THE STATUS OF TUPAIA SPLENDIDULA GRAY (PRIMATES TUPAIIDAE)

abstract not availabl

Next generation sequencing of CLU, PICALM and CR1: pitfalls and potential solutions

CLU, PICALM and CR1 were identified as genetic risk factors for late onset Alzheimer’s disease (AD) in two large genome wide association studies (GWAS) published in 2009, but the variants that convey this alteration in disease risk, and how the genes relate to AD pathology is yet to be discovered. A next generation sequencing (NGS) project was conducted targeting CLU, CR1 and PICALM, in 96 AD samples (8 pools of 12), in an attempt to discover rare variants within these AD associated genes. Inclusion of repetitive regions in the design of the SureSelect capture lead to significant issues in alignment of the data, leading to poor specificity and a lower than expected depth of coverage. A strong positive correlation (0.964, p<0.001) was seen between NGS and 1000 genome project frequency estimates. Of the ~170 “novel” variants detected in the genes, seven SNPs, all of which were present in multiple sample pools, were selected for validation by Sanger sequencing. Two SNPs were successfully validated by this method, and shown to be genuine variants, while five failed validation. These spurious SNP calls occurred as a result of the presence of small indels and mononucleotide repeats, indicating such features should be regarded with caution, and validation via an independent method is important for NGS variant calls

Next generation sequencing of CLU, PICALM and CR1: pitfalls and potential solutions

CLU, PICALM and CR1 were identified as genetic risk factors for late onset Alzheimer’s disease (AD) in two large genome wide association studies (GWAS) published in 2009, but the variants that convey this alteration in disease risk, and how the genes relate to AD pathology is yet to be discovered. A next generation sequencing (NGS) project was conducted targeting CLU, CR1 and PICALM, in 96 AD samples (8 pools of 12), in an attempt to discover rare variants within these AD associated genes. Inclusion of repetitive regions in the design of the SureSelect capture lead to significant issues in alignment of the data, leading to poor specificity and a lower than expected depth of coverage. A strong positive correlation (0.964, p<0.001) was seen between NGS and 1000 genome project frequency estimates. Of the ~170 “novel” variants detected in the genes, seven SNPs, all of which were present in multiple sample pools, were selected for validation by Sanger sequencing. Two SNPs were successfully validated by this method, and shown to be genuine variants, while five failed validation. These spurious SNP calls occurred as a result of the presence of small indels and mononucleotide repeats, indicating such features should be regarded with caution, and validation via an independent method is important for NGS variant calls

Influence of coding variability in APP-Aß metabolism genes in sporadic Alzheimer's disease

The cerebral deposition of Aß42, a neurotoxic proteolitic derivate of amyloid precursor protein (APP), is a central event in Alzheimer’s disease (AD)(Amyloid hypothesis). Given the key role of APP-Aß metabolism in AD pathogenesis, we selected 29 genes involved in APP processing, Aß degradation and clearance. We then used exome and genome sequencing to investigate the single independent (single-variant association test) and cumulative (gene-based association test) effect of coding variants in these genes as potential susceptibility factors for AD, in a cohort composed of 435 sporadic and mainly late-onset AD cases and 801 elderly controls from North America and the UK. Our study shows that common coding variability in these genes does not play a major role for the disease development. In the single-variant association analysis, the main hits, which were nominally significant, were found to be very rare coding variants (MAF 0.3%-0.8%) that map to genes involved in APP processing (MEP1B), trafficking and recycling (SORL1), Aß extracellular degradation (ACE) and clearance (LRP1). Moreover, four genes (ECE1, LYZ, TTR and MME) have been found as nominally associated to AD using c-alpha and SKAT tests. We suggest that Aβ degradation and clearance, rather than Aβ production, may play a crucial role in the etiology of sporadic AD

Influence of coding variability in APP-Aβ metabolism genes in sporadic Alzheimer’s disease

This is the final version of the article. Available from the publisher via the DOI in this record.The cerebral deposition of Aβ42, a neurotoxic proteolytic derivate of amyloid precursor protein (APP), is a central event in Alzheimer's disease (AD)(Amyloid hypothesis). Given the key role of APP-Aβ metabolism in AD pathogenesis, we selected 29 genes involved in APP processing, Aβ degradation and clearance. We then used exome and genome sequencing to investigate the single independent (single-variant association test) and cumulative (gene-based association test) effect of coding variants in these genes as potential susceptibility factors for AD, in a cohort composed of 332 sporadic and mainly late-onset AD cases and 676 elderly controls from North America and the UK. Our study shows that common coding variability in these genes does not play a major role for the disease development. In the single-variant association analysis, the main hits, none of which statistically significant after multiple testing correction (1.9e-4<p-value<0.05), were found to be rare coding variants (0.009%<MAF<1.4%) with moderate to strong effect size (1.84<OR<Inf) that map to genes mainly involved in Aβ extracellular degradation (TTR, ACE), clearance (LRP1) and APP trafficking and recycling (SORL1). These results were partially replicated in the gene-based analysis (c-alpha and SKAT tests), that reports ECE1, LYZ and TTR as nominally associated to AD (1.7e-3 <p-value <0.05). In concert with previous studies, we suggest that 1) common coding variability in APP-Aβ genes is not a critical factor for AD development and 2) Aβ degradation and clearance, rather than Aβ production, may play a key role in the etiology of sporadic AD.This study was supported by the Alzheimer's Research UK, the Medical Research Council (MRC), the Wellcome Trust/MRC Joint Call in Neurodegeneration Award (WT089698) to the UK Parkinson's Disease Consortium (whose members are from the University College London Institute of Neurology, the University of Sheffield, and the MRC Protein Phosphorylation Unit at the University of Dundee), grants (P50 AG016574, U01 AG006786, and R01 AG18023), the National Institute for Health Research Biomedical Research Unit in Dementia at University College London Hospitals, University College London; the Big Lottery (to Dr. Morgan); a fellowship from Alzheimer's Research UK (to Dr. Guerreiro); and the Intramural Research Programs of the National Institute on Aging and the National Institute of Neurological Disease and Stroke, National Institutes of Health (Department of Health and Human Services Project number, ZO1 AG000950-10). The MRC London Neurodegenerative Diseases Brain Bank and the Manchester Brain Bank from Brains for Dementia Research are jointly funded from ARUK and AS. Tissue samples were supplied by The London Neurodegenerative Diseases Brain Bank, which receives funding from the MRC and as part of the Brains for Dementia Research programme, jointly funded by Alzheimer’s Research UK and Alzheimer’s Society

Investigating splicing variants uncovered by next-generation sequencing the Alzheimer’s disease candidate genes, CLU, PICALM, CR1, ABCA7, BIN1, the MS4A locus, CD2AP, EPHA1 and CD33

Late onset Alzheimer’s disease (LOAD), the most common cause of late onset dementia, has a strong genetic component. To date, 21 disease-risk loci have been identified through genome wide association studies (GWAS). However, the causative functional variant(s) within these loci are yet to be discovered. This study aimed to identify potential functional splicing mutations in the nine original GWAS-risk genes: CLU, PICALM, CR1, ABCA7, BIN1, the MS4A locus, CD2AP, EPHA1 and CD33. Target enriched next generation sequencing (NGS) was used to resequence the entire genetic region for each of these GWAS-risk loci in 96 LOAD patients and in silico databases were used to annotate the variants for functionality. Predicted splicing variants were further functionally characterised using splicing prediction software and minigene splicing assays. Following in silico annotation, 21 variants were predicted to influence splicing and, upon further annotation, four of these were examined utilising the in vitro minigene assay. Two variants, rs881768 A>G in ABCA7 and a novel variant 11: 60179827 T>G in MS4A6A were shown, in these cell assays, to affect the splicing of these genes. The method employed in the paper successfully identified potential splicing variants in GWAS-risk genes. Further investigation will be needed to understand the full effect of these variants on LOAD risk. However, these results suggest a possible pipeline in order to identify putative functional variants as a result of NGS in disease-associated loci although improvements are needed within the current prediction programme in order to reduce the number of false positives

Influence of coding variability in APP-Aß metabolism genes in sporadic Alzheimer's disease

The cerebral deposition of Aß42, a neurotoxic proteolitic derivate of amyloid precursor protein (APP), is a central event in Alzheimer’s disease (AD)(Amyloid hypothesis). Given the key role of APP-Aß metabolism in AD pathogenesis, we selected 29 genes involved in APP processing, Aß degradation and clearance. We then used exome and genome sequencing to investigate the single independent (single-variant association test) and cumulative (gene-based association test) effect of coding variants in these genes as potential susceptibility factors for AD, in a cohort composed of 435 sporadic and mainly late-onset AD cases and 801 elderly controls from North America and the UK. Our study shows that common coding variability in these genes does not play a major role for the disease development. In the single-variant association analysis, the main hits, which were nominally significant, were found to be very rare coding variants (MAF 0.3%-0.8%) that map to genes involved in APP processing (MEP1B), trafficking and recycling (SORL1), Aß extracellular degradation (ACE) and clearance (LRP1). Moreover, four genes (ECE1, LYZ, TTR and MME) have been found as nominally associated to AD using c-alpha and SKAT tests. We suggest that Aβ degradation and clearance, rather than Aβ production, may play a crucial role in the etiology of sporadic AD