24-Hour Rhythms of DNA Methylation and Their Relation with Rhythms of RNA Expression in the Human Dorsolateral Prefrontal Cortex

Circadian rhythms modulate the biology of many human tissues, including brain tissues, and are driven by a near 24-hour transcriptional feedback loop. These rhythms are paralleled by 24-hour rhythms of large portions of the transcriptome. The role of dynamic DNA methylation in influencing these rhythms is uncertain. While recent work in Neurospora suggests that dynamic site-specific circadian rhythms of DNA methylation may play a role in modulating the fungal molecular clock, such rhythms and their relationship to RNA expression have not, to our knowledge, been elucidated in mammalian tissues, including human brain tissues. We hypothesized that 24-hour rhythms of DNA methylation exist in the human brain, and play a role in driving 24-hour rhythms of RNA expression. We analyzed DNA methylation levels in post-mortem human dorsolateral prefrontal cortex samples from 738 subjects. We assessed for 24-hour rhythmicity of 420,132 DNA methylation sites throughout the genome by considering methylation levels as a function of clock time of death and parameterizing these data using cosine functions. We determined global statistical significance by permutation. We then related rhythms of DNA methylation with rhythms of RNA expression determined by RNA sequencing. We found evidence of significant 24-hour rhythmicity of DNA methylation. Regions near transcription start sites were enriched for high-amplitude rhythmic DNA methylation sites, which were in turn time locked to 24-hour rhythms of RNA expression of nearby genes, with the nadir of methylation preceding peak transcript expression by 1–3 hours. Weak ante-mortem rest-activity rhythms were associated with lower amplitude DNA methylation rhythms as were older age and the presence of Alzheimer's disease. These findings support the hypothesis that 24-hour rhythms of DNA methylation, particularly near transcription start sites, may play a role in driving 24-hour rhythms of gene expression in the human dorsolateral prefrontal cortex, and may be affected by age and Alzheimer's disease

Modeling Disease Severity in Multiple Sclerosis Using Electronic Health Records

Objective: To optimally leverage the scalability and unique features of the electronic health records (EHR) for research that would ultimately improve patient care, we need to accurately identify patients and extract clinically meaningful measures. Using multiple sclerosis (MS) as a proof of principle, we showcased how to leverage routinely collected EHR data to identify patients with a complex neurological disorder and derive an important surrogate measure of disease severity heretofore only available in research settings. Methods: In a cross-sectional observational study, 5,495 MS patients were identified from the EHR systems of two major referral hospitals using an algorithm that includes codified and narrative information extracted using natural language processing. In the subset of patients who receive neurological care at a MS Center where disease measures have been collected, we used routinely collected EHR data to extract two aggregate indicators of MS severity of clinical relevance multiple sclerosis severity score (MSSS) and brain parenchymal fraction (BPF, a measure of whole brain volume). Results: The EHR algorithm that identifies MS patients has an area under the curve of 0.958, 83% sensitivity, 92% positive predictive value, and 89% negative predictive value when a 95% specificity threshold is used. The correlation between EHR-derived and true MSSS has a mean R[superscript 2] = 0.38±0.05, and that between EHR-derived and true BPF has a mean R[superscript 2] = 0.22±0.08. To illustrate its clinical relevance, derived MSSS captures the expected difference in disease severity between relapsing-remitting and progressive MS patients after adjusting for sex, age of symptom onset and disease duration (p = 1.56×10[superscript −12]). Conclusion: Incorporation of sophisticated codified and narrative EHR data accurately identifies MS patients and provides estimation of a well-accepted indicator of MS severity that is widely used in research settings but not part of the routine medical records. Similar approaches could be applied to other complex neurological disorders.National Institute of General Medical Sciences (U.S.) (NIH U54-LM008748

CD33 Alzheimer’s disease locus: Altered monocyte function and amyloid biology

In our functional dissection of the CD33 Alzheimer’s disease susceptibility locus, we find that the rs3865444C risk allele is associated with greater cell surface expression of CD33 in monocytes (t50 = 10.06, pjoint=1.3×10–13) of young and older individuals. It is also associated with (1) diminished internalization of Aβ42) (2) accumulation of neuritic amyloid pathology and fibrillar amyloid on in vivo imaging and (3), increased numbers of activated human microglia

Analysis of the putative role of CR1 in Alzheimer’s disease: Genetic association, expression and function

Chronic activation of the complement system and induced inflammation are associated with neuropathology in Alzheimer's disease (AD). Recent large genome wide association studies (GWAS) have identified single nucleotide polymorphisms (SNPs) in the C3b/C4b receptor (CR1 or CD35) that are associated with late onset AD. Here, anti-CR1 antibodies (Abs) directed against different epitopes of the receptor, were used to localize CR1 in brain, and relative binding affinities of the CR1 ligands, C1q and C3b, were assessed by ELISA. Most Abs tested stained red blood cells in blood vessels but showed no staining in brain parenchyma. However, two monoclonal anti-CR1 Abs labeled astrocytes in all of the cases tested, and this reactivity was preabsorbed by purified recombinant human CR1. Human brain-derived astrocyte cultures were also reactive with both mAbs. The amount of astrocyte staining varied among the samples, but no consistent difference was conferred by diagnosis or the GWAS-identified SNPs rs4844609 or rs6656401. Plasma levels of soluble CR1 did not correlate with diagnosis but a slight increase was observed with rs4844609 and rs6656401 SNP. There was also a modest but statistically significant increase in relative binding activity of C1q to CR1 with the rs4844609 SNP compared to CR1 without the SNP, and of C3b to CR1 in the CR1 genotypes containing the rs6656401 SNP (also associated with the larger isoform of CR1) regardless of clinical diagnosis. These results suggest that it is unlikely that astrocyte CR1 expression levels or C1q or C3b binding activity are the cause of the GWAS identified association of CR1 variants with AD. Further careful functional studies are needed to determine if the variant-dictated number of CR1 expressed on red blood cells contributes to the role of this receptor in the progression of AD, or if another mechanism is involved

GWAS for executive function and processing speed suggests involvement of the CADM2 gene

To identify common variants contributing to normal variation in two specific domains of cognitive functioning, we conducted a genome-wide association study (GWAS) of executive functioning and information processing speed in non-demented older adults from the CHARGE (Cohorts for Heart and Aging Research in Genomic Epidemiology) consortium. Neuropsychological testing was available for 5429-32 070 subjects of European ancestry aged 45 years or older, free of dementia and clinical stroke at the time of cognitive testing from 20 cohorts in the discovery phase. We analyzed performance on the Trail Making Test parts A and B, the Letter Digit Substitution Test (LDST), the Digit Symbol Substitution Task (DSST), semantic and phonemic fluency tests, and the Stroop Color and Word Test. Replication was sought in 1311-21860 subjects from 20 independent cohorts. A significant association was observed in the discovery cohorts for the single-nucleotide polymorphism (SNP) rs17518584 (discovery P-value=3.12 × 10(-8)) and in the joint discovery and replication meta-analysis (P-value=3.28 × 10(-9) after adjustment for age, gender and education) in an intron of the gene cell adhesion molecule 2 (CADM2) for performance on the LDST/DSST. Rs17518584 is located about 170 kb upstream of the transcription start site of the major transcript for the CADM2 gene, but is within an intron of a variant transcript that includes an alternative first exon. The variant is associated with expression of CADM2 in the cingulate cortex (P-value=4 × 10(-4)). The protein encoded by CADM2 is involved in glutamate signaling (P-value=7.22 × 10(-15)), gamma-aminobutyric acid (GABA) transport (P-value=1.36 × 10(-11)) and neuron cell-cell adhesion (P-value=1.48 × 10(-13)). Our findings suggest that genetic variation in the CADM2 gene is associated with individual differences in information processing speed.Molecular Psychiatry advance online publication, 14 April 2015; doi:10.1038/mp.2015.37

Integration of sequence data from a consanguineous family with genetic data from an outbred population identifies PLB1 as a candidate rheumatoid arthritis risk gene

Integrating genetic data from families with highly penetrant forms of disease together with genetic data from outbred populations represents a promising strategy to uncover the complete frequency spectrum of risk alleles for complex traits such as rheumatoid arthritis (RA). Here, we demonstrate that rare, low-frequency and common alleles at one gene locus, phospholipase B1 (PLB1), might contribute to risk of RA in a 4-generation consanguineous pedigree (Middle Eastern ancestry) and also in unrelated individuals from the general population (European ancestry). Through identity-by-descent (IBD) mapping and whole-exome sequencing, we identified a non-synonymous c.2263G>C (p.G755R) mutation at the PLB1 gene on 2q23, which significantly co-segregated with RA in family members with a dominant mode of inheritance (P = 0.009). We further evaluated PLB1 variants and risk of RA using a GWAS meta-analysis of 8,875 RA cases and 29,367 controls of European ancestry. We identified significant contributions of two independent non-coding variants near PLB1 with risk of RA (rs116018341 [MAF = 0.042] and rs116541814 [MAF = 0.021], combined P = 3.2×10-6). Finally, we performed deep exon sequencing of PLB1 in 1,088 RA cases and 1,088 controls (European ancestry), and identified suggestive dispersion of rare protein-coding variant frequencies between cases and controls (P = 0.049 for C-alpha test and P = 0.055 for SKAT). Together, these data suggest that PLB1 is a candidate risk gene for RA. Future studies to characterize the full spectrum of genetic risk in the PLB1 genetic locus are warranted. © 2014 Plenge et al

Alzheimer’s loci: epigenetic associations and interaction with genetic factors

Objective: We explore the role of DNA methylation in Alzheimer’s disease (AD). To elucidate where DNA methylation falls along the causal pathway linking risk factors to disease, we examine causal models to assess its role in the pathology of AD. Methods: DNA methylation profiles were generated in 740 brain samples using the Illumina HumanMet450K beadset. We focused our analysis on CpG sites from 11 AD susceptibility gene regions. The primary outcome was a quantitative measure of neuritic amyloid plaque (NP), a key early element of AD pathology. We tested four causal models: (1) independent associations, (2) CpG mediating the association of a variant, (3) reverse causality, and (4) genetic variant by CpG interaction. Results: Six genes regions (17 CpGs) showed evidence of CpG associations with NP, independent of genetic variation – BIN1 (5), CLU (5), MS4A6A (3), ABCA7 (2), CD2AP (1), and APOE (1). Together they explained 16.8% of the variability in NP. An interaction effect was seen in the CR1 region for two CpGs, cg10021878 (P = 0.01) and cg05922028 (P = 0.001), in relation to NP. In both cases, subjects with the risk allele rs6656401AT/AA display more methylation being associated with more NP burden, whereas subjects with the rs6656401TT protective genotype have an inverse association with more methylation being associated with less NP. Interpretation These observations suggest that, within known AD susceptibility loci, methylation is related to pathologic processes of AD and may play a largely independent role by influencing gene expression in AD susceptibility loci

The Challenge of Examining Social Determinants of Health in People Living With Tourette Syndrome

Background: To examine the association between race, ethnicity, and parental educational attainment on tic-related outcomes among Tourette Syndrome (TS) participants in the Tourette Association of America International Consortium for Genetics (TAAICG) database. Methods: 723 participants in the TAAICG dataset aged ≤21 years were included. The relationships between tic-related outcomes and race and ethnicity were examined using linear and logistic regressions. Parametric and nonparametric tests were performed to examine the association between parental educational attainment and tic-related outcomes. Results: Race and ethnicity were collapsed as non-Hispanic white (N=566, 88.0%) versus Other (N=77, 12.0%). Tic symptom onset was earlier by 1.1 years (P &lt; 0.0001) and TS diagnosis age was earlier by 0.9 years (P = 0.0045) in the Other group (versus non-Hispanic white). Sex and parental education as covariates did not contribute to the differences observed in TS diagnosis age. There were no significant group differences observed across the tic-related outcomes in parental education variable. Conclusions: Our study was limited by the low number of nonwhite or Hispanic individuals in the cohort. Racial and ethnic minoritized groups experienced an earlier age of TS diagnosis than non-Hispanic white individuals. Tic severity did not differ between the two groups, and parental educational attainment did not affect tic-related outcomes. There remain significant disparities and gaps in knowledge regarding TS and associated comorbid conditions. Our study suggests the need for more proactive steps to engage individuals with tic disorders from all racial and ethnic minoritized groups to participate in research studies.</p

Susceptibility to neurofibrillary tangles: role of the PTPRD locus and limited pleiotropy with other neuropathologies

Tauopathies, including Alzheimer’s disease (AD) and other neurodegenerative conditions, are defined by a pathological hallmark: neurofibrillary tangles (NFT). NFT accumulation is thought to be closely linked to cognitive decline in AD. Here, we perform a genome-wide association study for NFT pathologic burden and report the association of the PTPRD locus (rs560380, p=3.8×10−8) in 909 prospective autopsies. The association is replicated in an independent dataset of 369 autopsies. The association of PTPRD with NFT is not dependent on the accumulation of amyloid pathology. In contrast, we find that the ZCWPW1 AD susceptibility variant influences NFT accumulation and that this effect is mediated by an accumulation of amyloid β plaques. We also performed complementary analyses to identify common pathways that influence multiple neuropathologies which co-exist with NFT and found suggestive evidence that certain loci may influence multiple different neuropathological traits, including tau, amyloid β plaques, vascular injury and Lewy bodies. Overall, these analyses offer an evaluation of genetic susceptibility to NFT, a common endpoint for multiple different pathologic processes