Primary focal hyperhidrosis (PFH, OMIM %144110) is a genetically influenced condition characterised by excessive sweating. Prevalence varies between 1.0–6.1% in the general population, dependent on ethnicity. The aetiology of PFH remains unclear but an autosomal dominant mode of inheritance, incomplete penetrance and variable phenotypes have been reported. In our study, nine pedigrees (50 affected, 53 non-affected individuals) were included. Clinical characterisation was performed at the German Hyperhidrosis Centre, Munich, by using physiological and psychological questionnaires. Genome-wide parametric linkage analysis with GeneHunter was performed based on the Illumina genome-wide SNP arrays. Haplotypes were constructed using easyLINKAGE and visualised via HaploPainter. Whole-exome sequencing (WES) with 100x coverage in 31 selected members (24 affected, 7 non-affected) from our pedigrees was achieved by next generation sequencing. We identified four genome-wide significant loci, 1q41-1q42.3, 2p14-2p13.3, 2q21.2-2q23.3 and 15q26.3-15q26.3 for PFH. Three pedigrees map to a shared locus at 2q21.2-2q23.3, with a genome-wide significant LOD score of 3.45. The chromosomal region identified here overlaps with a locus at chromosome 2q22.1-2q31.1 reported previously. Three families support 1q41-1q42.3 (LOD = 3.69), two families share a region identical by descent at 2p14-2p13.3 (LOD = 3.15) and another two families at 15q26.3 (LOD = 3.01). Thus, our results point to considerable genetic heterogeneity. WES did not reveal any causative variants, suggesting that variants or mutations located outside the coding regions might be involved in the molecular pathogenesis of PFH. We suggest a strategy based on whole-genome or targeted next generation sequencing to identify causative genes or variants for PFH.
The contribution of three genes (C15orf53, OXTR and MLC1) to the etiology of chromosome 15-bound schizophrenia (SCZD10), bipolar disorder (BD) and autism spectrum disorder (ASD) were studied. At first, the uncharacterized gene C15orf53 was comprehensively analyzed. Previous genome-wide association studies (GWAS) in bipolar disorder samples have identified an association signal in close vicinity to C15orf53 on chromosome 15q14. This gene is located in exactly the genomic region, which is segregating in our SCZD10 families. An association study with bipolar disorder (BD) and SCZD10 individual samples did not reveal any association of single nucleotide polymorphisms (SNPs) in C15orf53. Mutational analysis of C15orf53 in SCZD10-affected individuals from seven multiplex families did not show any mutations in the 5'-untranslated region, the coding region and the intron-exon boundaries. Gene expression analysis revealed that C15orf53 was expressed in a subpopulation of leukocytes, but not in human post-mortem limbic brain tissue. Summarizing these studies, C15orf53 is unlikely to be a strong candidate gene for the etiology of BD or SCZD10. The second investigated gene was the human oxytocin receptor gene (OXTR). Five well described SNPs located in the OXTR gene were taken for a transmission-disequilibrium test (TDT) in parents-child trios with ASD-affected children. Neither in the complete sample nor in a subgroup with children that had an intelligence quotient (IQ) above 70, association was found, independent from the application of Haploview or UNPHASED for analysis. The third gene, MLC1, was investigated with regards to its implication in the etiology of SCZD10. Mutations in the MLC1 gene lead to megalencephalic leukoencephalopathy with subcortical cysts (MLC) and one variant coding for the amino acid methionine (Met) instead of leucine (Leu) at position 309 was identified to segregate in a family affected with SCZD10. For further investigation of MLC1 and its possible implication in the etiology of SCZD10, a constitutive Mlc1 knockout mouse model should be created. Mouse embryonic stem cells (mES) were electroporated with a knockout vector construct and analyzed with respect to homologous recombination of the knockout construct with the genomic DNA (gDNA) of the mES. Polymerase chain reaction (PCR) on the available stem cell clones did not reveal any homologous recombined ES. Additionally, we conducted experiments to knockdown MLC1 and using microRNAs. The 3'-untranslated region of the MLC1 gene was analyzed with the bioinformatics tool TargetScan to screen for potential microRNA target sites. In the 3'-untranslated region of the MLC1 gene, a potential binding site for miR-137 was identified. The gene expression level of genes that had been linked to psychiatric disorders and carried a predicated miR-137 binding site has been proven to be immediately responsive to miR-137. Thus, there is new evidence that MLC1 is a candidate gene for the etiology of SCZD10.