Molecular genetic studies of Creutzfeldt-Jakob disease

Genetic study of over 200 cases of Creutzfeldt-Jakob disease (CJD), Gerstmann-Sträussler-Scheinker disease (GSS), fatal familial insomnia (FFI), and kuru have brought a reliable body of evidence that the familial forms of CJD and all known cases of GSS and FFI are linked to germline mutations in the coding region of the PRNP gene on chromosome 20, either point substitutions or expansion of the number of repeat units. No pathogenic mutations have so far been found in sporadic or infectious forms of CJD, although there are features of genetic predisposition in iatrogenic CJD and kuru. In FFI and familial CJD, clinically and pathologically distinct syndromes that are both linked to the 178^Asp→Asn substitution, phenotypic expression is dependent on a polymorphism at codon 129. Synthetic peptides homologous to several regions of PrP spontaneously form insoluble amyloid fibrils with unique morphological characteristics and polymerization tendencies. Peptides homologous to mutated regions of PrP exhibit enhanced fibrilogenic properties and, if mixed with the wild-type peptide, produce even more abundant and larger fibrous aggregates. A similar process in vivo may lead to amyloid accumulation and disease, and transmission of “baby fibrils” may induce disease in other hosts.     

Prion protein gene sequence and chronic wasting disease susceptibility in white-tailed deer (Odocoileus virginianus)

ABSTRACT

The sequence of the prion protein gene (PRNP) affects susceptibility to spongiform encephalopathies, or prion diseases in many species. In white-tailed deer, both coding and non-coding single nucleotide polymorphisms have been identified in this gene that correlate to chronic wasting disease (CWD) susceptibility. Previous studies examined individual nucleotide or amino acid mutations; here we examine all nucleotide polymorphisms and their combined effects on CWD. A 626 bp region of PRNP was examined from 703 free-ranging white-tailed deer. Deer were sampled between 2002 and 2010 by hunter harvest or government culling in Illinois and Wisconsin. Fourteen variable nucleotide positions were identified (4 new and 10 previously reported). We identified 68 diplotypes comprised of 24 predicted haplotypes, with the most common diplotype occurring in 123 individuals. Diplotypes that were found exclusively among positive or negative animals were rare, each occurring in less than 1% of the deer studied. Only one haplotype (C, odds ratio 0.240) and 2 diplotypes (AC and BC, odds ratios of 0.161 and 0.108 respectively) has significant associations with CWD resistance. Each contains mutations (one synonymous nucleotide 555C/T and one nonsynonymous nucleotide 286G/A) at positions reported to be significantly associated with reduced CWD susceptibility. Results suggest that deer populations with higher frequencies of haplotype C or diplotypes AC and BC might have a reduced risk for CWD infection – while populations with lower frequencies may have higher risk for infection. Understanding the genetic basis of CWD has improved our ability to assess herd susceptibility and direct management efforts within CWD infected areas.     

Genotype patterns and characteristics of PRNP in the Korean population

Creutzfeldt-Jakob disease (CJD), included in the human transmissible spongiform encephalopathies (TSE), is widely known to be caused by an abnormal accumulation of misfolding prion protein in the brain. Human prion protein gene (PRNP) is mapped in chromosome 20p13 and many single nucleotide polymorphisms (SNPs) in PRNP have been discovered. However, the functionality of SNPs in PRNP is yet unclear, though several SNPs have been known as important mutation related with susceptibility human prion diseases. Our aim is to identify specific genotype patterns and characteristics in the PRNP genomic region and to understand susceptibility among Korean discriminated prion disease patients, suspected CJD patients and the KARE data group. Here, we have researched genotypes and SNPs allele frequencies in PRNP in discriminated prion disease patients group (n = 22), suspected prion diseases patients group (n = 163) and the Korea Association REsource (KARE) data group (n = 296) in Korea. The sequencing regions were promoter region, exon1 and exon2 with their junction parts among 481 samples. A total of 25 SNPs were shown in this study. Nucleotide frequencies of all SNPs are exceedingly tended to bias toward dominant homozygote types except in rs2756271. Genotype frequencies at codon 129 and 219 coding region were similar with previous studies in Korea and Japan. Pathogenic mutations such as 102P/L, 200E/K and 203V/I were observed in discriminated CJD patients group, and 180V/I and 232M/R were shown in suspected prion disease patients group and the KARE data group. A total of 10 SNPs were newly identified, six in the promoter region, one in exon 2 and three in the 3′ UTR. The strong and unique linkage disequilibrium (D' = 0.94, r² = 0.89) was observed between rs57633656 and rs1800014 which is located in codon 219 coding region. We expect that these data can be provided to determine specific susceptibility and a protective factor of prion diseases not only in Koreans but also in East Asians.     

Prion protein gene polymorphisms in sheep in the state of Paraná, Brazil

To determine the polymorphisms of the prion protein gene in sheep from the state of Paraná, Brazil, 323 animals of meat breeds (Suffolk, Hampshire Down, Texel, Ile de France, Dorper, Dorset, Santa Inês and crossbreds) were genotyped by restriction fragment length polymorphism (RFLP) analysis. The most frequent allele was ARQ, with a frequency of 0.61, followed by ARR (0.30). VRQ and AHQ alleles were present at very low frequencies (0.13 and 0.05 respectively), and the ARH allele was not found. Seven genotypes were identified (ARR/ARR, ARR/ARQ, ARQ/ARQ, ARR/VRQ, ARR/AHQ, ARQ/VRQ and ARQ/AHQ), of which ARQ/ARQ was the most frequent (0.41). The Santa Inês breed and crossbred animals showed the highest genotypic variability.     

The first report of genetic variations in the chicken prion protein gene

ABSTRACT

Abnormal structural changes of the prion protein (PrP) are the cause of prion disease in a wide range of mammals. However, spontaneous infected cases have not been reported in chicken. Genetic variations of the prion protein gene (PRNP) may impact susceptibility to prion disease but have not been investigated thus far. Because an investigation of the chicken PRNP can improve the understanding of characteristics related to resistance to prion disease, research on the chicken PRNP is highly desirable. In this study, we investigated the genetic characteristics of the chicken PRNP gene. For this, we performed direct sequencing in 106 Dekalb White chickens and analyzed the genotype and allele frequencies of chicken PRNP gene. We found two insertion and deletion polymorphisms in the chicken PRNP: c.163_180delAACCCAGGGTACCCCCAT and c.268_269insC. The former is a U2 hexapeptide deletion polymorphism. Of the 106 samples, 13 (12.26%) were insertion homozygotes, 89 (83.96%) were heterozygotes, and 4 (3.77%) were deletion homozygotes in c.163_180delAACCCAGGGTACCCCCAT. In the c.268_269insC polymorphism, 102 (96.23%) were deletion homozygotes, and 4 (3.77%) were heterozygotes. Insertion homozygotes of c.268_269insC were not detected. Two polymorphisms were in perfect linkage disequilibrium (LD) with a D’ value of 1.0, and three haplotypes were identified. Furthermore, PROVEAN evaluates 163_180delAACCCAGGGTACCCCCAT as ‘deleterious’ with a score of – 13.173. Furthermore, single nucleotide polymorphisms (SNPs) in the open reading frame (ORF) of the PRNP gene were not found in the chicken. To the best of our knowledge, this was the first report on the genetic variations of the chicken PRNP gene.     

PRNP haplotype distribution in Moroccan goats

Susceptibility/resistance to scrapie in sheep and goats is influenced by host prion protein gene ( PRNP ) genotype. In this study, we report the analysis of prion protein gene polymorphisms in 137 goats of two Moroccan populations: D'man and Chaouni. We found seven previously described amino acid polymorphisms at codons 37, 127, 137, 142, 154, 222 and 240, as well as three known silent mutations. In addition, we identified three new allelic variants: 101R and 139S in D'man goats and 145D in D'man and Chaouni individuals. The high frequency of the resistant allele 154H could offer genetic protection against the disease to the analysed animals. A total of 12 haplotypes and 28 genotypes were found, the distribution of which shows significant differences between both groups. Moreover, haplotype frequencies were compared with bibliographic data showing that the haplotype distribution of PRNP in Moroccan populations is genetically similar to Southern Italian and Greek goats.     

Analysis of polymorphic microsatellites within the bovine and ovine prion protein (PRNP) genes

Twenty-four microsatellite sites with at least three repeats were found in the bovine prion protein gene (PRNP) and 23 in the ovine PRNP gene. Eight microsatellite sites were polymorphic in cattle and six in sheep with up to 10 alleles per site. In many cases allelic DNA fragments had variants in microsatellite sites and in flanking regions. Distances between microsatellite sites in eight genes from cattle and sheep occurred on average every 0.9 kb. The numerous polymorphic microsatellite sites will improve analysis of phylogenetic origin of different PRNP alleles and trait association studies for bovine spongiform encephalopathy (BSE) and scrapie.     

Genotype-phenotype analysis in inherited prion disease with eight octapeptide repeat insertional mutation

A minority of inherited prion diseases (IPD) are caused by four to 12 extra octapeptide repeat insertions (OPRI) in the prion protein gene (PRNP). Only four families affected by IPD with 8-OPRI have been reported, one of them was a three-generation Swedish kindred in which four of seven affected subjects had chorea which was initially attributed to Huntington’s disease (HD). Following the exclusion of HD, this phenotype was labeled Huntington disease-like 1 (HDL1). Here, we provide an update on the Swedish 8-OPRI family, describe the clinical features of one of its affected members with video-recordings, compare the four 8-OPRI families and study the effect of PRNP polymorphic codon 129 and gender on phenotype. Surprisingly, the Swedish kindred displayed the longest survival of all of the 8-OPRI families with a mean of 15.1 years from onset of symptoms. Subjects with PRNP polymorphic codon 129M in the mutated allele had significantly earlier age of onset, longer survival and earlier age of death than 129V subjects. Homozygous 129MM had earlier age of onset than 129VV. Females had a significantly earlier age of onset and earlier age of death than males. Up to 50% of variability in age of onset was conferred by the combined effect of PRNP polymorphic codon 129 and gender. An inverse correlation between early age of onset and long survival was found for this mutation.     

Bison PRNP genotyping and potential association with Brucella spp. seroprevalence

The implication that host cellular prion protein (PrP(C)) may function as a cell surface receptor and/or portal protein for Brucella abortus in mice prompted an evaluation of nucleotide and amino acid variation within exon 3 of the prion protein gene (PRNP) for six US bison populations. A non-synonymous single nucleotide polymorphism (T50C), resulting in the predicted amino acid replacement M17T (Met --> Thr), was identified in each population. To date, no variation (T50; Met) has been detected at the corresponding exon 3 nucleotide and/or amino acid position for domestic cattle. Notably, 80% (20 of 25) of the Yellowstone National Park bison possessing the C/C genotype were Brucella spp. seropositive, representing a significant (P = 0.021) association between seropositivity and the C/C genotypic class. Moreover, significant differences in the distribution of PRNP exon 3 alleles and genotypes were detected between Yellowstone National Park bison and three bison populations that were either founded from seronegative stock or previously subjected to test-and-slaughter management to eradicate brucellosis. Unlike domestic cattle, no indel polymorphisms were detected within the corresponding regions of the putative bison PRNP promoter, intron 1, octapeptide repeat region or 3'-untranslated region for any population examined. This study provides the first evidence of a potential association between nucleotide variation within PRNP exon 3 and the presence of Brucella spp. antibodies in bison, implicating PrP(C) in the natural resistance of bison to brucellosis infection.