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Huntington's disease (HD) is a dominantly inherited neurodegenerative disease caused by an expanded CAG repeat in HTT. Many clinical characteristics of HD such as age at motor onset are determined largely by the size of HTT CAG repeat. However, emerging evidence strongly supports a role for other genetic factors in modifying the disease pathogenesis driven by mutant huntingtin. A recent genome-wide association analysis to discover genetic modifiers of HD onset age provided initial evidence for modifier loci on chromosomes 8 and 15 and suggestive evidence for a locus on chromosome 3. Here, genotyping of candidate single nucleotide polymorphisms in a cohort of 3,314 additional HD subjects yields independent confirmation of the former two loci and moves the third to genome-wide significance at MLH1, a locus whose mouse orthologue modifies CAG length-dependent phenotypes in a Htt-knock-in mouse model of HD. Both quantitative and dichotomous association analyses implicate a functional variant on ∼32% of chromosomes with the beneficial modifier effect that delays HD motor onset by 0.7 years/allele. Genomic DNA capture and sequencing of a modifier haplotype localize the functional variation to a 78 kb region spanning the 3'end of MLH1 and the 5'end of the neighboring LRRFIP2, and marked by an isoleucine-valine missense variant in MLH1. Analysis of expression Quantitative Trait Loci (eQTLs) provides modest support for altered regulation of MLH1 and LRRFIP2, raising the possibility that the modifier affects regulation of both genes. Finally, polygenic modification score and heritability analyses suggest the existence of additional genetic modifiers, supporting expanded, comprehensive genetic analysis of larger HD datasets., (© The Author 2017. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

To investigate epigenetic contributions to Huntington's disease (HD) pathogenesis, we carried out genome-wide mapping of the transcriptional mark, trimethyl-histone H3-lysine 4 (H3K4me3) in neuronal nuclei extracted from prefrontal cortex of HD cases and controls using chromatin immunoprecipitation followed by deep-sequencing. Neuron-specific mapping of the genome-wide distribution of H3K4me3 revealed 136 differentially enriched loci associated with genes implicated in neuronal development and neurodegeneration, including GPR3, TMEM106B, PDIA6 and the Notch signaling genes hairy and enhancer of split 4 (HES4) and JAGGED2, supporting the view that the neuronal epigenome is affected in HD. Importantly, loss of H3K4me3 at CpG-rich sequences on the HES4 promoter was associated with excessive DNA methylation, reduced binding of nuclear proteins to the methylated region and altered expression of HES4 and HES4 targeted genes MASH1 and P21 involved in striatal development. Moreover, hypermethylation of HES4 promoter sequences was strikingly correlated with measures of striatal degeneration and age-of-onset in a cohort of 25 HD brains (r = 0.56, P = 0.006). Lastly, shRNA knockdown of HES4 in human neuroblastoma cells altered MASH1 and P21 mRNA expression and markedly increased mutated HTT-induced aggregates and cell death. These findings, taken together, suggest that epigenetic dysregulation of HES4 could play a critical role in modifying HD disease pathogenesis and severity., (© The Author 2014. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Although family history is a well-established risk factor for Parkinson's disease (PD), fewer than 5% of PD cases can be attributed to known genetic mutations. The etiology for the remainder of PD cases is unclear; however, neuronal accumulation of the protein α-synuclein is common to nearly all patients, implicating pathways that influence α-synuclein in PD pathogenesis. We report a genome-wide significant association (P = 3.97 × 10(-8)) between a polymorphism, rs1564282, in the cyclin-G-associated kinase (GAK) gene and increased PD risk, with a meta-analysis odds ratio of 1.48. This association result is based on the meta-analysis of three publicly available PD case-control genome-wide association study and genotyping from a new, independent Italian cohort. Microarray expression analysis of post-mortem frontal cortex from PD and control brains demonstrates a significant association between rs1564282 and higher α-synuclein expression, a known cause of early onset PD. Functional knockdown of GAK in cell culture causes a significant increase in toxicity when α-synuclein is over-expressed. Furthermore, knockdown of GAK in rat primary neurons expressing the A53T mutation of α-synuclein, a well-established model for PD, decreases cell viability. These observations provide evidence that GAK is associated with PD risk and suggest that GAK and α-synuclein interact in a pathway involved in PD pathogenesis. The GAK protein, a serine/threonine kinase, belongs to a family of proteins commonly targeted for drug development. This, combined with GAK's observed relationship to the levels of α-synuclein expression and toxicity, suggests that the protein is an attractive therapeutic target for the treatment of PD.

The age of onset of Huntington's disease (HD) is determined primarily by the length of the HD CAG repeat mutation, but is also influenced by other modifying factors. Delineating these modifiers is a critical step towards developing validated therapeutic targets in HD patients. The HD CAG repeat is somatically unstable, undergoing progressive length increases over time, particularly in brain regions that are the targets of neurodegeneration. Here, we have explored the hypothesis that somatic instability of the HD CAG repeat is itself a modifier of disease. Using small-pool PCR, we quantified somatic instability in the cortex region of the brain from a cohort of HD individuals exhibiting phenotypic extremes of young and old disease onset as predicted by the length of their constitutive HD CAG repeat lengths. After accounting for constitutive repeat length, somatic instability was found to be a significant predictor of onset age, with larger repeat length gains associated with earlier disease onset. These data are consistent with the hypothesis that somatic HD CAG repeat length expansions in target tissues contribute to the HD pathogenic process, and support pursuing factors that modify somatic instability as viable therapeutic targets.

Genetically precise models of Huntington's disease (HD), Hdh CAG knock-in mice, are powerful systems in which phenotypes associated with expanded HD CAG repeats are studied. To dissect the genetic pathways that underlie such phenotypes, we have generated Hdh(Q111) knock-in mouse lines that are congenic for C57BL/6, FVB/N and 129Sv inbred genetic backgrounds and investigated four Hdh(Q111) phenotypes in these three genetic backgrounds: the intergenerational instability of the HD CAG repeat and the striatal-specific somatic HD CAG repeat expansion, nuclear mutant huntingtin accumulation and intranuclear inclusion formation. Our results reveal increased intergenerational and somatic instability of the HD CAG repeat in C57BL/6 and FVB/N backgrounds compared with the 129Sv background. The accumulation of nuclear mutant huntingtin and the formation of intranuclear inclusions were fastest in the C57BL/6 background, slowest in the 129Sv background and intermediate in the FVB/N background. Inbred strain-specific differences were independent of constitutive HD CAG repeat size and did not correlate with Hdh mRNA levels. These data provide evidence for genetic modifiers of both intergenerational HD CAG repeat instability and striatal-specific phenotypes. Different relative contributions of C57BL/6 and 129Sv genetic backgrounds to the onset of nuclear mutant huntingtin and somatic HD CAG repeat expansion predict that the initiation of each of these two phenotypes is modified by different genes. Our findings set the stage for defining disease-related genetic pathways that will ultimately provide insight into disease mechanism.

The 'expanded' HD CAG repeat that causes Huntington's disease (HD) encodes a polyglutamine tract in huntingtin, which first targets the death of medium-sized spiny striatal neurons. Mitochondrial energetics, related to N-methyl-d-aspartate (NMDA) Ca2+-signaling, has long been implicated in this neuronal specificity, implying an integral role for huntingtin in mitochondrial energy metabolism. As a genetic test of this hypothesis, we have looked for a relationship between the length of the HD CAG repeat, expressed in endogenous huntingtin, and mitochondrial ATP production. In STHdhQ111 knock-in striatal cells, a juvenile onset HD CAG repeat was associated with low mitochondrial ATP and decreased mitochondrial ADP-uptake. This metabolic inhibition was associated with enhanced Ca2+-influx through NMDA receptors, which when blocked resulted in increased cellular [ATP/ADP]. We then evaluated [ATP/ADP] in 40 human lymphoblastoid cell lines, bearing non-HD CAG lengths (9-34 units) or HD-causing alleles (35-70 units). This analysis revealed an inverse association with the longer of the two allelic HD CAG repeats in both the non-HD and HD ranges. Thus, the polyglutamine tract in huntingtin appears to regulate mitochondrial ADP-phosphorylation in a Ca2+-dependent process that fulfills the genetic criteria for the HD trigger of pathogenesis, and it thereby determines a fundamental biological parameter--cellular energy status, which may contribute to the exquisite vulnerability of striatal neurons in HD. Moreover, the evidence that this polymorphism can determine energy status in the non-HD range suggests that it should be tested as a potential physiological modifier in both health and disease.

Spirometric measures of pulmonary function have been shown to be highly heritable and evidence for major genes influencing forced expiratory volume in 1 s (FEV1) and forced vital capacity (FVC) have been reported. A genome scan of pulmonary traits in the Framingham Heart Study identified a region on chromosome 6qter with evidence for linkage to FEV1 and the FEV1/FVC ratio. For this study, additional markers were genotyped in the region to refine the location of linkage and test for association. Variance component linkage analysis was performed using GENEHUNTER, and family-based association tests were performed using FBAT. The chromosome 6 telomeric region provided significant evidence of linkage with the additional markers, resulting in a maximum multipoint LOD score of 5.0 for FEV1 at 184.5 cM. LOD scores for FVC and the FEV1/FVC ratio were also above 1.0 in this region. Evidence for association with FEV1 and FVC was observed with D6S281 at 190 cM. The strongest effect was seen with the 224 allele, which was associated with higher levels of FEV1 and FVC in allele carriers compared with those carrying other alleles. This study supports the presence of a gene influencing pulmonary function on the q-terminus of chromosome 6 in the region of 184 cM (D6S503) to 190 cM (D6S281).

Some studies show that plasma triglyceride (TG) levels are a significant independent risk factor for cardiovascular disease (CVD). TG levels are inversely correlated with high density lipoprotein cholesterol (HDL-C) levels, and their metabolism may be closely interrelated. Therefore, the TG/HDL-C ratio may be a relevant CVD risk factor. Our analysis of families in the Framingham Heart Study gave a genetic heritability estimate for log(TG) of 0.40 and for log(TG/HDL-C) of 0.49, demonstrating an important genetic component for both. A 10 cM genome-wide scan for log(TG) level and log(TG/HDL-C) was carried out for the largest 332 extended families of the Framingham Heart Study (1702 genotyped individuals). The highest multipoint variance component LOD scores obtained for both log(TG) and log(TG/HDL-C) were on chromosome 7 (at 155 cM), where the results for the two phenotypes were 1.8 and 2.5, respectively. The 7q32.3-qter region contains several candidate genes. Four other regions with multipoint LOD scores greater than one were identified on chromosome 3 [LOD score for log(TG/HDL-C) = 1.8 at 140 cM], chromosome 11 [LOD score for log(TG/HDL-C) = 1.1 at 125 cM], chromosome 16 [LOD score for log(TG) = 1.5 at 70 cM, LOD score for log(TG/HDL-C) = 1.1 at 75 cM] and chromosome 20 [LOD score for log(TG/HDL-C) = 1.7 at 35 cM, LOD score for log(TG) = 1.3 at 40 cM]. These results identify loci worthy of further study.

Controversy persists concerning the significance of Huntington disease (HD) alleles in the 36-39 repeat range. Although some clinically affected persons have been documented with repeats in this range, elderly unaffected individuals have also been reported. We examined 10 paternal transmissions of HD alleles of 37-39 repeats in collateral branches of families with de novo HD. All 10 descendants, including many who are elderly, are without symptoms of HD. Forty percent of the transmissions were unstable, although none varied by more than one repeat. The observation that individuals with alleles of 37-39 repeats may survive unaffected beyond common life expectancy supports the presence of reduced penetrance for HD among some persons with repeat sizes which overlap the clinical range. Non-penetrance may be increased in the collateral branches of de novo mutation families when compared to penetrance estimates from patient series. There was no CAA-->CAG mutation for the penultimate glutamine in either a de novo expanded 42 repeat allele or the corresponding non-penetrant 38 repeat allele in a family with fresh mutation to HD.

The CAG triplet repeat region of the Huntington's disease gene was amplified in 923 single sperm from three affected and two normal individuals. Average-size alleles (15-18 repeats) showed only three contraction mutations among 475 sperm (0.6%). A 30 repeat normal allele showed an 11% mutation frequency. The mutation frequency of a 36 repeat intermediate allele was 53% with 8% of all gametes having expansions which brought the allele size into the HD disease range (> or = 38 repeats). Disease alleles (38-51 repeats) showed a very high mutation frequency (92-99%). As repeat number increased there was a marked elevation in the frequency of expansions, in the mean number of repeats added per expansion and the size of the largest observed expansion. Contraction frequencies also appeared to increase with allele size but decreased as repeat number exceeded 36. Our sperm typing data are of a discrete nature rather than consisting of smears of PCR product from pooled sperm. This allowed the observed mutation frequency spectra to be compared to the distribution calculated using discrete stochastic models based on current molecular ideas of the expansion process. An excellent fit was found when the model specified that a random number of repeats are added during the progression of the polymerase through the repeated region.