A familial missense variant in the Alzheimer's disease gene SORL1 impairs its maturation and endosomal sorting.

The SORL1 gene has recently emerged as a strong Alzheimer's Disease (AD) risk gene. Over 500 different variants have been identified in the gene and the contribution of individual variants to AD development and progression is still largely unknown. Here, we describe a family consisting of 2 parents and 5 offspring. Both parents were affected with dementia and one had confirmed AD pathology with an age of onset > 75 years. All offspring were affected with AD with ages at onset ranging from 53 years to 74 years. DNA was available from the parent with confirmed AD and 5 offspring. We identified a coding variant, p.(Arg953Cys), in SORL1 in 5 of 6 individuals affected by AD. Notably, variant carriers had severe AD pathology, and the SORL1 variant segregated with TDP-43 pathology (LATE-NC). We further characterized this variant and show that this Arginine substitution occurs at a critical position in the YWTD-domain of the SORL1 translation product, SORL1. Functional studies further show that the p.R953C variant leads to retention of the SORL1 protein in the endoplasmic reticulum which leads to decreased maturation and shedding of the receptor and prevents its normal endosomal trafficking. Together, our analysis suggests that p.R953C is a pathogenic variant of SORL1 and sheds light on mechanisms of how missense SORL1 variants may lead to AD.

Evolution of a Human-Specific Tandem Repeat Associated with ALS.

Tandem repeats are proposed to contribute to human-specific traits, and more than 40 tandem repeat expansions are known to cause neurological disease. Here, we characterize a human-specific 69 bp variable number tandem repeat (VNTR) in the last intron of WDR7, which exhibits striking variability in both copy number and nucleotide composition, as revealed by long-read sequencing. In addition, greater repeat copy number is significantly enriched in three independent cohorts of individuals with sporadic amyotrophic lateral sclerosis (ALS). Each unit of the repeat forms a stem-loop structure with the potential to produce microRNAs, and the repeat RNA can aggregate when expressed in cells. We leveraged its remarkable sequence variability to align the repeat in 288 samples and uncover its mechanism of expansion. We found that the repeat expands in the 3'-5' direction, in groups of repeat units divisible by two. The expansion patterns we observed were consistent with duplication events, and a replication error called template switching. We also observed that the VNTR is expanded in both Denisovan and Neanderthal genomes but is fixed at one copy or fewer in non-human primates. Evaluating the repeat in 1000 Genomes Project samples reveals that some repeat segments are solely present or absent in certain geographic populations. The large size of the repeat unit in this VNTR, along with our multiplexed sequencing strategy, provides an unprecedented opportunity to study mechanisms of repeat expansion, and a framework for evaluating the roles of VNTRs in human evolution and disease.

APOE Locus-Associated Mitochondrial Function and Its Implication in Alzheimer's Disease and Aging.

The Apolipoprotein E (APOE) locus has garnered significant clinical interest because of its association with Alzheimer's disease (AD) and longevity. This genetic association appears across multiple genes in the APOE locus. Despite the apparent differences between AD and longevity, both conditions share a commonality of aging-related changes in mitochondrial function. This commonality is likely due to accumulative biological effects partly exerted by the APOE locus. In this study, we investigated changes in mitochondrial structure/function-related markers using oxidative stress-induced human cellular models and postmortem brains (PMBs) from individuals with AD and normal controls. Our results reveal a range of expressional alterations, either upregulated or downregulated, in these genes in response to oxidative stress. In contrast, we consistently observed an upregulation of multiple APOE locus genes in all cellular models and AD PMBs. Additionally, the effects of AD status on mitochondrial DNA copy number (mtDNA CN) varied depending on APOE genotype. Our findings imply a potential coregulation of APOE locus genes possibly occurring within the same topologically associating domain (TAD) of the 3D chromosome conformation. The coordinated expression of APOE locus genes could impact mitochondrial function, contributing to the development of AD or longevity. Our study underscores the significant role of the APOE locus in modulating mitochondrial function and provides valuable insights into the underlying mechanisms of AD and aging, emphasizing the importance of this locus in clinical research.

TOMM40 genetic variants associated with healthy aging and longevity: a systematic review.

INTRODUCTION: Healthy aging relies on mitochondrial functioning because this organelle provides energy and diminishes oxidative stress. Single nucleotide polymorphisms (SNPs) in TOMM40, a critical gene that produces the outer membrane protein TOM40 of mitochondria, have been associated with mitochondrial dysfunction and neurodegenerative processes. Yet it is not clear whether or how the mitochondria may impact human longevity. We conducted this review to ascertain which SNPs have been associated with markers of healthy aging. METHODS: Using the PRISMA methodology, we conducted a systematic review on PubMed and Embase databases to identify associations between TOMM40 SNPs and measures of longevity and healthy aging. RESULTS: Twenty-four articles were selected. The TOMM40 SNPs rs2075650 and rs10524523 were the two most commonly identified and studied SNPs associated with longevity. The outcomes associated with the TOMM40 SNPs were changes in BMI, brain integrity, cognitive functions, altered inflammatory network, vulnerability to vascular risk factors, and longevity. DISCUSSIONS: Our systematic review identified multiple TOMM40 SNPs potentially associated with healthy aging. Additional research can help to understand mechanisms in aging, including resilience, prevention of disease, and adaptation to the environment.

DNA methylation of TOMM40-APOE-APOC2 in Alzheimer's disease.

The apolipoprotein E (APOE) ε4 allele is the major genetic risk factor for Alzheimer's disease (AD). Multiple regulatory elements, spanning the extended TOMM40-APOE-APOC2 region, regulate gene expression at this locus. Regulatory element DNA methylation changes occur under different environmental conditions, such as disease. Our group and others have described an APOE CpG island as hypomethylated in AD, compared to cognitively normal controls. However, little is known about methylation of the larger TOMM40-APOE-APOC2 region. The hypothesis of this investigation was that regulatory element methylation levels of the larger TOMM40-APOE-APOC2 region are associated with AD. The aim was to determine whether DNA methylation of the TOMM40-APOE-APOC2 region differs in AD compared to cognitively normal controls in post-mortem brain and peripheral blood. DNA was extracted from human brain (n = 12) and peripheral blood (n = 67). A methylation array was used for this analysis. Percent methylation within the TOMM40-APOE-APOC2 region was evaluated for differences according to tissue type, disease state, AD-related biomarkers, and gene expression. Results from this exploratory analysis suggest that regulatory element methylation levels within the larger TOMM40-APOE-APOC2 gene region correlate with AD-related biomarkers and TOMM40 or APOE gene expression in AD.

APOE DNA methylation is altered in Lewy body dementia.

INTRODUCTION: Inheritance of the ε4 allele of apolipoprotein E (APOE) increases a person's risk of developing both Alzheimer's disease (AD) and Lewy body dementia (LBD), yet the underlying mechanisms behind this risk are incompletely understood. The recent identification of reduced APOE DNA methylation in AD postmortem brains prompted this study to investigate APOE methylation in LBD. METHODS: Genomic DNA from postmortem brain tissues (frontal lobe and cerebellum) of neuropathological pure (np) controls and npAD, LBD + AD, and npLBD subjects were bisulfite pyrosequenced. DNA methylation levels of two APOE subregions were then compared for these groups. RESULTS: APOE DNA methylation was significantly reduced in npLBD compared with np controls, and methylation levels were lowest in the LBD + AD group. DISCUSSION: Given that npLBD and npAD postmortem brains shared a similar reduction in APOE methylation, it is possible that an aberrant epigenetic change in APOE is linked to risk for both diseases.

Epigenetic signature and enhancer activity of the human APOE gene.

The human apolipoprotein E (APOE) gene plays an important role in lipid metabolism. It has three common genetic variants, alleles ε2/ε3/ε4, which translate into three protein isoforms of apoE2, E3 and E4. These isoforms can differentially influence total serum cholesterol levels; therefore, APOE has been linked with cardiovascular disease. Additionally, its ε4 allele is strongly associated with the risk of Alzheimer's disease (AD), whereas the ε2 allele appears to have a modest protective effect for AD. Despite decades of research having illuminated multiple functional differences among the three apoE isoforms, the precise mechanisms through which different APOE alleles modify diseases risk remain incompletely understood. In this study, we examined the genomic structure of APOE in search for properties that may contribute novel biological consequences to the risk of disease. We identify one such element in the ε2/ε3/ε4 allele-carrying 3'-exon of APOE. We show that this exon is imbedded in a well-defined CpG island (CGI) that is highly methylated in the human postmortem brain. We demonstrate that this APOE CGI exhibits transcriptional enhancer/silencer activity. We provide evidence that this APOE CGI differentially modulates expression of genes at the APOE locus in a cell type-, DNA methylation- and ε2/ε3/ε4 allele-specific manner. These findings implicate a novel functional role for a 3'-exon CGI and support a modified mechanism of action for APOE in disease risk, involving not only the protein isoforms but also an epigenetically regulated transcriptional program at the APOE locus driven by the APOE CGI.

APOE loss-of-function variants: Compatible with longevity and associated with resistance to Alzheimer's disease pathology.

The ε4 allele of apolipoprotein E (APOE) is the strongest genetic risk factor for sporadic Alzheimer's disease (AD). Knockdown of ε4 may provide a therapeutic strategy for AD, but the effect of APOE loss of function (LoF) on AD pathogenesis is unknown. We searched for APOE LoF variants in a large cohort of controls and patients with AD and identified seven heterozygote carriers of APOE LoF variants. Five carriers were controls (aged 71-90 years), one carrier was affected by progressive supranuclear palsy, and one carrier was affected by AD with an unremarkable age at onset of 75 years. Two APOE ε3/ε4 controls carried a stop-gain affecting ε4: one was cognitively normal at 90 years and had no neuritic plaques at autopsy; the other was cognitively healthy at 79 years, and lumbar puncture at 76 years showed normal levels of amyloid. These results suggest that ε4 drives AD risk through the gain of abnormal function and support ε4 knockdown as a viable therapeutic option.

Inheritance model introduces differential bias in CNV calls between parents and offspring.

Copy Number Variation (CNV) is increasingly implicated in disease pathogenesis. CNVs are often identified by statistical models applied to data from single nucleotide polymorphism panels. Family information for samples provides additional information for CNV inference. Two modes of PennCNV (the Joint-call and Posterior-call), which are some of the most well-developed family-based CNV calling methods, use a "Joint-model" as a main component. This models all family members' CNV states together with Mendelian inheritance. Methods based on the Joint-model are used to infer CNV calls of cases and controls in a pedigree, which may be compared to each other to test an association. Although benefits from the Joint-model have been shown elsewhere, equality of call rates in parents and offspring has not been evaluated previously. This can affect downstream analyses in studies that compare CNV rates in cases vs. controls in pedigrees. In this paper, we show that the Joint-model can introduce different CNV call rates among family members in the absence of a true difference. We show that the Joint-model may analytically introduce differential CNV calls because of asymmetry of the model. We demonstrate these differential call rates using single-marker simulations. We show that call rates using the two modes of PennCNV also differ between parents and offspring in one multimarker simulated dataset and two real datasets. Our results advise need for caution in use of the Joint-model calls in CNV association studies with family-based datasets.

MicroRNA in Alzheimer's disease: an exploratory study in brain, cerebrospinal fluid and plasma.

MicroRNA (miRNA) may be potential biomarkers of Alzheimer's disease (AD). The objective of this investigation was to demonstrate that miRNAs in human brain or biofluids are differentially expressed according to disease status, tissue type, neuritic plaque score or Braak stage. Post-mortem brain (PMB) miRNA were profiled using arrays and validated using quantitative RT-PCR (qRT-PCR). Five qRT-PCR-validated miRNAs were measured in an independent sample of PMB, cerebrospinal fluid and plasma from the same subjects. Plasma miR-15a was found to be associated with plaque score in the independent sample. In conclusion, miRNA present in human biofluids may offer utility as biomarkers of AD.

TOMM40 intron 6 poly-T length, age at onset, and neuropathology of AD in individuals with APOE ε3/ε3.

BACKGROUND: This study investigates the association between TOMM40 poly-T length, age at onset, and neuropathology in individuals with Alzheimer's disease (AD) with the apolipoprotein E (APOE) ε3/ε3 allele. METHODS: Thirty-two presenilin 1 (PSEN1) mutation carriers with AD, 27 presenilin 2 (PSEN2) mutation carriers with AD, 59 participants with late-onset AD (LOAD), and 168 autopsied subjects from a community-based cohort were genotyped for TOMM40 intron 6 poly-T (rs10524523) length using short tandem repeat assays. RESULTS: Among AD individuals with PSEN2 mutations, the presence of a long poly-T was associated with an earlier age at onset, whereas there were no such associations for subjects with PSEN1 mutations or LOAD. In community-based participants, the presence of a long poly-T was associated with increased neuritic tangles and a greater likelihood of pathologically diagnosed AD. CONCLUSION: TOMM40 intron 6 poly-T length may explain some of the variation in age at onset in PSEN2 familial AD and may be associated with AD neuropathology in persons with APOE ε3/ε3.

A Preliminary Comparison of the Methylome and Transcriptome from the Prefrontal Cortex Across Alzheimer's Disease and Lewy Body Dementia.

Background: Pathological amyloid-ß and α-synuclein are associated with a spectrum of related dementias, ranging from Alzheimer's disease (AD) and dementia with Lewy bodies (DLB) to Parkinson disease dementia (PDD). While these diseases share clinical and pathological features, they also have unique patterns of pathology. However, epigenetic factors that contribute to these pathological differences remain unknown. Objective: In this preliminary study, we explore differences in DNA methylation and transcription in five neuropathologically defined groups: cognitively unimpaired controls, AD, pure DLB, DLB with concomitant AD (DLBAD), and PDD. Methods: We employed an Illumina Infinium 850k array and RNA-seq to quantify these differences in DNA methylation and transcription, respectively. We then used Weighted Gene Co-Network Expression Analysis (WGCNA) to determine transcriptional modules and correlated these with DNA methylation. Results: We found that PDD was transcriptionally unique and correlated with an unexpected hypomethylation pattern compared to the other dementias and controls. Surprisingly, differences between PDD and DLB were especially notable with 197 differentially methylated regions. WGCNA yielded numerous modules associated with controls and the four dementias: one module was associated with transcriptional differences between controls and all the dementias as well as having significant overlap with differentially methylated probes. Functional enrichment demonstrated that this module was associated with responses to oxidative stress. Conclusion: Future work that extends these joint DNA methylation and transcription analyses will be critical to better understanding of differences that contribute to varying clinical presentation across dementias.

Weight Loss Is a Strong Predictor of Memory Disorder Independent of Genetic Influences.

BACKGROUND: Past studies identified a link between weight loss and dementia, but lacked consistent conclusions. We sought to establish this link by examining the weight change profiles before and after dementia diagnosis. METHODS: Using data from the Health and Retirement Study (1996-2020), we examined 13,123 participants. We conducted a nested case-control analysis to assess differences in biennial weight change profile while controlling for BMI, longevity polygenic risk scores, and APOE gene variants. RESULTS: Participants with a memory disorder lost weight (-0.63%) biennially, whereas those without a diagnosis did not (+0.013%, p-value < 0.0001). Our case-control study shows a significant difference (p-value < 0.01) in pre-dementia % weight changes between the cases (-0.29%) and controls (0.19%), but not in post-dementia weight changes. The weight loss group have the highest risk (OR = 2.01; p-value < 0.0001) of developing a memory disorder compared to the stable weight and weight gain groups. The observations hold true after adjusting for BMI, longevity polygenic risk scores, and APOE variant in a multivariable model. CONCLUSIONS: We observe that weight loss in dementia is a physiological process independent of genetic factors associated with BMI and longevity. Pre-dementia weight loss may be an important prognostic criterion to assess a person's risk of developing a memory disorder.

A familial missense variant in the Alzheimer's Disease gene SORL1 impairs its maturation and endosomal sorting.

The SORL1 gene has recently emerged as a strong Alzheimer's Disease (AD) risk gene. Over 500 different variants have been identified in the gene and the contribution of individual variants to AD development and progression is still largely unknown. Here, we describe a family consisting of 2 parents and 5 offspring. Both parents were affected with dementia and one had confirmed AD pathology with an age of onset >75 years. All offspring were affected with AD with ages at onset ranging from 53yrs-74yrs. DNA was available from the parent with confirmed AD and 5 offspring. We identified a coding variant, p.(Arg953Cys), in SORL1 in 5 of 6 individuals affected by AD. Notably, variant carriers had severe AD pathology, and the SORL1 variant segregated with TDP-43 pathology (LATE-NC). We further characterized this variant and show that this Arginine substitution occurs at a critical position in the YWTD-domain of the SORL1 translation product, SORL1. Functional studies further show that the p.R953C variant leads to retention of the SORL1 protein in the endoplasmic reticulum which leads to decreased maturation and shedding of the receptor and prevents its normal endosomal trafficking. Together, our analysis suggests that p.R953C is a pathogenic variant of SORL1 and sheds light on mechanisms of how missense SORL1 variants may lead to AD.

APOE loss-of-function variants: Compatible with longevity and associated with resistance to Alzheimer's Disease pathology.

The ε4 allele of apolipoprotein E (APOE) is the strongest genetic risk factor for sporadic Alzheimer's Disease (AD). Knockdown of this allele may provide a therapeutic strategy for AD, but the effect of APOE loss-of-function (LoF) on AD pathogenesis is unknown. We searched for APOE LoF variants in a large cohort of older controls and patients with AD and identified six heterozygote carriers of APOE LoF variants. Five carriers were controls (ages 71-90) and one was an AD case with an unremarkable age-at-onset between 75-79. Two APOE ε3/ε4 controls (Subjects 1 and 2) carried a stop-gain affecting the ε4 allele. Subject 1 was cognitively normal at 90+ and had no neuritic plaques at autopsy. Subject 2 was cognitively healthy within the age range 75-79 and underwent lumbar puncture at between ages 75-79 with normal levels of amyloid. The results provide the strongest human genetics evidence yet available suggesting that ε4 drives AD risk through a gain of abnormal function and support knockdown of APOE ε4 or its protein product as a viable therapeutic option.

Functional analysis of APOE locus genetic variation implicates regional enhancers in the regulation of both TOMM40 and APOE.

Genetic variation within the apolipoprotein E gene (APOE) locus is associated with late-onset Alzheimer's disease risk and quantitative traits as well as apoE expression in multiple tissues. The aim of this investigation was to explore the influence of APOE locus cis-regulatory element enhancer region genetic variation on regional gene promoter activity. Luciferase reporter constructs containing haplotypes of APOE locus gene promoters; APOE, APOC1 and TOMM40, and regional putative enhancers; TOMM40 intervening sequence (IVS)2-4, TOMM40 IVS6 poly-T, as well as previously described enhancers; multienhancer 1 (ME1), or brain control region (BCR), were evaluated for their effects on luciferase activity in three human cell lines. Results of this investigation demonstrate that in SHSY5Y cells, the APOE promoter is significantly influenced by the TOMM40 IVS2-4 and ME1, and the TOMM40 promoter is significantly influenced by the TOMM40 IVS6 poly-T, ME1 and BCR. In HepG2 cells, theTOMM40 promoter is significantly influenced by all four enhancers, whereas the APOE promoter is not influenced by any of the enhancers. The main novel finding of this investigation was that multiple APOE locus cis-elements influence both APOE and TOMM40 promoter activity according to haplotype and cell type, suggesting that a complex transcriptional regulatory structure modulates regional gene expression.

Tau phosphorylation pathway genes and cerebrospinal fluid tau levels in Alzheimer's disease.

Alzheimer's disease (AD) is characterized by the presence in the brain of amyloid plaques, consisting predominately of the amyloid ß peptide (Aß), and neurofibrillary tangles, consisting primarily of tau. Hyper-phosphorylated-tau (p-tau) contributes to neuronal damage, and both p-tau and total-tau (t-tau) levels are elevated in AD cerebrospinal fluid (CSF) compared to cognitively normal controls. Our hypothesis was that increased ratios of CSF phosphorylated-tau levels relative to total-tau levels correlate with regulatory region genetic variation of kinase or phosphatase genes biologically associated with the phosphorylation status of tau. Eighteen SNPs located within 5' and 3' regions of 5 kinase and 4 phosphatase genes, as well as two SNPs within regulatory regions of the MAPT gene were chosen for this analysis. The study sample consisted of 101 AD patients and 169 cognitively normal controls. Rs7768046 in the FYN kinase gene and rs913275 in the PPP2R4 phosphatase gene were both associated with CSF p-tau and t-tau levels in AD. These SNPs were also differentially associated with either CSF t-tau (rs7768046) or CSF p-tau (rs913275) relative to t-tau levels in AD compared to controls. These results suggest that rs7768046 and rs913275 both influence CSF tau levels in an AD-associated manner.

Cerebrocerebellar hypometabolism associated with repetitive blast exposure mild traumatic brain injury in 12 Iraq war Veterans with persistent post-concussive symptoms.

Disagreement exists regarding the extent to which persistent post-concussive symptoms (PCS) reported by Iraq combat Veterans with repeated episodes of mild traumatic brain injury (mTBI) from explosive blasts represent structural or functional brain damage or an epiphenomenon of comorbid depression or posttraumatic stress disorder (PTSD). Objective assessment of brain function in this population may clarify the issue. To this end, twelve Iraq war Veterans (32.0 ± 8.5 [mean ± standard deviation (SD)] years of age) reporting one or more blast exposures meeting American Congress of Rehabilitation Medicine criteria for mTBI and persistent PCS and 12 cognitively normal community volunteers (53.0 ± 4.6 years of age) without history of head trauma underwent brain fluorodeoxyglucose positron emission tomography (FDG-PET) and neuropsychological assessments and completed PCS and psychiatric symptom rating scales. Compared to controls, Veterans with mTBI (with or without PTSD) exhibited decreased cerebral metabolic rate of glucose in the cerebellum, vermis, pons, and medial temporal lobe. They also exhibited subtle impairments in verbal fluency, cognitive processing speed, attention, and working memory, similar to those reported in the literature for patients with cerebellar lesions. These FDG-PET imaging findings suggest that regional brain hypometabolism may constitute a neurobiological substrate for chronic PCS in Iraq combat Veterans with repetitive blast-trauma mTBI. Given the potential public health implications of these findings, further investigation of brain function in these Veterans appears warranted.

TOMM40 RNA Transcription in Alzheimer's Disease Brain and Its Implication in Mitochondrial Dysfunction.

Increasing evidence suggests that the Translocase of Outer Mitochondria Membrane 40 (TOMM40) gene may contribute to the risk of Alzheimer's disease (AD). Currently, there is no consensus as to whether TOMM40 expression is up- or down-regulated in AD brains, hindering a clear interpretation of TOMM40's role in this disease. The aim of this study was to determine if TOMM40 RNA levels differ between AD and control brains. We applied RT-qPCR to study TOMM40 transcription in human postmortem brain (PMB) and assessed associations of these RNA levels with genetic variants in APOE and TOMM40. We also compared TOMM40 RNA levels with mitochondrial functions in human cell lines. Initially, we found that the human genome carries multiple TOMM40 pseudogenes capable of producing highly homologous RNAs that can obscure precise TOMM40 RNA measurements. To circumvent this obstacle, we developed a novel RNA expression assay targeting the primary transcript of TOMM40. Using this assay, we showed that TOMM40 RNA was upregulated in AD PMB. Additionally, elevated TOMM40 RNA levels were associated with decreases in mitochondrial DNA copy number and mitochondrial membrane potential in oxidative stress-challenged cells. Overall, differential transcription of TOMM40 RNA in the brain is associated with AD and could be an indicator of mitochondrial dysfunction.

Genetics of Alzheimer disease.

Alzheimer disease (AD) is the most common causes of neurodegenerative disorder in the elderly individuals. Clinically, patients initially present with short-term memory loss, subsequently followed by executive dysfunction, confusion, agitation, and behavioral disturbances. Three causative genes have been associated with autosomal dominant familial AD (APP, PSEN1, and PSEN2) and 1 genetic risk factor (APOEε4 allele). Identification of these genes has led to a number of animal models that have been useful to study the pathogenesis underlying AD. In this article, we provide an overview of the clinical and genetic features of AD.