An epigenome-wide association study meta-analysis of educational attainment.

The epigenome is associated with biological factors, such as disease status, and environmental factors, such as smoking, alcohol consumption and body mass index. Although there is a widespread perception that environmental influences on the epigenome are pervasive and profound, there has been little evidence to date in humans with respect to environmental factors that are biologically distal. Here we provide evidence on the associations between epigenetic modifications-in our case, CpG methylation-and educational attainment (EA), a biologically distal environmental factor that is arguably among the most important life-shaping experiences for individuals. Specifically, we report the results of an epigenome-wide association study meta-analysis of EA based on data from 27 cohort studies with a total of 10 767 individuals. We find nine CpG probes significantly associated with EA. However, robustness analyses show that all nine probes have previously been found to be associated with smoking. Only two associations remain when we perform a sensitivity analysis in the subset of never-smokers, and these two probes are known to be strongly associated with maternal smoking during pregnancy, and thus their association with EA could be due to correlation between EA and maternal smoking. Moreover, the effect sizes of the associations with EA are far smaller than the known associations with the biologically proximal environmental factors alcohol consumption, body mass index, smoking and maternal smoking during pregnancy. Follow-up analyses that combine the effects of many probes also point to small methylation associations with EA that are highly correlated with the combined effects of smoking. If our findings regarding EA can be generalized to other biologically distal environmental factors, then they cast doubt on the hypothesis that such factors have large effects on the epigenome.

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.

Genetic effects on information processing speed are moderated by age--converging results from three samples.

Information processing is a cognitive trait forming the basis of complex abilities like executive function. The Trail Making Test (TMT) is a well-established test of information processing with moderate to high heritability. Age of the individual also plays an important role. A number of genetic association studies with the TMT have been performed, which, however, did not consider age as a moderating factor. We report the results of genome-wide association studies (GWASs) on age-independent and age-dependent TMT performance in two population-representative community samples (Munich Antidepressant Response Signature, MARS: N1 = 540; Ludwig Maximilians University, LMU: N2 = 350). Age-dependent genome-wide findings were then evaluated in a third sample of healthy elderly subjects (Sydney Memory and Ageing Study, Sydney MAS: N3 = 448). While a meta-analysis on the GWAS findings did not reveal age-independent TMT associations withstanding correction for multiple testing, we found a genome-wide significant age-moderated effect between variants in the DSG1 gene region and TMT-A performance predominantly reflecting visual processing speed (rs2199301, P(meta-analysis) = 1.3 × 10(-7)). The direction of the interaction suggests for the minor allele a beneficial effect in younger adults turning into a detrimental effect in older adults. The detrimental effect of the missense single nucleotide polymorphism rs1426310 within the same DSG1 gene region could be replicated in Sydney MAS participants aged 70-79, but not in those aged 80 years and older, presumably a result of survivor bias. Our findings demonstrate opposing effects of DSG1 variants on information processing speed depending on age, which might be related to the complex processes that DSG1 is involved with, including cell adhesion and apoptosis.

Memory complaints in a community sample aged 60-64 years: associations with cognitive functioning, psychiatric symptoms, medical conditions, APOE genotype, hippocampus and amygdala volumes, and white-matter hyperintensities.

BACKGROUND: Previous research has found that depression is a major cause of memory complaints. However, there is evidence that memory complaints also weakly predict cognitive decline and dementia. The present study examined a range of possible determinants of memory complaints, covering psychiatric and personality factors, medical history, cognitive test performance, and biological risk factors for dementia (APOE genotype, hippocampus and amygdala volumes, and white-matter hyperintensities). METHOD: A community survey was carried out with 2546 persons aged 60-64 years living in Canberra and Queanbeyan, Australia. Participants were asked about memory problems which interfered with daily life and whether medical help had been sought. A randomly selected subsample of 476 persons was given a brain MRI scan. RESULTS: Participants with memory complaints were found to have poorer memory test performance, more depression and anxiety symptoms, have higher scores on personality traits involving negative affect, and to have worse physical health. Multivariate analyses showed that measures of cognitive performance did not make a unique contribution to the prediction of memory complaints above that of the other categories of predictors. Those with memory complaints did not differ on any of the biological risk factors for dementia. CONCLUSION: In a community sample aged 60-64 years, memory complaints were most closely related to psychiatric symptoms, personality characteristics and poor physical health. There was no evidence of brain changes indicating early dementia.

HLA-H: a pseudogene with increased variation due to balancing selection at neighboring loci.

The HLA complex includes the most polymorphic genes in the human genome. However, the HLA class Ib loci have little, if any, nucleotide variation, presumably due to their specialized functions or perhaps due to a lack of function. This population genetic study of HLA-H, a class I pseudogene, was initiated to determine the pattern of variation at neutral sites within the HLA complex. We found that the pattern of variation for HLA-H is consistent with the neutral model. However, the amount of variation in HLA-H is significantly greater than estimates for other silent sites within the human genome outside of the MHC (theta = 0.0144, P < 0.000001). Our study further indicates that other possible causes of increased variation such as a high mutation rate for HLA-H, interlocus gene conversion, increased diversity in the sample population in general, and selection acting directly on HLA-H are unlikely. Instead, these data suggest that HLA-H has increased variation as a result of balancing selection acting on nearby loci such as HLA-A.

Expression of influenza neuraminidase in baculovirus-infected cells.

Recombinant influenza neuraminidase (NA, subtype 2, A/NT/60/68) was produced by recombinant baculovirus-infected insect cells. The recombinant NA retained enzyme activity and was located on the cell surface. Enzyme activity was both cell-associated and in the cell free supernatant; maximal NA activity was found in the supernatant. Recombinant NA was recognised by polyclonal antisera and by three monoclonal antibodies specific for NA (subtype 2). Enzyme activity was also neutralised by polyclonal antisera. Recombinant NA thus retains most of the immunological and activity properties of authentic influenza NA. Immunoprecipitation of [35S]Methionine-labelled cells and supernatant and partial purification of NA indicated that a approximately 50-kDa form of NA was present in the supernatant, whilst the expected size (approximately 67-kDa) was cell-associated. Purified recombinant extracellular virus was also enzymatically active, and contained the 67-kDa NA which was located on the membrane capsule of the virus. This suggests that the virus had acquired the cell-associated form of recombinant NA during the budding process from the cell.

Characterisation of an avian influenza virus nucleoprotein expressed in E. coli and in insect cells.

The nucleoprotein (NP) gene from influenza virus A/Shearwater/Australia/72 has been expressed intracellularly in both E. coli and insect cells. E. coli-derived NP was identified by Western blot analysis as a 56 kDa protein which co-migrates with virion-derived NP. This protein was purified by immunoaffinity chromatography and a nitrocellulose binding assay showed that NP formed complexes with positive- and negative-sense influenza neuraminidase RNA transcribed in vitro. ELISA and Western blot analysis revealed that recombinant NP of 56 kDa was produced in high yields in insect cells using a baculovirus vector. Immunofluorescence microscopy revealed that NP was localised to the nucleus of infected insect cells.

Structure of the 5' flanking region of the gene encoding human parathyroid-hormone-related protein (PTHrP).

We have characterized a human genomic clone that contains the 5' coding and 5' flanking sequences of the human parathyroid hormone-related protein gene (PTHrP). The 5' end of the gene contains three exons separated by two small introns of 60 and 165 bp, respectively. The coding region of the PTHrP gene exhibits significant structural homology to the human parathyroid hormone gene (PTH), including the position of at least two introns. However, there is no significant nucleotide sequence homology to the PTH gene within the intragenic region nor in the flanking genomic sequences. The PTHrP gene has been localized, by chromosomal in situ hybridization to bands p11 or p12, on human chromosome 12. Analysis of the 5'-noncoding DNA reveals a complex, putative regulatory region, with multiple potential transcription start points. Nucleotide sequence analysis shows the position of one consensus TATA sequence, at -514 bp, from the start of translation whereas the other regulatory domain is located at least 1 kb further 5' to this consensus TATA sequence. Evidence from the structure of a number of cDNA clones, as well as S1 nuclease and primer extension studies supports the hypothesis that the PTHrP gene contains at least two mRNA transcription start points that define two putative regulatory domains. The result of expression from these different promoters combined with an alternative splicing event would be to produce multiple forms of PTHrP mRNA that differ in the 5'-untranslated region. This analysis of the human PTHrP gene is the first report of a PTHrP gene for any species.