Luba M. Pardo

The Effect of Genetic Drift in a Young Genetically Isolated Population: Genetic Drift in a Dutch Isolated Population.

The genetic make‐up of genetically isolated populations may differ from a general population as a result of genetic drift and founder effects. We assessed the extent of this deviation in a recently isolated population located in the southwest of the Netherlands and studied as part of the Genetic Research in Isolated Population (GRIP) program. A gene‐dropping experiment was performed in a large pedigree from this isolate, assuming different initial frequencies in the population founders came from. Allelic frequencies in the last generations of this pedigree were estimated. Simulation analysis showed large fluctuations, as measured by variation coefficient and sufficient loss probability, when initial frequencies were lower than or equal to 1%. For initial frequencies larger than 1% the fluctuations were small. We also analyzed mean heterozygosity and allele diversity of 592 markers in a random sample from the GRIP population. The results were compared with a general population (CEPH sample), old large isolate (Icelandic sample) and the small‐sized population of Talana (Sardinia). GRIP mean heterozygosity and mean number of alleles were significantly lower as compared with CEPH and Iceland, but much higher when compared with the Talana population. We also concluded that the findings from the GRIP population for common variants (>1%) are likely to be extendable to other young isolates in Europe as well as to outbred populations.

Mutations in a TGF-β Ligand, TGFB3, Cause Syndromic Aortic Aneurysms and Dissections

Background Aneurysms affecting the aorta are a common condition associated with high mortality as a result of aortic dissection or rupture. Investigations of the pathogenic mechanisms involved in syndromic types of thoracic aortic aneurysms, such as Marfan and Loeys-Dietz syndromes, have revealed an important contribution of disturbed transforming growth factor (TGF)-β signaling. Objectives This study sought to discover a novel gene causing syndromic aortic aneurysms in order to unravel the underlying pathogenesis. Methods We combined genome-wide linkage analysis, exome sequencing, and candidate gene Sanger sequencing in a total of 470 index cases with thoracic aortic aneurysms. Extensive cardiological examination, including physical examination, electrocardiography, and transthoracic echocardiography was performed. In adults, imaging of the entire aorta using computed tomography or magnetic resonance imaging was done. Results Here, we report on 43 patients from 11 families with syndromic presentations of aortic aneurysms caused by TGFB3 mutations. We demonstrate that TGFB3 mutations are associated with significant cardiovascular involvement, including thoracic/abdominal aortic aneurysm and dissection, and mitral valve disease. Other systemic features overlap clinically with Loeys-Dietz, Shprintzen-Goldberg, and Marfan syndromes, including cleft palate, bifid uvula, skeletal overgrowth, cervical spine instability and clubfoot deformity. In line with previous observations in aortic wall tissues of patients with mutations in effectors of TGF-β signaling (TGFBR1/2, SMAD3, and TGFB2), we confirm a paradoxical up-regulation of both canonical and noncanonical TGF-β signaling in association with up-regulation of the expression of TGF-β ligands. Conclusions Our findings emphasize the broad clinical variability associated with TGFB3 mutations and highlight the importance of early recognition of the disease because of high cardiovascular risk.

Sex-specific genetic effects influence variation in body composition

Aims/hypothesisDespite well-known sex differences in body composition it is not known whether sex-specific genetic or environmental effects contribute to these differences.MethodsWe assessed body composition in 2,506 individuals, from a young Dutch genetic isolate participating in the Erasmus Rucphen Family study, by dual-energy X-ray absorptiometry and anthropometry. We used variance decomposition procedures to partition variation of body composition into genetic and environmental components common to both sexes and to men and women separately and calculated the correlation between genetic components in men and women.ResultsAfter accounting for age, sex and inbreeding, heritability ranged from 0.39 for fat mass index to 0.84 for height. We found sex-specific genetic effects for fat percentage (fat%), lean mass, lean mass index (LMI) and fat distribution, but not for BMI and height. Genetic correlations between sexes were significantly different from 1 for fat%, lean mass, LMI, android fat, android:gynoid fat ratio and WHR, indicating that there are sex-specific genes contributing to variation of these traits. Genetic variance was significantly higher in women for the waist, hip and thigh circumference and WHR, implying that genes account for more variance of fat distribution in women than in men. Environmental variance was significantly higher in men for the android:gynoid fat ratio.Conclusions/interpretationSex-specific genetic effects underlie sexual dimorphism in several body composition traits. The findings are relevant for studies on the relationship of body composition with common diseases like cardiovascular disease and type 2 diabetes and for genetic association studies.

The apolipoprotein E gene and its age-specific effects on cognitive function

The E4 allele of the apolipoprotein E gene (APOE) is a well-established determinant of Alzheimers disease but its relation to cognitive function is much less understood. We studied the age-specific effects of the APOE*E4 allele on cognitive function and cardiovascular risk factors in 2208 related individuals. APOE*E4 allele was significantly associated with reduced test scores for Adult Verbal Learning Test, particularly on the memory and learning sub domains, in persons older than 50 years of age. The effect of APOE*E4 was independent of the effect of APOE*E4 on vascular risk factors and most pronounced on learning ability. Our findings suggest that APOE*E4 has an effect on cognitive function predominantly in the elderly, independent of vascular risk factors.

Relationship of the Ubiquilin 1 gene with Alzheimer's and Parkinson's disease and cognitive function

Ubiquilin 1 (UBQLN1) is involved in the ubiquitination machinery, which has been implicated in Alzheimers disease (AD) as well as Parkinsons disease (PD). A polymorphism in the gene encoding for UBQLN1 has been previously associated with a higher risk of AD. We studied the role of the SNP rs12344615 on the UBQLN 1 gene in AD, PD and cognitive function in a population-based study, the Rotterdam Study, and a family-based study embedded in the genetic research in isolated population (GRIP) program. The Rotterdam Study includes 549 patients with AD and 157 patients with PD. The GRIP program includes a series of 123 patients with AD and a study of 1049 persons who are characterized for cognitive function. Data were analysed using logistic and multiple regression analysis. We found no significant difference in risk of AD or PD by the UBQLN1 SNP rs12344615 in our overall and stratified analyses in the Rotterdam Study. In our family-based study, we did not find evidence for linkage of AD to the region including the UBQLN1 gene. In the family-based study we also failed to detect an effect of this polymorphism on cognitive function. Our results suggest that it is unlikely that the SNP rs12344615 of the UBQLN1 gene is related to the onset of AD, PD or cognitive function.

IRF4, MC1R, and TYR genes are risk factors for actinic keratosis independent of skin color

Actinic keratosis (AK) is a pre-malignant skin disease, highly prevalent in elderly Europeans. This study investigates genetic susceptibility to AK with a genome-wide association study (GWAS). A full body skin examination was performed in 3194 elderly individuals from the Rotterdam Study (RS) of exclusive north-western European origin (aged 51-99 years, 45% male). Physicians graded the number of AK into four severity levels: none (76%), 1-3 (14%), 4-9 (6%) and ≥10 (5%), and skin color was quantified using a spectrophotometer on sun-unexposed skin. A GWAS for AK severity was conducted, where promising signals at IRF4 and MC1R (P < 4.2 × 10(-7)) were successfully replicated in an additional cohort of 623 RS individuals (IRF4, rs12203592, Pcombined = 6.5 × 10(-13) and MC1R, rs139810560, Pcombined = 4.1 × 10(-9)). Further, in an analysis of ten additional well-known human pigmentation genes, TYR also showed significant association with AK (rs1393350, P = 5.3 × 10(-4)) after correction for multiple testing. Interestingly, the strength and significance of above-mentioned associations retained largely the same level after skin color adjustment. Overall, our data strongly suggest that IRF4, MC1R and TYR genes likely have pleiotropic effects, a combination of pigmentation and oncogenic functions, resulting in an increased risk of AK.

Dissecting the genetic make-up of North-East Sardinia using a large set of haploid and autosomal markers

Sardinia has been used for genetic studies because of its historical isolation, genetic homogeneity and increased prevalence of certain rare diseases. Controversy remains concerning the genetic substructure and the extent of genetic homogeneity, which has implications for the design of genome-wide association studies (GWAS). We revisited this issue by examining the genetic make-up of a sample from North-East Sardinia using a dense set of autosomal, Y chromosome and mitochondrial markers to assess the potential of the sample for GWAS and fine mapping studies. We genotyped individuals for 500K single-nucleotide polymorphisms, Y chromosome markers and sequenced the mitochondrial hypervariable (HVI–HVII) regions. We identified major haplogroups and compared these with other populations. We estimated linkage disequilibrium (LD) and haplotype diversity across autosomal markers, and compared these with other populations. Our results show that within Sardinia there is no major population substructure and thus it can be considered a genetically homogenous population. We did not find substantial differences in the extent of LD in Sardinians compared with other populations. However, we showed that at least 9% of genomic regions in Sardinians differed in LD structure, which is helpful for identifying functional variants using fine mapping. We concluded that Sardinia is a powerful setting for genetic studies including GWAS and other mapping approaches.

Evidence for novel loci for late-onset Parkinson’s disease in a genetic isolate from the Netherlands

We studied patients with idiopathic Parkinson’s disease (PD) from an isolated population in the Netherlands aiming to map gene(s) involved in PD susceptibility. A total of 109 parkinsonism patients were independently ascertained, of whom 62 presented late-onset, idiopathic PD. Genealogical research showed that 45 index cases with idiopathic PD were linked to a common ancestor, indicating familiar clustering among the patients. This strong familial clustering was highly significant (P=0.005) when compared to random controls from the same population. We performed a genome wide scan using 382 polymorphic markers in 44 distantly related PD patients plus 112 unaffected first-degree relatives and spouses. Our genome wide association analysis (DISLAMB) revealed evidence of association at a nominal P-value<0.01 for markers D2S2333, D4S405, D9S158, D13S153. Other regions on chromosomes 3p, 4q, 14q, 17p and 17q were found at a significance level of P<0.05. In a follow-up study, we investigated all the positive regions using a denser marker set and a larger sample (total of 630 individuals including all late-onset PD patients). The strongest evidence for association remained for the 9q and 14q region. A significant association was found for marker D9S1838 (OR=2.0, 95% CI 1.1–3.5, P=0.014) and D14S65 (OR=3.2, 95% CI 1.7–6.1, P<0.001). Moreover, a common haplotype with excess of sharing among late-onset PD cases was observed on both regions. Our results suggest the existence of two loci influencing PD susceptibility on chromosome 9q and 14q.

Perceived skin colour seems a swift, valid and reliable measurement

DEAR EDITOR, Skin colour is an important trait in dermatological research because it modifies the risk of many skin diseases. In clinical practice dermatologists evaluate skin colour by a quick visual assessment of the sun-unexposed skin. Because visual assessment might be subjective, Fitzpatrick proposed a sun-sensitivity skin-type (Fitzpatrick skin type, FST) classification as a better alternative to quantify skin colour. However, this self-reported skin type might change in time and could be biased because light-skinned individuals tend to overestimate their skin colour. Alternatively, a spectrophotometer measures skin colour objectively, but might be influenced by the seasonal variations and colour inequalities. Visual assessment of perceived skin colour (PSC) allows physicians to combine different clues besides the colour (such as freckles), and to exclude tanning influences, into one skin-colour category. PSC has been used in several studies, but it has not yet been validated. We investigated the reliability of physicians’ perception of skin colour, and how PSC relates to the widely used FST and to unexposed skin colour quantified by a spectrophotometer. This study included adult volunteers visiting the dermatology department of Erasmus MC Rotterdam, the Netherlands, in December 2014 (exclusion criteria: albinism and erythroderma). A sample-size calculation showed 80% power to detect an inter-rater correlation of 0 8 with a maximum deviation of 0 1 when 80 participants were included. The local medical ethics committee approved this study and all participants signed informed consent. Skin colour was assessed using a six-point scale: 1 very white, 2 white, 3 white to olive, 4 light brown, 5 brown and 6 dark brown to black. Three physicians (L.C.J., M.A.H. and J.A.C.V.) independently graded the colour of sun-unexposed skin of the upper body (abdomen and inner upper arm) of each participant, without discussing their assessments. The agreement in grading was tested using the intraclass correlation coefficient (ICC; two-way-mixed, single measures). Subsequently, skin colour was measured as colour saturation at the inner upper arm using a spectrophotometer (CM-600d; Konica-Minolta, Osaka, Japan), and the FST was assessed by combining the answers to the questions: ‘What is your skin colour?’ (type I–IV white, type V brown, type VI black) and, ‘If white, how does your unprotected skin react to sunlight?’ (type I always burns and never tans, type II usually burns and afterwards lightly tans, type III sometimes burns and always tans, type IV never burns and tans deeply). Spearman’s q was used to test the correlation of the spectrophotometer skin colour with PSC and FST. We studied 117 individuals (mean age 45 7 18 6 years, 31% men) of different ancestry. The mean PSC of the three graders included 28 individuals (24%) as very white, 58 (50%) white, 14 (12%) white to olive, six (5%) light brown, 11 (9%) brown and none dark brown to black. The three physicians showed an excellent agreement in grading (ICCabsolute agreement = 0 90). In 114 individuals (three answered ‘unknown’) the self-reported FST included 14 individuals (12%) of type I, 46 (40%) type II, 28 (25%) type III, four (4%) type IV, 22 (19%) type V and none type VI. Colour saturation of unexposed skin ranged from 0 17 (very white) to 0 45 (dark) and was highly correlated with mean PSC (q = 0 82, Fig. 1a, and less so with FST (q = 0 63, Fig. 1b). The FST was generally higher than the mean PSC (mean difference = 0 5, P < 0 001; Fig. 2), but consistency between PSC and FST was still high (ICCconsistency = 0 82). Five individuals were white or white to olive according to the physicians but judged themselves as brown. However, none of the individuals with light brown or brown PSC judged their own skin as white (Fig. 2). After excluding these five individuals, the consistency between PSC and FST increased to an ICC of 0 86. Comparing PSC in younger (≤ 45 years, n = 60) vs. older (> 45 years, n = 57) individuals, a similar agreement between the three physicians was seen (ICCabsolute agreement young = 0 89, ICC old = 0 90). However, the correlation between the mean PSC and the FST was higher in the younger individuals (ICCconsistency young = 0 89, ICC old = 0 76). To assess differences in sun reaction between the present and the teenage years, we asked the volunteers whether their answers to the FST questions would have been different for their teens. Seven individuals reported a higher FST in the past and 11 a lower FST. Most individuals with a higher FST in the past were aged < 45 years (six of seven), and most individuals with a lower FST in the past would have had an FST of I in their teens (seven of 11). Our results show that physician assessment of skin colour is reliable, because minimal interobserver variability was observed. The self-reported FST is higher than the PSC, but largely consistent with it. However, the spectrophotometer colour saturation correlated less with the FST than with the PSC. This suggests that PSC is a mix between true colour (spectrophotometer) and sun sensitivity (FST). Moreover, our data confirm that light-skinned individuals tend to overestimate

Profiling transcription initiation in human aged brain using deep-CAGE

The genome sequencing projects completed in recent years revealed that the number of protein-coding genes does not change appreciably with increasing complexity of the organisms, and it is now generally accepted that this divergence is largely due to variation at the regulatory level. Mechanisms such as alternative splicing, alternative promoters and antisense transcription allow to both obtain a high number of transcripts from a relatively small number of genes and to fine tune isoforms expression in a cell-specific or developmentally-restricted manner. It is likely that the extensive use of such mechanisms plays a pivotal role in development, adult function and ageing of complex tissues like brain. The aim of this study was to characterize transcription start sites (TSSs) in different areas of human aged brain and correlate expression with methylation and structural genomic variation. Since its ability to profile TSSs at high resolution and at a genome wide level, we used Cap Analysis of Gene Expression (CAGE) combined with high-throughput sequencing (deep-CAGE) to collect exact TSSs and their expression levels. We present here our findings on alternative promoters and antisense transcription. Post-mortem tissue from 5 different brain regions was collected from 5 human donors and used to prepare 25 libraries.