STK39 polymorphisms and blood pressure: an association study in British Caucasians and assessment of cis-acting influences on gene expression

<p>Abstract</p> <p>Background</p> <p>Blood pressure (BP) has significant heritability, but the genes responsible remain largely unknown. Single nucleotide polymorphisms (SNPs) at the <it>STK39 </it>locus were recently associated with hypertension by genome-wide association in an Amish population; <it>in vitro </it>data from transient transfection experiments using reporter constructs suggested that altered <it>STK39 </it>expression might mediate the effect. However, other large studies have not implicated <it>STK39 </it>in hypertension. We determined whether reported SNPs influenced <it>STK39 </it>expression <it>in vivo</it>, or were associated with BP in a large British Caucasian cohort.</p> <p>Methods</p> <p>1372 members of 247 Caucasian families ascertained through a hypertensive proband were genotyped for reported risk variants in <it>STK39 </it>(rs6749447, rs3754777, rs35929607) using Sequenom technology. MERLIN software was used for family-based association testing. <it>Cis</it>-acting influences on expression were assessed <it>in vivo </it>using allelic expression ratios in cDNA from peripheral blood cells in 35 South African individuals heterozygous for a transcribed SNP in <it>STK39 </it>(rs1061471) and quantified by mass spectrometry (Sequenom).</p> <p>Results</p> <p>No significant association was seen between the SNPs tested and systolic or diastolic BP in clinic or ambulatory measurements (all p > 0.05). The tested SNPs were all associated with allelic expression differences in peripheral blood cells (p < 0.05), with the most significant association for the intronic SNP rs6749447 (P = 9.9 × 10^-4). In individuals who were heterozygous for this SNP, on average the G allele showed 13% overexpression compared to the T allele.</p> <p>Conclusions</p> <p><it>STK39 </it>expression is modified by polymorphisms acting in <it>cis </it>and the typed SNPs are associated with allelic expression of this gene, but there is no evidence for an association with BP in a British Caucasian cohort.</p

Frequency of rare recessive mutations in unexplained late onset cerebellar ataxia.

Sporadic late onset cerebellar ataxia is a well-described clinical presentation with a broad differential diagnosis that adult neurologists should be familiar with. However, despite extensive clinical investigations, an acquired cause is identified in only a minority of cases. Thereafter, an underlying genetic basis is often considered, even in those without a family history. Here we apply whole exome sequencing to a cohort of 12 patients with late onset cerebellar ataxia. We show that 33% of 'idiopathic' cases harbor compound heterozygous mutations in known ataxia genes, including genes not included on multi-gene panels, or primarily associated with an ataxic presentation

Clinical and functional characterisation of the combined respiratory chain defect in two sisters due to autosomal recessive mutations in MTFMT.

Exome sequencing identified compound heterozygous mutations in the recently discovered mitochondrial methionyl-tRNA formyltransferase (MTFMT) gene in two sisters with mild Leigh syndrome and combined respiratory chain deficiency. The mutations lead to undetectable levels of the MTFMT protein. Blue native polyacrylamide gel electrophoresis showed decreased complexes I and IV, and additional products stained with complex V antibodies, however the overall steady state level of mt-tRNA(Met) was normal. Our data illustrate that exome sequencing is an excellent diagnostic tool, and its value in clinical medicine is enormous, however it can only be optimally exploited if combined with detailed phenotyping and functional studies

Universal heteroplasmy of human mitochondrial DNA.

Mammalian cells contain thousands of copies of mitochondrial DNA (mtDNA). At birth, these are thought to be identical in most humans. Here, we use long read length ultra-deep resequencing-by-synthesis to interrogate regions of the mtDNA genome from related and unrelated individuals at unprecedented resolution. We show that very low-level heteroplasmic variance is present in all tested healthy individuals, and is likely to be due to both inherited and somatic single base substitutions. Using this approach, we demonstrate an increase in mtDNA mutations in the skeletal muscle of patients with a proofreading-deficient mtDNA polymerase γ due to POLG mutations. In contrast, we show that OPA1 mutations, which indirectly affect mtDNA maintenance, do not increase point mutation load. The demonstration of universal mtDNA heteroplasmy has fundamental implications for our understanding of mtDNA inheritance and evolution. Ostensibly de novo somatic mtDNA mutations, seen in mtDNA maintenance disorders and neurodegenerative disease and aging, will partly be due to the clonal expansion of low-level inherited variants

Accurate mitochondrial DNA sequencing using off-target reads provides a single test to identify pathogenic point mutations.

PURPOSE: Mitochondrial disorders are a common cause of inherited metabolic disease and can be due to mutations affecting mitochondrial DNA or nuclear DNA. The current diagnostic approach involves the targeted resequencing of mitochondrial DNA and candidate nuclear genes, usually proceeds step by step, and is time consuming and costly. Recent evidence suggests that variations in mitochondrial DNA sequence can be obtained from whole-exome sequence data, raising the possibility of a comprehensive single diagnostic test to detect pathogenic point mutations. METHODS: We compared the mitochondrial DNA sequence derived from off-target exome reads with conventional mitochondrial DNA Sanger sequencing in 46 subjects. RESULTS: Mitochondrial DNA sequences can be reliably obtained using three different whole-exome sequence capture kits. Coverage correlates with the relative amount of mitochondrial DNA in the original genomic DNA sample, heteroplasmy levels can be determined using variant and total read depths, and-providing there is a minimum read depth of 20-fold-rare sequencing errors occur at a rate similar to that observed with conventional Sanger sequencing. CONCLUSION: This offers the prospect of using whole-exome sequence in a diagnostic setting to screen not only all protein coding nuclear genes but also all mitochondrial DNA genes for pathogenic mutations. Off-target mitochondrial DNA reads can also be used to assess quality control and maternal ancestry, inform on ethnic origin, and allow genetic disease association studies not previously anticipated with existing whole-exome data sets