Abstract
In neurodegenerative conditions such as Alzheimer's and prion disease it has been shown that host genetic background can have a significant effect on susceptibility. Indeed, human genome-wide association studies (GWAS) have implicated several candidate genes. Understanding such genetic susceptibility is relevant to risks of developing variant CJD (vCJD) in populations exposed to bovine spongiform encephalopathy (BSE) and understanding mechanisms of neurodegeneration. In mice, aspects of prion disease susceptibility can be modelled by examining the incubation period following experimental inoculation. Quantitative trait linkage studies have already identified multiple candidate genes; however, it is also possible to take an individual candidate gene approach. Rarb and Stmn2 were selected as candidates based on the known association with vCJD. Because of the increasing overlap described between prion and Alzheimer's diseases we also chose Clu, Picalm and Cr1, which were identified as part of Alzheimer's disease GWAS. Clusterin (Clu) was considered to be of particular interest as it has already been implicated in prion disease. Approximately 1,000 heterogeneous stock (HS) mice were inoculated intra-cerebrally with Chandler/RML prions and incubation times were recorded. Candidate genes were evaluated by sequencing the whole transcript including exon-intron boundaries and potential promoters in the parental lines of the HS mice. Representative SNPs were genotyped in the HS mice. No SNPs were identified in Cr1 and no statistical association with incubation time was seen for Clu (P = 0.96) and Picalm (P = 0.91). Significant associations were seen for both Stmn2 (P = 0.04) and Rarb (P = 0.0005), however, this was only highly significant for Rarb. This data provides significant further support for a role for the Rarb region of Mmu14 and Stmn2 in prion disease.
Highlights
Prion diseases, known as transmissible spongiform encephalopathies, are fatal neurodegenerative disorders of both humans and animals
No coding changes were observed in the Retinoic Acid Receptor Beta (Rarb) ORF and no clear functions were obviously associated with the non-coding SNPs, we looked at Rarb mRNA expression levels in mouse brains from the parental lines of the heterogeneous stock (HS) to see whether expression level could be associated with genotype. mRNA was prepared from whole brains of 6–8 week old mice and Rarb transcripts were quantified using real-time RT-PCR (Table S5 in File S1)
Failure to replicate human susceptibility factors in mouse may occur for several reasons, including prion strain differences and the fact that the mouse model we are using here only looks at one aspect of susceptibility, incubation time following intra-cerebral challenge
Summary
Known as transmissible spongiform encephalopathies, are fatal neurodegenerative disorders of both humans and animals. The first human genome wide association study (GWAS) for variant CJD (vCJD) confirmed this by implicating an additional two independent genetic loci [6]. The GWAS study of Mead et al found a genome-wide significant association between vCJD and the SNP rs6794719 which maps upstream of the gene RARB (retinoic acid receptor beta) on chromosome 3 (P = 1.961027) This was replicated in a small sample of patients with iatrogenic CJD (P = 0.03) but not with sporadic CJD or kuru. We have previously used a heterogeneous stock (HS) of mice inoculated with RML mouse adapted scrapie prions to fine map and identify prion disease incubation time candidate genes [16,17] This resource can be used in a genome-wide association study or to look at individual candidate genes [18]. To further verify our candidate genes we looked at mRNA expression levels by real time RT-PCR in both normal and endstage RML prion-infected mice
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