- Email: [email protected]
IDENTIFICATION OF NOVEL GENETIC RISK VARIANTS FOR ALZHEIMER’S DISEASE IN THE HMGCR GENE LOCUS
Poster Presentations: Sunday, July 16, 2017
efforts are underway to identify the causative gene and elucidate its contribution to the phenotypic variability in this family. P1-157
THE PREDICTION METHOD OF DELETERIOUS VARIANTS FOR ALZHEIMER’S DISEASE USING CHROMATIN HIGHER-ORDER STRUCTURE 1
2
3
2
Background: The most common cause of dementia is late-onset Alz-
heimer’s disease (AD), which occurs in individuals aged >65 years and leads to neuronal death. The heritability of AD was reported as approximately 60-80%, indicating that the genetic factors are strongly associated with the pathogenesis of AD. To identify the genetic variants associated with AD, genome-wide association studies (GWAS) and whole-exome sequencing have been performed across some cohorts and ethnic groups. However most of these loci are located within introns and intergenic regions. In this study, we explored whether AD-associated variants are in functional noncoding regions such as enhancers, and predicted a deleterious effect of AD-associated variants. Given that enhancers participate in longrange interactions via chromatin loops, we determined these longrange chromatin interactions for AD-associated variants in candidate regions of enhancers and searched for genes in the neighborhood of AD-associated variants. Methods: We collected 208 ADassociated variants from the GWAS catalog database and defined enhancers by transcription factor binding motifs, DNase-peaks and ChiP-seq data of enhancer marks (H3K4me3 and H3K27ac). We explored AD-associated variants close to the enhancers. To identify long-range chromatin interactions for AD-associated variants, we performed tethered conformation capture (TCC) in the human neuroblastoma SK-N-SH and human astrocytoma U-251 MG cell lines. Results: We found that 19 AD-associated variants lying in non-coding regions were localized in the proximity of regulatory elements. These variants are predicted to work as enhancers/silencers and may have a deleterious effect on the transcription of distant genes. For instance, 2 variants in the APOE gene cluster, which includes APOE, a strong risk factor for AD, were localized in transcription factor binding regions. Furthermore, analysis of the chromatin structure by TCC revealed that the APOE gene cluster conformationally interacted with some genes involved in AD pathology. The chromatin interactions of the APOE gene cluster were observed in the neuroblastoma and astrocytoma cell lines. Conclusions: Our study suggested that some AD-associated variants exert a deleterious effect on the transcriptional regulation of genes at distinct genomic regions through chromatin higher-order structure.
IDENTIFICATION OF NOVEL GENETIC RISK VARIANTS FOR ALZHEIMER’S DISEASE IN THE HMGCR GENE LOCUS
Nathalie I. V. Nilsson1,2, Cynthia Picard2, Judes Poirier3,4,5, 1 McGill University, Montreal, QC, Canada; 2Douglas Mental Health University Institute Research Centre, Montreal, QC, Canada; 3Centre for the Studies on the Prevention of Alzheimer’s
Disease, Douglas Mental Health University Institute, Montreal, QC, Canada; 4Douglas Mental Health University Institute, Montreal, QC, Canada; 5Centre for Studies on Prevention of Alzheimer’s Disease (StoP-AD Centre), Douglas Mental Health Institute, Montreal, QC, Canada. Contact e-mail: nathalie. [email protected] Background: Cholesterol lowering drugs inhibiting HMG-CoA
Masataka Kikuchi , Norikazu Hara , Mai Hasegawa , Akinori Miyashita , Ryozo Kuwano2, Takeshi Ikeuchi2, Akihiro Nakaya1, 1Graduate School of Medicine, Osaka University, Osaka, Japan; 2Brain Research Institute, Niigata University, Niigata, Japan; 3Research Institute for Biomedical Sciences (RIBS), Tokyo University of Science, Chiba, Japan. Contact e-mail: [email protected]
P1-158
P303
reductase activity (statins) have been found to be protective in retrospective analyses of sporadic AD. Recently, a genetic variant in the HMGCR gene, also associated with reduced reductase activity, was found to be protective in AD (Leduc et al., 2015). This led us to systematically investigate all the genetic polymorphisms at the HMGCR locus with a population frequency greater than 5%, and their relationship to AD risk and HMGCR expression. Methods: Whole-genome sequencing, plasma, and cerebrospinal fluid (CSF) biomarker data were obtained from the Alzheimer’s Disease Neuroimaging Initiative (ADNI) database (adni.loni.usc.edu). Genetic polymorphisms in and around the HMGCR gene were extracted, and logistic regressions were performed in plink (pngu. mgh.harvard.edu) to identify variants associated with either risk or protection. Further analyses were performed in SPSS; KaplanMeier for conversion rate analysis, ANCOVA for expression analysis and Mann-Whitney U test for CSF biomarker analyses. Results: We identified three new single nucleotide polymorphisms (SNPs) associated with marked increased risk of AD (ORs ¼ 2.4 - 2.7, ps < 0.05), however only in subjects also positive for the APOE4 allele. Survival analysis further revealed that carriers of the risk alleles exhibit an accelerated conversion rate, but again only in APOE4 carriers (X2s > 6.6, ps < 0.01). HMGCR risk alleles were also associated with an increased gene expression in peripheral lymphocytes (Fs > 6.7, ps < 0.01); with no interaction with APOE4 status. Finally, a significant increase in CSF phospho-tau/ tau ratio was observed in carriers of the risk alleles, specifically in APOE4 positive females (Us ¼ 245.5, ps ¼ 0.017), but not for Ab levels. Conclusions: We have identified three new SNPs in the HMGCR locus associated with risk of AD, accelerated conversion rate to AD, and increased CSF phospho-tau/tau ratio, specifically in APOE4 carriers. In line with our hypothesis that protection is mediated by reduction in HMG-CoA reductase activity, the identified risk alleles were shown to be significantly associated with increased HMGCR expression. We are now in the process of replicating these findings in an independent cohort composed of autopsy-confirmed AD cases and age-matched controls from a population isolate form eastern Canada.
P1-159
PPARg: ROLE IN THE REGULATION OF THE TOMM40-APOE-C1 GENES CLUSTER
Ornit Chiba-Falek, Shobana Subramanian, William Kirby Gottschalk, So Young Kim, Duke University, Durham, NC, USA. Contact e-mail: [email protected] Background: Chromosome 19q13.32 is a gene rich region, and has been implicated in multiple human phenotypes in adulthood including lipid traits, Alzheimer’s disease, and longevity. Peroxisome Proliferator Activated Receptor Gamma (PPARg) is a ligand-activated nuclear transcription factor that plays a role in human complex traits that are also genetically associated with the chromosome 19q13.32 region. Methods: Here, we studied the effects of PPARg on the regional expression regulation of the genes clustered within chromosome 19q13.32, specifically TOMM40, APOE, and APOC1, applying two complementary approaches. Using the short hairpin RNA (shRNA) method in the HepG2 cell-line
efforts are underway to identify the causative gene and elucidate its contribution to the phenotypic variability in this family. P1-157
THE PREDICTION METHOD OF DELETERIOUS VARIANTS FOR ALZHEIMER’S DISEASE USING CHROMATIN HIGHER-ORDER STRUCTURE 1
2
3
2
Background: The most common cause of dementia is late-onset Alz-
heimer’s disease (AD), which occurs in individuals aged >65 years and leads to neuronal death. The heritability of AD was reported as approximately 60-80%, indicating that the genetic factors are strongly associated with the pathogenesis of AD. To identify the genetic variants associated with AD, genome-wide association studies (GWAS) and whole-exome sequencing have been performed across some cohorts and ethnic groups. However most of these loci are located within introns and intergenic regions. In this study, we explored whether AD-associated variants are in functional noncoding regions such as enhancers, and predicted a deleterious effect of AD-associated variants. Given that enhancers participate in longrange interactions via chromatin loops, we determined these longrange chromatin interactions for AD-associated variants in candidate regions of enhancers and searched for genes in the neighborhood of AD-associated variants. Methods: We collected 208 ADassociated variants from the GWAS catalog database and defined enhancers by transcription factor binding motifs, DNase-peaks and ChiP-seq data of enhancer marks (H3K4me3 and H3K27ac). We explored AD-associated variants close to the enhancers. To identify long-range chromatin interactions for AD-associated variants, we performed tethered conformation capture (TCC) in the human neuroblastoma SK-N-SH and human astrocytoma U-251 MG cell lines. Results: We found that 19 AD-associated variants lying in non-coding regions were localized in the proximity of regulatory elements. These variants are predicted to work as enhancers/silencers and may have a deleterious effect on the transcription of distant genes. For instance, 2 variants in the APOE gene cluster, which includes APOE, a strong risk factor for AD, were localized in transcription factor binding regions. Furthermore, analysis of the chromatin structure by TCC revealed that the APOE gene cluster conformationally interacted with some genes involved in AD pathology. The chromatin interactions of the APOE gene cluster were observed in the neuroblastoma and astrocytoma cell lines. Conclusions: Our study suggested that some AD-associated variants exert a deleterious effect on the transcriptional regulation of genes at distinct genomic regions through chromatin higher-order structure.
IDENTIFICATION OF NOVEL GENETIC RISK VARIANTS FOR ALZHEIMER’S DISEASE IN THE HMGCR GENE LOCUS
Nathalie I. V. Nilsson1,2, Cynthia Picard2, Judes Poirier3,4,5, 1 McGill University, Montreal, QC, Canada; 2Douglas Mental Health University Institute Research Centre, Montreal, QC, Canada; 3Centre for the Studies on the Prevention of Alzheimer’s
Disease, Douglas Mental Health University Institute, Montreal, QC, Canada; 4Douglas Mental Health University Institute, Montreal, QC, Canada; 5Centre for Studies on Prevention of Alzheimer’s Disease (StoP-AD Centre), Douglas Mental Health Institute, Montreal, QC, Canada. Contact e-mail: nathalie. [email protected] Background: Cholesterol lowering drugs inhibiting HMG-CoA
Masataka Kikuchi , Norikazu Hara , Mai Hasegawa , Akinori Miyashita , Ryozo Kuwano2, Takeshi Ikeuchi2, Akihiro Nakaya1, 1Graduate School of Medicine, Osaka University, Osaka, Japan; 2Brain Research Institute, Niigata University, Niigata, Japan; 3Research Institute for Biomedical Sciences (RIBS), Tokyo University of Science, Chiba, Japan. Contact e-mail: [email protected]
P1-158
P303
reductase activity (statins) have been found to be protective in retrospective analyses of sporadic AD. Recently, a genetic variant in the HMGCR gene, also associated with reduced reductase activity, was found to be protective in AD (Leduc et al., 2015). This led us to systematically investigate all the genetic polymorphisms at the HMGCR locus with a population frequency greater than 5%, and their relationship to AD risk and HMGCR expression. Methods: Whole-genome sequencing, plasma, and cerebrospinal fluid (CSF) biomarker data were obtained from the Alzheimer’s Disease Neuroimaging Initiative (ADNI) database (adni.loni.usc.edu). Genetic polymorphisms in and around the HMGCR gene were extracted, and logistic regressions were performed in plink (pngu. mgh.harvard.edu) to identify variants associated with either risk or protection. Further analyses were performed in SPSS; KaplanMeier for conversion rate analysis, ANCOVA for expression analysis and Mann-Whitney U test for CSF biomarker analyses. Results: We identified three new single nucleotide polymorphisms (SNPs) associated with marked increased risk of AD (ORs ¼ 2.4 - 2.7, ps < 0.05), however only in subjects also positive for the APOE4 allele. Survival analysis further revealed that carriers of the risk alleles exhibit an accelerated conversion rate, but again only in APOE4 carriers (X2s > 6.6, ps < 0.01). HMGCR risk alleles were also associated with an increased gene expression in peripheral lymphocytes (Fs > 6.7, ps < 0.01); with no interaction with APOE4 status. Finally, a significant increase in CSF phospho-tau/ tau ratio was observed in carriers of the risk alleles, specifically in APOE4 positive females (Us ¼ 245.5, ps ¼ 0.017), but not for Ab levels. Conclusions: We have identified three new SNPs in the HMGCR locus associated with risk of AD, accelerated conversion rate to AD, and increased CSF phospho-tau/tau ratio, specifically in APOE4 carriers. In line with our hypothesis that protection is mediated by reduction in HMG-CoA reductase activity, the identified risk alleles were shown to be significantly associated with increased HMGCR expression. We are now in the process of replicating these findings in an independent cohort composed of autopsy-confirmed AD cases and age-matched controls from a population isolate form eastern Canada.
P1-159
PPARg: ROLE IN THE REGULATION OF THE TOMM40-APOE-C1 GENES CLUSTER
Ornit Chiba-Falek, Shobana Subramanian, William Kirby Gottschalk, So Young Kim, Duke University, Durham, NC, USA. Contact e-mail: [email protected] Background: Chromosome 19q13.32 is a gene rich region, and has been implicated in multiple human phenotypes in adulthood including lipid traits, Alzheimer’s disease, and longevity. Peroxisome Proliferator Activated Receptor Gamma (PPARg) is a ligand-activated nuclear transcription factor that plays a role in human complex traits that are also genetically associated with the chromosome 19q13.32 region. Methods: Here, we studied the effects of PPARg on the regional expression regulation of the genes clustered within chromosome 19q13.32, specifically TOMM40, APOE, and APOC1, applying two complementary approaches. Using the short hairpin RNA (shRNA) method in the HepG2 cell-line