Jeremy R.B. Newman

Haplotype Analysis of the PARK 11 Gene, GIGYF2, in Sporadic Parkinson's Disease

Familial Parkinsonism (PARK) genes are strong candidates for conferring susceptibility to common forms of PD. However, most studies to date have provided little evidence that their common variants substantially influence disease risk. Recently, mutations were described in the gene, GIGYF2 (TNRC15), located at the PARK11 locus (2q37.1). Here, we use a haplotype tagging approach to examine common variation in the GIGYF2 gene and PD risk. PD cases (n = 568) and age and gender‐matched control subjects (n = 568) were recruited from three specialist movement disorder clinics in Brisbane (Australia) and the Australian electoral roll. Twelve tagging SNPs were assessed in all subjects and haplotype and genotype associations were explored. Overall our findings suggest that common genetic variants of GIGYF2 do not significantly affect sporadic PD risk in Australian Caucasians.

Common polymorphisms in dystonia-linked genes and susceptibility to the sporadic primary dystonias.

Genes involved in familial dystonia syndromes (DYT genes) are ideal candidates for investigating whether common genetic variants influence the susceptibility to sporadic primary dystonia. To date, there have been few candidate gene studies for primary dystonia and only two DYT genes, TOR1A and THAP1, have been assessed. We therefore employed a haplotype-tagging strategy to comprehensively assess if common polymorphisms in eight DYT genes (TOR1A, TAF1, GCH1, THAP1, MR-1 (PNKD), SGCE, ATP1A3 and PRKRA) confer risk for sporadic primary dystonia. The 230 primary dystonia cases were matched for age and gender to 228 controls, recruited from movement disorder clinics in Brisbane, Australia and the Australian electoral roll. All subjects were genotyped for 56 tagging SNPs and genotype associations were investigated. Modest genotypic associations (P<0.05) were observed for three GCH1 SNPs (rs12147422, rs3759664 and rs10483639) when comparing all cases against controls. Associations were also seen when the cases were stratified based on presentation. Overall, our findings do not support the hypothesis that common TOR1A variants affect susceptibility for sporadic primary dystonia, and that it is unlikely that common variants around the DYT genes confer substantial risk for sporadic primary dystonia. Further work is warranted to follow up the GCH1 SNPs and the subgroup analyses.

Comprehensive assessment of genetic sequence variants in the antioxidant 'master regulator' Nrf2 in idiopathic parkinson's disease

Parkinson’s disease (PD) is a complex neurodegenerative disorder influenced by a combination of genetic and environmental factors. The molecular mechanisms that underlie PD are unknown; however, oxidative stress and impairment of antioxidant defence mechanisms have been implicated as major contributors to disease pathogenesis. Previously, we have reported a PD patient-derived cellular model generated from biopsies of the olfactory mucosa, termed hONS cells, in which the NRF2-mediated antioxidant response pathway genes were among the most differentially-expressed. To date, few studies have examined the role of the NRF2 encoding gene, NFE2L2, and PD. In this study, we comprehensibly assessed whether rare and common NFE2L2 genetic variations modify susceptibility to PD using a large Australian case-control sample (PD=1338, controls=1379). We employed a haplotype-tagging approach that identified an association with the tagging SNP rs2364725 and PD (OR = 0.849 (0.760-0.948), P = 0.004). Further genetic screening in hONS cell lines produced no obvious pathogenic variants in the coding regions of NFE2L2. Finally, we investigated the relationship between xenobiotic exposures and NRF2 function, through gene-environment interactions, between NFE2L2 SNPs and smoking or pesticide exposure. Our results demonstrated a significant interaction between rs2706110 and pesticide exposure (OR = 0.597 (0.393-0.900), P = 0.014). In addition, we were able to identify some age-at-onset modifying SNPs and replicate an ‘early-onset’ haplotype that contains a previously identified ‘functional promoter’ SNP (rs6721961). Our results suggest a role of NFE2L2 genetic variants in modifying PD susceptibility and onset. Our findings also support the utility of testing gene-environment interactions in genetic studies of PD.

Haplotype analysis of the IGF2-INS-TH gene cluster in Parkinson's disease†

Idiopathic Parkinsons disease is a common movement disorder characterized by a loss of dopaminergic neurons in the substantia nigra. Its pathogenesis is postulated to involve complex interactions between genetic susceptibility and environmental exposures. The IGF2‐INS‐TH gene cluster on the telomeric end of human chromosome 11 is a gene rich region expressing several proteins important for dopamine neuron homeostasis. We used a haplotyping approach to determine whether common genetic variation in the IGF2‐INS‐TH cluster influences the risk of idiopathic Parkinsons disease in a Caucasian case–control group recruited from Brisbane, Australia. Three tagging polymorphisms, the SNPs, rs680 and rs689 and the microsatellite, HUMTH01, were genotyped in 215 cases and 215 age‐ and gender‐matched controls. Eight common haplotypes accounted for 91% of the genetic variation in our control group and one haplotype, IGF2‐INS‐TH*6, was significantly under‐represented among the cases with idiopathic Parkinsons disease (OR = 0.42, 95% CI = 0.25–0.72, P‐value = 0.001). Analysis of the individual polymorphisms showed that the IGF2‐rs680 alternate ‘A’ allele accounted for the majority of the protective effect. Our findings suggest that common genetic variants in the IGF2‐INS‐TH cluster modify susceptibility to idiopathic Parkinsons disease.

UBASH3A Mediates Risk for Type 1 Diabetes through Inhibition of T-Cell Receptor-Induced NF-κB Signaling

Although over 40 type 1 diabetes (T1D) risk loci have been mapped in humans, the causative genes and variants for T1D are largely unknown. Here, we investigated a candidate gene in the 21q22.3 risk locus—UBASH3A, which is primarily expressed in T cells where it is thought to play a largely redundant role. Genetic variants in UBASH3A have been shown to be associated with several autoimmune diseases in addition to T1D. However, the molecular mechanism underlying these genetic associations is unresolved. Our study reveals a previously unrecognized role of UBASH3A in human T cells: UBASH3A attenuates the NF-κB signal transduction upon T-cell receptor (TCR) stimulation by specifically suppressing the activation of the IκB kinase complex. We identify novel interactions of UBASH3A with nondegradative polyubiquitin chains, TAK1 and NEMO, suggesting that UBASH3A regulates the NF-κB signaling pathway by an ubiquitin-dependent mechanism. Finally, we show that risk alleles at rs11203203 and rs80054410, two T1D-associated variants in UBASH3A, increase UBASH3A expression in human primary CD4+ T cells upon TCR stimulation, inhibiting NF-κB signaling via its effects on the IκB kinase complex and resulting in reduced IL2 gene expression.

Lack of reproducibility in re-evaluating associations between GCH1 polymorphisms and Parkinson's disease and isolated dystonia in an Australian case–control group

AN US CR IP T AC CE PT ED ACCEPTED MANUSCRIPT Lack of reproducability in re-evaluating associations between GCH1 polymorphisms and Parkinson’s disease and isolated dystonia in an Australian case-control group Jeremy R. B. Newman , Michael Todorovic, Peter A. Silburn , Greg T. Sutherland , George D. Mellick 1. Eskitis Institute for Drug Discovery, Griffith University, Nathan, Queensland, Australia 2. Department of Neurology, Princess Alexandra Hospital, Woolloongabba, Queensland, Australia 3. Department of Neurology, Royal Brisbane and Women’s Hospital, Herston, Queensland, Australia 4. University of Queensland Centre for Clinical Research, School of Medicine, Queensland, Royal Brisbane and Women’s Hospital, Herston, Australia 5. Discipline of Pathology, Sydney Medical School, University of Sydney, Camperdown, New South Wales, Australia

Screening for rare sequence variants in the THAP1 gene in a primary dystonia cohort.

The screening of primary dystonia cases has identified >50 potential disease-causing sequence variants in THAP1, the causative gene for DYT6 dystonia. One-third of these mutations occur in patients with adult-onset forms of dystonia. We report on the identification of a novel variant following a comprehensive examination of the THAP1 gene in a series of adult-onset primary dystonia patients from Queensland, Australia.

Disease-specific biases in alternative splicing and tissue-specific dysregulation revealed by multitissue profiling of lymphocyte gene expression in type 1 diabetes

Genome-wide association studies (GWAS) have identified multiple, shared allelic associations with many autoimmune diseases. However, the pathogenic contributions of variants residing in risk loci remain unresolved. The location of the majority of shared disease-associated variants in noncoding regions suggests they contribute to risk of autoimmunity through effects on gene expression in the immune system. In the current study, we test this hypothesis by applying RNA sequencing to CD4+, CD8+, and CD19+ lymphocyte populations isolated from 81 subjects with type 1 diabetes (T1D). We characterize and compare the expression patterns across these cell types for three gene sets: all genes, the set of genes implicated in autoimmune disease risk by GWAS, and the subset of these genes specifically implicated in T1D. We performed RNA sequencing and aligned the reads to both the human reference genome and a catalog of all possible splicing events developed from the genome, thereby providing a comprehensive evaluation of the roles of gene expression and alternative splicing (AS) in autoimmunity. Autoimmune candidate genes displayed greater expression specificity in the three lymphocyte populations relative to other genes, with significantly increased levels of splicing events, particularly those predicted to have substantial effects on protein isoform structure and function (e.g., intron retention, exon skipping). The majority of single-nucleotide polymorphisms within T1D-associated loci were also associated with one or more cis-expression quantitative trait loci (cis-eQTLs) and/or splicing eQTLs. Our findings highlight a substantial, and previously underrecognized, role for AS in the pathogenesis of autoimmune disorders and particularly for T1D.

Risk factors for idiopathic dystonia in Queensland, Australia

It is currently hypothesised that a combination of genetic and environmental factors underlies the development of idiopathic isolated dystonia (IID). In this study, we examined several possible environmental or other non-genetic factors that may influence the risk for IID in Queensland, Australia. We surveyed several environmental exposures, lifestyle factors, medical and family histories to investigate potential risk factors for IID. Associations between putative risk factors and IID were assessed using a total of 184 dystonia patients and 1048 neurologically-normal control subjects sampled from Queensland between 2005 and 2012. Our analyses revealed that anxiety disorders, depression, tremor, cigarette smoking and head injuries with a loss of consciousness were associated with increased risk for IID (p<0.05), all of which remained statistically significant following an adjustment for multiple hypothesis testing except for depression. We also observed that the risk for dystonia increased with higher cigarette smoking pack-year quartiles in our analyses. Our results suggest possible environmental factors that influence the development of IID and complement the findings of similar dystonia risk factor studies. Further investigation defining the environmental and other non-genetic risk factors for IID may provide insight into the development of the disorder in genetically-susceptible individuals.

Variable selection in omics data: A practical evaluation of small sample sizes

In omics experiments, variable selection involves a large number of metabolites/ genes and a small number of samples (the n < p problem). The ultimate goal is often the identification of one, or a few features that are different among conditions- a biomarker. Complicating biomarker identification, the p variables often contain a correlation structure due to the biology of the experiment making identifying causal compounds from correlated compounds difficult. Additionally, there may be elements in the experimental design (blocks, batches) that introduce structure in the data. While this problem has been discussed in the literature and various strategies proposed, the over fitting problems concomitant with such approaches are rarely acknowledged. Instead of viewing a single omics experiment as a definitive test for a biomarker, an unrealistic analytical goal, we propose to view such studies as screening studies where the goal of the study is to reduce the number of features present in the second round of testing, and to limit the Type II error. Using this perspective, the performance of LASSO, ridge regression and Elastic Net was compared with the performance of an ANOVA via a simulation study and two real data comparisons. Interestingly, a dramatic increase in the number of features had no effect on Type I error for the ANOVA approach. ANOVA, even without multiple test correction, has a low false positive rates in the scenarios tested. The Elastic Net has an inflated Type I error (from 10 to 50%) for small numbers of features which increases with sample size. The Type II error rate for the ANOVA is comparable or lower than that for the Elastic Net leading us to conclude that an ANOVA is an effective analytical tool for the initial screening of features in omics experiments.