A mutation equivalent to P102L in the human PrP gene, associated with Gerstmann–Straussler syndrome (GSS), has been introduced into the murine PrP gene by gene targeting. Mice homozygous for this ...mutation (101LL) showed no spontaneous transmissible spongiform encephalopathy (TSE) disease, but had incubation times dramatically different from wild‐type mice following inoculation with different TSE sources. Inoculation with GSS produced disease in 101LL mice in 288 days. Disease was transmitted from these mice to both wild‐type (226 days) and 101LL mice (148 days). In contrast, 101LL mice infected with ME7 had prolonged incubation times (338 days) compared with wild‐type mice (161 days). The 101L mutation does not, therefore, produce any spontaneous genetic disease in mice but significantly alters the incubation time of TSE infection. Additionally, a rapid TSE transmission was demonstrated despite extremely low levels of disease‐associated PrP.
Two monoclonal antibodies that specifically detect human prion protein (PrP) were developed. The epitope of both antibodies was mapped using fusion proteins of glutathione-
S-transferase and PrP ...peptides to the C-terminal region encompassing the polymorphic 219 residue. The antibodies recognized human PrP with 219Glu but not that with 219Lys. The unique property of the antibodies was utilized to determine the allelic origin of abnormal PrP deposited in the brain of a patient with Gerstmann-Straussler syndrome (GSS) with 102Leu/219Lys encoded by the same allele. Abnormal PrP was exclusively of mutant allelic origin, suggesting that 219Lys may be permissive to the formation of abnormal PrP in GSS. The antibodies may help to explore the relationship of 219Glu/Lys polymorphism to the pathogenesis of human prion diseases.
A hallmark event in transmissible spongiform encephalopathies is the conversion of the physiological prion protein into the disease-associated isoform. A natural polymorphism at codon 129 of the ...human prion gene, resulting in either methionine or valine, has profound influence on susceptibility and phenotypic expression of the disease in humans. In this study, we investigated the local propensity of synthetic peptides, corresponding to the region of the polymorphism and containing either methionine or valine, to adopt a β-sheet-rich structure similar to the pathological protein. Circular dichroism studies showed that the methionine-containing peptide has a greater propensity to adopt a β-sheet conformation in a variety of experimental conditions. The higher β-sheet tendency of this peptide was also associated with an increased ability to aggregate into amyloid-like fibrils. These results suggest that methionine at position 129 of the prion protein increases its susceptibility to switch to the abnormal conformation, in comparison with the presence of valine at the same position.
Chickens were immunized with human prion protein (PrP) peptide H25 (amino acid residues 25–49) coupled to keyhole limpet hemocyanin. From a fusion experiment using the chicken fusion partner cell ...line MuH1 and immune spleen cells, one mAb, HUC2-13, was generated which reacted with the peptide. HUC2-13 was specific for a pentapeptide (RPKPG) of the N-terminal of the peptide H25. In Western blotting analysis, the mAb reacted with PrP materials from a human Creutzfeldt–Jakob disease (CJD) case and the membrane fraction from normal murine brain, but not with the same materials pretreated with proteinase K. When compared with the HUC2-13 and the conventional mouse mAb 3F4, the background stainings using the HUC2-13 were minimal. In immunohistochemistry, the HUC2-13 stained positively with kuru plaques in brain sections from patients with Gerstmann–Straussler syndrome (GSS), and also reacted with synaptic structures of the CJD patients. However, any immunolabelings using the HUC2-13 were not observed in the section from a patient with amyotrophic lateral sclerosis (ALS) as CJD-negative control. These results indicate that the mAb HUC2-13 is a suitable tool for immunological and diagnostic analyses of prion disease in humans and other mammals.
We present here a case of variant Gerstmann-Sträussler syndrome (GSS) with a codon 105 mutation of the prion protein gene. A 57-year-old woman developed dementia and gait disturbance dissimilar to ...the spastic paraparesis that is observed in most cases with codon 105 mutation. The clinical course of the disease in this case was 12 years. The brain weighed 900 g, and the frontal lobe, pallidum and thalamus were markedly atrophic. Severe neuronal loss was observed in the deep layer of the frontal and temporal cortices, and fibrillary gliosis and a marked loss of neurons was observed in the globus pallidus, thalamus and substantia nigra. Many amyloid plaques and some ballooned neurons were present in the frontal, temporal and parietal cortices. However, no spongiform changes were seen. The cerebellum was relatively well preserved. Numerous neurofibrillary tangles (NFTs) were recognized in the cerebral cortices, and scattered NFTs were observed in the basal nucleus of Meynert, thalamus, substantia nigra, periaqueductal gray matter, raphe nuclei and locus ceruleus. The case presented here indicates the presence of variations in the pathological findings of cases with codon 105 mutation, and that the formation of cortical and brain stem NFTs might have something to do with the duration of illness and/or the degree of brain tissue destruction that had occurred.
Assessment of neurochemical markers in the frontal cortex indicates that choline acetyltransferase is significantly decreased in Alzheimer's and Gerstmann-Straussler dementias but not in Pick's ...dementia. It therefore appears that the cholinergic innervation of the cortex from the basal forebrain is intact in Pick's disease. Cortical somatostatin was decreased only in Alzheimer's disease (AD), indicating that loss of somatostatin is not a constant feature in different forms of dementia. Muscarinic binding sites were unaltered in Pick's disease and Gerstmann-Straussler syndrome but were decreased in a subpopulation of AD patients. These data suggest that in some cases of AD a significant loss of cholinoceptive neurones in the cortex is evident.
Abstract
Chickens were immunized with human prion protein (PrP) peptide H25 (amino acid residues 25–49) coupled to keyhole limpet hemocyanin. From a fusion experiment using the chicken fusion partner ...cell line MuH1 and immune spleen cells, one mAb, HUC2–13, was generated which reacted with the peptide. HUC2–13 was specific for a pentapeptide (RPKPG) of the N-terminal of the peptide H25. In Western blotting analysis, the mAb reacted with PrP materials from a human Creutzfeldt—Jakob disease (CJD) case and the membrane fraction from normal murine brain, but not with the same materials pretreated with proteinase K. When compared with the HUC2–13 and the conventional mouse mAb 3F4, the background stainings using the HUC2–13 were minimal. In immunohistochemistry, the HUC2–13 stained positively with kuru plaques in brain sections from patients with Gerstmann—Straussler syndrome (GSS), and also reacted with synaptic structures of the CJD patients. However, any immunolabelings using the HUC2–13 were not observed in the section from a patient with amyotrophic lateral sclerosis (ALS) as CJD-negative control. These results indicate that the mAb HUC2–13 is a suitable tool for immunological and diagnostic analyses of prion disease in humans and other mammals.
Prion diseases appear to be caused by the aggregation of the cellular prion protein (PrPC) into an infectious form denoted PrPSc. The in vitro aggregation of the prion protein has been extensively ...investigated, yet many of these studies utilize truncated polypeptides. Because the C‐terminal portion of PrPSc is protease‐resistant and retains infectivity, it is assumed that studies on this fragment are most relevant. The full‐length protein can be distinguished from the truncated protein because it contains a largely structured, α‐helical, C‐terminal region in addition to an N terminus that is unstructured in the absence of metal ion binding. Herein, the in vitro aggregation of a truncated portion of the prion protein (PrP 90–231) and a full‐length version (PrP 23–231) were compared. In each case, concentration‐dependent aggregation was analyzed to discern whether it proceeds by a nucleation‐dependent pathway. Both protein constructs appear to aggregate via a nucleated polymerization with a small nucleus size, yet the later steps differ. The full‐length protein forms larger aggregates than the truncated protein, indicating that the N terminus may mediate higher‐order aggregation processes. In addition, the N terminus has an influence on the assembly state of PrP before aggregation begins, causing the full‐length protein to adopt several oligomeric forms in a neutral pH buffer. Our results emphasize the importance of studying the full‐length protein in addition to the truncated forms for in vitro aggregation studies in order to make valid hypotheses about the mechanisms of prion aggregation and the distribution of aggregates in vivo.
In order to clarify the relationship between cerebellar degeneration and prion protein (PrP) accumulation, 17 cases of sporadic Creutzfeldt‐Jakob disease (CJD) and three cases of Gerstmann–Sträussler ...syndrome (GSS) were analyzed neuropathologically. Immunohistochemical staining for PrP showed synaptic‐type deposits in all of the CJD cases. The punctate PrP stains in CJD patients were almost identical with those of synaptophysin, suggesting that PrP had accumulated in the synaptic structures. Synapses damage due to PrP accumulation seemed to develop prior to granule cell loss. PrP accumulation was inversely correlated with granule cell loss. The degree of atrophy in the molecular layer was compatible with granule cell loss. The nodulus showed severe PrP accumulation when compared with other sites in the cerebellum. How‐ever, in GSS cases, the loss of Purkinje cells and dentate nucleus neurons was obvious. Further, immunohisto‐chemical staining for PrP demonstrated Kuru plaque‐type and synaptic‐type depositions in the molecular layer, the granular cell layer, and the dentate nucleus, which was different from CJD cases.
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