but may also be found in cases of sporadic SE without known mutations or base-pair insertions in the PrP gene.

A “presymptomatic” case of Gerstmann-Sträussler-Scheinker disease from the indiana kindred has cerebellar deposits of prion protein amyloid and a mutation in the prion protein gene

A “presymptomatic” case of Gerstmann-Sträussler-Scheinker disease from the indiana kindred has cerebellar deposits of prion protein amyloid and a mutation in the prion protein gene

Insertions in the prion protein gene in atypical dementias

A number of mutations have been demonstrated in the open reading frame (ORF) of the prion protein (PrP) gene in patients with familial Creutzfeldt-Jakob disease or Gerstmann-Sträussler syndrome. On the basis of detecting an insertion in the ORF of the PrP gene in a patient originally suspected to be suffering from familial Alzheimer-type dementia, we screened 101 individuals with atypical dementias for the known PrP gene mutations. Insertions were found in five individuals, whereas none of the other reported mutations in the PrP gene was detected in the present study. One of the five insertions was larger than that described previously, suggesting that the individuals with these mutations are unlikely to be all lineally related and that insertions in the PrP gene may not be uncommon in prion diseases.