A First Step for the Molecular Characterization of Neurological Involvement of Behçet Syndrome: an Italian Pivotal Study

Behçet syndrome (BS) is a vasculitis characterized by several clinical manifestations including the rare neurological involvement (neuro-BS, NBS). The aim of our pivotal study was to investigate the mutational status of several inflammation-related genes in a cohort of Italian patients with and without the neurological involvement (20 NBS vs 40 no-NBS patients). The preliminary in silico single nucleotide polymorphism (SNP) selection and primer design were performed by NCBI Primer-Blast tool. Genomic DNA was isolated and amplified using PCR. PCR amplicons were sequenced and bioinformatically analysed. Twelve tagSNPs were selected and genotyped: ERAP1 rs30187, rs17482078, and rs27044; IL10 rs1800872 and rs1518111, IL12A rs17810546, IL23R rs17375018, IL23R-IL12RB2 rs924080, STAT4 rs7572482, CCR1 rs7616215, KLRC4 rs2617170, and UBAC2 rs3825427. ERAP1 and IL23R SNPs showed statistically significant higher frequencies in NBS group than no-NBS. ERAP1 rs30187 AA was more common in no-NBS patients (20.0% NBS vs 47.5% no-NBS; p < 0.05), while rs17482078 GA frequency was higher in NBS patients (55.0% NBS vs 22.5% no-NBS; p < 0.05, OR: 4.21). IL23R rs17375018 GG was more frequent in NBS group (65.0% NBS vs 40.0% no-NBS; p < 0.05), according to a previous finding. No other statistically significant differences were found. In conclusion, ERAP1 and IL23R SNPs were found associated with neurological involvement of BS. Additional and larger analyses were required to verify our preliminary findings

The reference human nuclear mitochondrial sequences compilation validated and implemented on the UCSC genome browser

<p>Abstract</p> <p>Background</p> <p>Eukaryotic nuclear genomes contain fragments of mitochondrial DNA called NumtS (Nuclear mitochondrial Sequences), whose mode and time of insertion, as well as their functional/structural role within the genome are debated issues. Insertion sites match with chromosomal breaks, revealing that micro-deletions usually occurring at non-homologous end joining <it>loci </it>become reduced in presence of NumtS. Some NumtS are involved in recombination events leading to fragment duplication. Moreover, NumtS are polymorphic, a feature that renders them candidates as population markers. Finally, they are a cause of contamination during human mtDNA sequencing, leading to the generation of false heteroplasmies.</p> <p>Results</p> <p>Here we present RHNumtS.2, the most exhaustive human NumtSome catalogue annotating 585 NumtS, 97% of which were here validated in a European individual and in HapMap samples. The NumtS complete dataset and related features have been made available at the UCSC Genome Browser. The produced sequences have been submitted to INSDC databases. The implementation of the RHNumtS.2 tracks within the UCSC Genome Browser has been carried out with the aim to facilitate browsing of the NumtS tracks to be exploited in a wide range of research applications.</p> <p>Conclusions</p> <p>We aimed at providing the scientific community with the most exhaustive overview on the human NumtSome, a resource whose aim is to support several research applications, such as studies concerning human structural variation, diversity, and disease, as well as the detection of false heteroplasmic mtDNA variants. Upon implementation of the NumtS tracks, the application of the BLAT program on the UCSC Genome Browser has now become an additional tool to check for heteroplasmic artefacts, supported by data available through the NumtS tracks.</p

Genome Digging: Insight into the Mitochondrial Genome of Homo

A fraction of the Neanderthal mitochondrial genome sequence has a similarity with a 5,839-bp nuclear DNA sequence of mitochondrial origin (numt) on the human chromosome 1. This fact has never been interpreted. Although this phenomenon may be attributed to contamination and mosaic assembly of Neanderthal mtDNA from short sequencing reads, we explain the mysterious similarity by integration of this numt (mtAncestor-1) into the nuclear genome of the common ancestor of Neanderthals and modern humans not long before their reproductive split.Exploiting bioinformatics, we uncovered an additional numt (mtAncestor-2) with a high similarity to the Neanderthal mtDNA and indicated that both numts represent almost identical replicas of the mtDNA sequences ancestral to the mitochondrial genomes of Neanderthals and modern humans. In the proteins, encoded by mtDNA, the majority of amino acids distinguishing chimpanzees from humans and Neanderthals were acquired by the ancestral hominins. The overall rate of nonsynonymous evolution in Neanderthal mitochondrial protein-coding genes is not higher than in other lineages. The model incorporating the ancestral hominin mtDNA sequences estimates the average divergence age of the mtDNAs of Neanderthals and modern humans to be 450,000-485,000 years. The mtAncestor-1 and mtAncestor-2 sequences were incorporated into the nuclear genome approximately 620,000 years and 2,885,000 years ago, respectively.This study provides the first insight into the evolution of the mitochondrial DNA in hominins ancestral to Neanderthals and humans. We hypothesize that mtAncestor-1 and mtAncestor-2 are likely to be molecular fossils of the mtDNAs of Homo heidelbergensis and a stem Homo lineage. The d(N)/d(S) dynamics suggests that the effective population size of extinct hominins was low. However, the hominin lineage ancestral to humans, Neanderthals and H. heidelbergensis, had a larger effective population size and possessed genetic diversity comparable with those of chimpanzee and gorilla

Human mtDNA site-specific variability values can act as haplogroup markers

Sequencing of entire human mtDNA genomes has become rapid and efficient, leading to the production of a great number of complete mtDNA sequences from a wide range of human populations. We introduce here a new statistical approach for classifying mtDNA nucleotide sites, simply by comparing the mean simple deviation (MSD) of their specific variability values estimated on continent-specific dataset sequences, without the need for any reference sequence. Excellent correspondence was observed between sites with the highest MSD values and those marking known mtDNA haplogroups. This in turn supports the classification of 81 sites (23 in Africa, eight in Asia, eight in Europe, 34 in Oceania, and eight in America) as novel markers of 47 mtDNA haplogroups not yet identified by phylogeographic studies. Not only does this approach allow refinement of mtDNA phylogeny, an essential requirement also for mitochondrial disease studies, but may greatly facilitate the discrimination of candidate disease-causing mutations from haplogroup-specific polymorphisms in mtDNA sequences of patients affected by mitochondrial disorders

“Human mtDNA site specific variability values can act as haplogroup markers”

Sequencing of entire human mtDNA genomes has become rapid and efficient, leading to the production of a great number of complete mtDNA sequences from a wide range of human populations. We introduce here a new statistical approach for classifying mtDNA nucleotide sites, simply by comparing the mean simple deviation (MSD) of their specific variability values estimated on continent-specific dataset sequences, without the need for any reference sequence. Excellent correspondence was observed between sites with the highest MSD values and those marking known mtDNA haplogroups. This in turn supports the classification of 81 sites (23 in Africa, eight in Asia, eight in Europe, 34 in Oceania, and eight in America) as novel markers of 47mtDNA haplogroups not yet identified by phylogeographic studies. Not only does this approach allow refinement of mtDNA phylogeny, an essential requirement also for mitochondrial disease studies, but may greatly facilitate the discrimination of candidate disease-causing mutations from haplogroup-specific polymorphisms in mtDNA sequences of patients affected by mitochondrial disorders

A data mining approach to retrieve mitochondrial variability data associated to clinical phenotypes

The maintenance of biological databases is at present a problem of great interest since the progress made in many experimental procedures has led to an ever increasing amount of data. These data need to be structured and stored in databases and made accessible to the biological community in user-friendly ways. Although both the interest and the need of accessing biological databases are high, the mechanisms to fund their maintenance are unclear. Funding agencies cannot support data annotation in terms of labour costs and hence the development of new tools based on “data miming” technologies could greatly contribute to keep biological databases updated. Here we present a new approach aimed to contribute to the annotation in the HmtDB resource (http://www.hmdb.uniba.it/) of variability data associated to clinical phenotypes [1]. These data are prevalently available in literature where they are reported in a completely free style. Thus, we suggest the construction of a knowledge base derived from browsing papers on web and to be used in the retrieval phase. Nevertheless, problems in extracting data from literature come not only from the heterogeneity of presentation styles but mainly from the unstructured format (i.e. the natural language) in which they are represented. In this scenario, the goal is to feed a knowledge base by identifying occurrences of specific biological entities and their features as well as the particular method and experimental setting of the scientific study adopted in the publication. In this work, we describe some solutions to the problem of structuring information contained in scientific literature in digital (i.e., pdf) or paper format

Genotyping of Italian patients with Behçet syndrome identified two novel ERAP1 polymorphisms using sequencing-based approach

The endoplasmic reticulum aminopeptidase protein 1 gene (ERAP1) is related to several human diseases, including Behçet syndrome (BS), a multisystemic disorder with unknown etiology. ERAP1 is involved in immune response and its role can be influenced by gene single nucleotide variations (SNVs). We genotyped the ERAP1 whole structure in 50 consecutive BS patients and 50 ethnically-matched healthy controls using both bioinformatics and molecular methodologies. We identified two novel heterozygous missense SNVs of ERAP1 exon3 responsible for the p.Glu183Val and p.Phe199Ser changes. The first variation was recognized in 7/50 (14%) BS patients and involved the substrate binding site (p.Glu183) required for the anchorage of the peptide N-terminal group. The SNV was predicted to be a damaging variation, as well as the p.Phe199Ser substitution (PolyPhen-2 and SIFT on line software). 3D protein structure prediction showed a change in energy score when the wild-type and the variant states were compared, probably influencing the substrate binding and the protein folding. The first variation was associated to a more stable protein chain, while the second polymorphism was related to a less stable protein chain. Our data need to be tested in larger genetic studies