Florey Department of Neuroscience and Mental Health - Research Publications
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ItemThe TAM Receptor Tyro3 Regulates Myelination in the Central Nervous System(WILEY, 2017-04)Myelin is an essential component of the mammalian nervous system, facilitating rapid conduction of electrical impulses by axons, as well as providing trophic support to neurons. Within the central nervous system, the oligodendrocyte is the specialized neural cell responsible for producing myelin by a process that is thought to be regulated by both activity dependent and independent mechanisms but in incompletely understood ways. We have previously identified that the protein Gas6, a ligand for a family of tyrosine kinase receptors known as the TAM (Tyro3, Axl, and Mertk) receptors, directly increases oligodendrocyte induced myelination in vitro. Gas6 can bind to and activate all three TAM receptors, but the high level of expression of Tyro3 on oligodendrocytes makes this receptor the principal candidate for transducing the pro-myelinating effect of Gas6. In this study, we establish that in the absence of Tyro3, the pro-myelinating effect of Gas6 is lost, that developmental myelination is delayed and that the myelin produced is thinner than normal. We show that this effect is specific to the myelination process and not due to changes in the proliferation or differentiation of oligodendrocyte precursor cells. We have further demonstrated that the reduction in myelination is due to the loss of Tyro3 on oligodendrocytes, and this effect may be mediated by activation of Erk1. Collectively, our findings indicate the critical importance of Tyro3 in potentiating central nervous system myelination. GLIA 2017 GLIA 2017;65:581-591.
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ItemNo Preview AvailableFine-Mapping the Genetic Association of the Major Histocompatibility Complex in Multiple Sclerosis: HLA and Non-HLA Effects(PUBLIC LIBRARY SCIENCE, 2013-11)The major histocompatibility complex (MHC) region is strongly associated with multiple sclerosis (MS) susceptibility. HLA-DRB1*15:01 has the strongest effect, and several other alleles have been reported at different levels of validation. Using SNP data from genome-wide studies, we imputed and tested classical alleles and amino acid polymorphisms in 8 classical human leukocyte antigen (HLA) genes in 5,091 cases and 9,595 controls. We identified 11 statistically independent effects overall: 6 HLA-DRB1 and one DPB1 alleles in class II, one HLA-A and two B alleles in class I, and one signal in a region spanning from MICB to LST1. This genomic segment does not contain any HLA class I or II genes and provides robust evidence for the involvement of a non-HLA risk allele within the MHC. Interestingly, this region contains the TNF gene, the cognate ligand of the well-validated TNFRSF1A MS susceptibility gene. The classical HLA effects can be explained to some extent by polymorphic amino acid positions in the peptide-binding grooves. This study dissects the independent effects in the MHC, a critical region for MS susceptibility that harbors multiple risk alleles.
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ItemIdentity-by-Descent Mapping to Detect Rare Variants Conferring Susceptibility to Multiple Sclerosis(PUBLIC LIBRARY SCIENCE, 2013-03-05)Genome-wide association studies (GWAS) have identified around 60 common variants associated with multiple sclerosis (MS), but these loci only explain a fraction of the heritability of MS. Some missing heritability may be caused by rare variants that have been suggested to play an important role in the aetiology of complex diseases such as MS. However current genetic and statistical methods for detecting rare variants are expensive and time consuming. 'Population-based linkage analysis' (PBLA) or so called identity-by-descent (IBD) mapping is a novel way to detect rare variants in extant GWAS datasets. We employed BEAGLE fastIBD to search for rare MS variants utilising IBD mapping in a large GWAS dataset of 3,543 cases and 5,898 controls. We identified a genome-wide significant linkage signal on chromosome 19 (LODā=ā4.65; pā=ā1.9Ć10(-6)). Network analysis of cases and controls sharing haplotypes on chromosome 19 further strengthened the association as there are more large networks of cases sharing haplotypes than controls. This linkage region includes a cluster of zinc finger genes of unknown function. Analysis of genome wide transcriptome data suggests that genes in this zinc finger cluster may be involved in very early developmental regulation of the CNS. Our study also indicates that BEAGLE fastIBD allowed identification of rare variants in large unrelated population with moderate computational intensity. Even with the development of whole-genome sequencing, IBD mapping still may be a promising way to narrow down the region of interest for sequencing priority.
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ItemModeling the cumulative genetic risk for multiple sclerosis from genome-wide association data(BMC, 2011)BACKGROUND: Multiple sclerosis (MS) is the most common cause of chronic neurologic disability beginning in early to middle adult life. Results from recent genome-wide association studies (GWAS) have substantially lengthened the list of disease loci and provide convincing evidence supporting a multifactorial and polygenic model of inheritance. Nevertheless, the knowledge of MS genetics remains incomplete, with many risk alleles still to be revealed. METHODS: We used a discovery GWAS dataset (8,844 samples, 2,124 cases and 6,720 controls) and a multi-step logistic regression protocol to identify novel genetic associations. The emerging genetic profile included 350 independent markers and was used to calculate and estimate the cumulative genetic risk in an independent validation dataset (3,606 samples). Analysis of covariance (ANCOVA) was implemented to compare clinical characteristics of individuals with various degrees of genetic risk. Gene ontology and pathway enrichment analysis was done using the DAVID functional annotation tool, the GO Tree Machine, and the Pathway-Express profiling tool. RESULTS: In the discovery dataset, the median cumulative genetic risk (P-Hat) was 0.903 and 0.007 in the case and control groups, respectively, together with 79.9% classification sensitivity and 95.8% specificity. The identified profile shows a significant enrichment of genes involved in the immune response, cell adhesion, cell communication/signaling, nervous system development, and neuronal signaling, including ionotropic glutamate receptors, which have been implicated in the pathological mechanism driving neurodegeneration. In the validation dataset, the median cumulative genetic risk was 0.59 and 0.32 in the case and control groups, respectively, with classification sensitivity 62.3% and specificity 75.9%. No differences in disease progression or T2-lesion volumes were observed among four levels of predicted genetic risk groups (high, medium, low, misclassified). On the other hand, a significant difference (F = 2.75, P = 0.04) was detected for age of disease onset between the affected misclassified as controls (mean = 36 years) and the other three groups (high, 33.5 years; medium, 33.4 years; low, 33.1 years). CONCLUSIONS: The results are consistent with the polygenic model of inheritance. The cumulative genetic risk established using currently available genome-wide association data provides important insights into disease heterogeneity and completeness of current knowledge in MS genetics.
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ItemCommon and Low Frequency Variants in MERTK Are Independently Associated with Multiple Sclerosis Susceptibility with Discordant Association Dependent upon HLA-DRB1*15:01 Status(PUBLIC LIBRARY SCIENCE, 2016-03)Multiple Sclerosis (MS) is a chronic inflammatory demyelinating disease of the central nervous system. The risk of developing MS is strongly influenced by genetic predisposition, and over 100 loci have been established as associated with susceptibility. However, the biologically relevant variants underlying disease risk have not been defined for the vast majority of these loci, limiting the power of these genetic studies to define new avenues of research for the development of MS therapeutics. It is therefore crucial that candidate MS susceptibility loci are carefully investigated to identify the biological mechanism linking genetic polymorphism at a given gene to the increased chance of developing MS. MERTK has been established as an MS susceptibility gene and is part of a family of receptor tyrosine kinases known to be involved in the pathogenesis of demyelinating disease. In this study we have refined the association of MERTK with MS risk to independent signals from both common and low frequency variants. One of the associated variants was also found to be linked with increased expression of MERTK in monocytes and higher expression of MERTK was associated with either increased or decreased risk of developing MS, dependent upon HLA-DRB1*15:01 status. This discordant association potentially extended beyond MS susceptibility to alterations in disease course in established MS. This study provides clear evidence that distinct polymorphisms within MERTK are associated with MS susceptibility, one of which has the potential to alter MERTK transcription, which in turn can alter both susceptibility and disease course in MS patients.
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ItemClosing the case of APOE in multiple sclerosis: no association with disease risk in over 29 000 subjects(BMJ PUBLISHING GROUP, 2012-09)BACKGROUND: Single nucleotide polymorphisms (SNPs) rs429358 (Īµ4) and rs7412 (Īµ2), both invoking changes in the amino-acid sequence of the apolipoprotein E (APOE) gene, have previously been tested for association with multiple sclerosis (MS) risk. However, none of these studies was sufficiently powered to detect modest effect sizes at acceptable type-I error rates. As both SNPs are only imperfectly captured on commonly used microarray genotyping platforms, their evaluation in the context of genome-wide association studies has been hindered until recently. METHODS: We genotyped 12 740 subjects hitherto not studied for their APOE status, imputed raw genotype data from 8739 subjects from five independent genome-wide association studies datasets using the most recent high-resolution reference panels, and extracted genotype data for 8265 subjects from previous candidate gene assessments. RESULTS: Despite sufficient power to detect associations at genome-wide significance thresholds across a range of ORs, our analyses did not support a role of rs429358 or rs7412 on MS susceptibility. This included meta-analyses of the combined data across 13 913 MS cases and 15 831 controls (OR=0.95, p=0.259, and OR 1.07, p=0.0569, for rs429358 and rs7412, respectively). CONCLUSION: Given the large sample size of our analyses, it is unlikely that the two APOE missense SNPs studied here exert any relevant effects on MS susceptibility.
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ItemComparing genotyping algorithms for Illumina's Infinium whole-genome SNP BeadChips(BMC, 2011-03-08)BACKGROUND: Illumina's Infinium SNP BeadChips are extensively used in both small and large-scale genetic studies. A fundamental step in any analysis is the processing of raw allele A and allele B intensities from each SNP into genotype calls (AA, AB, BB). Various algorithms which make use of different statistical models are available for this task. We compare four methods (GenCall, Illuminus, GenoSNP and CRLMM) on data where the true genotypes are known in advance and data from a recently published genome-wide association study. RESULTS: In general, differences in accuracy are relatively small between the methods evaluated, although CRLMM and GenoSNP were found to consistently outperform GenCall. The performance of Illuminus is heavily dependent on sample size, with lower no call rates and improved accuracy as the number of samples available increases. For X chromosome SNPs, methods with sex-dependent models (Illuminus, CRLMM) perform better than methods which ignore gender information (GenCall, GenoSNP). We observe that CRLMM and GenoSNP are more accurate at calling SNPs with low minor allele frequency than GenCall or Illuminus. The sample quality metrics from each of the four methods were found to have a high level of agreement at flagging samples with unusual signal characteristics. CONCLUSIONS: CRLMM, GenoSNP and GenCall can be applied with confidence in studies of any size, as their performance was shown to be invariant to the number of samples available. Illuminus on the other hand requires a larger number of samples to achieve comparable levels of accuracy and its use in smaller studies (50 or fewer individuals) is not recommended.
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ItemAnalysis of immune-related loci identifies 48 new susceptibility variants for multiple sclerosis(NATURE PUBLISHING GROUP, 2013-11)Using the ImmunoChip custom genotyping array, we analyzed 14,498 subjects with multiple sclerosis and 24,091 healthy controls for 161,311 autosomal variants and identified 135 potentially associated regions (P < 1.0 Ć 10(-4)). In a replication phase, we combined these data with previous genome-wide association study (GWAS) data from an independent 14,802 subjects with multiple sclerosis and 26,703 healthy controls. In these 80,094 individuals of European ancestry, we identified 48 new susceptibility variants (P < 5.0 Ć 10(-8)), 3 of which we found after conditioning on previously identified variants. Thus, there are now 110 established multiple sclerosis risk variants at 103 discrete loci outside of the major histocompatibility complex. With high-resolution Bayesian fine mapping, we identified five regions where one variant accounted for more than 50% of the posterior probability of association. This study enhances the catalog of multiple sclerosis risk variants and illustrates the value of fine mapping in the resolution of GWAS signals.
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ItemThe MS Risk Allele of CD40 Is Associated with Reduced Cell-Membrane Bound Expression in Antigen Presenting Cells: Implications for Gene Function(PUBLIC LIBRARY SCIENCE, 2015-06-11)Human genetic and animal studies have implicated the costimulatory molecule CD40 in the development of multiple sclerosis (MS). We investigated the cell specific gene and protein expression variation controlled by the CD40 genetic variant(s) associated with MS, i.e. the T-allele at rs1883832. Previously we had shown that the risk allele is expressed at a lower level in whole blood, especially in people with MS. Here, we have defined the immune cell subsets responsible for genotype and disease effects on CD40 expression at the mRNA and protein level. In cell subsets in which CD40 is most highly expressed, B lymphocytes and dendritic cells, the MS-associated risk variant is associated with reduced CD40 cell-surface protein expression. In monocytes and dendritic cells, the risk allele additionally reduces the ratio of expression of full-length versus truncated CD40 mRNA, the latter encoding secreted CD40. We additionally show that MS patients, regardless of genotype, express significantly lower levels of CD40 cell-surface protein compared to unaffected controls in B lymphocytes. Thus, both genotype-dependent and independent down-regulation of cell-surface CD40 is a feature of MS. Lower expression of a co-stimulator of T cell activation, CD40, is therefore associated with increased MS risk despite the same CD40 variant being associated with reduced risk of other inflammatory autoimmune diseases. Our results highlight the complexity and likely individuality of autoimmune pathogenesis, and could be consistent with antiviral and/or immunoregulatory functions of CD40 playing an important role in protection from MS.
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ItemCeruloplasmin gene-deficient mice with experimental autoimmune encephalomyelitis show attenuated early disease evolution(WILEY, 2014-06)We conducted a microarray study to identify genes that are differentially regulated in the spinal cords of mice with the inflammatory disease experimental autoimmune encephalomyelitis (EAE) relative to healthy mice. In total 181 genes with at least a two-fold increase in expression were identified, and most of these genes were associated with immune function. Unexpectedly, ceruloplasmin (Cp), a ferroxidase that converts toxic ferrous iron to its nontoxic ferric form and also promotes the efflux of iron from astrocytes in the CNS, was shown to be highly upregulated (13.2-fold increase) in EAE spinal cord. Expression of Cp protein is known to be increased in several neurological conditions, but the role of Cp regulation in CNS autoimmune disease is not known. To investigate this, we induced EAE in Cp gene knockout, heterozygous, and wild-type mice. Cp knockout mice were found to have slower disease evolution than wild-type mice (EAE days 13-17; Pā=ā0.05). Interestingly, Cp knockout mice also exhibited a significant increase in the number of astrocytes with reactive morphology in early EAE compared with wild-type mice at the same stage of disease. CNS iron levels were not increased with EAE in these mice. Based on these observations, we propose that an increase in Cp expression could contribute to tissue damage in early EAE. In addition, endogenous CP either directly or indirectly inhibits astrocyte reactivity during early disease, which could also worsen early disease evolution.