Positive Selektion in langlebigen Nagern und kurzlebigen Fischen: eine bioinformatische Suche nach der genetischen Basis des Alterns

Zahlreiche Geschichten und Legenden, angefangen von den frühen Hochkulturen bis hinein in die moderne Populärliteratur, zeugen von dem Wunsch der Menschen das Altern zu verlangsamen. Viele Forscher gehen davon aus, dass genau dies notwendig sein wird, um den seit einigen Jahrzehnten anhaltenden Trend steigender Lebenserwartungen auch in Zukunft fortsetzen zu können und halten es für wahrscheinlich, dass ein verbessertes Verständnis des Alternsprozesses uns dahin führen wird. Um zu einem solchen Verständnis beizutragen, habe ich in dieser Arbeit mit bioinformatischen Mitteln nach der genetischen Basis natürlicherweise Weise vorhandener Unterschiede in der Lebenserwartung verschiedener Spezies gefahndet. Ich bediente mich dabei der Methode der genomweiten Suche nach positiv selektierten Genen (PSGs), die auf dem Vergleich der proteinkodierenden Sequenzen verschiedener Spezies beruht. Dazu habe ich verschiedene Software-Werkzeuge entwickelt und diese in dem Programm PosiGene zusammengefasst. PosiGene ist das erste öffentliche verfügbare Programm, das es ermöglicht PSGs genomweit auf beliebigen, vom Nutzer ausgewählten evolutionären Zweigen zu detektieren. Das erlaubt es allgemein also über die in dieser Arbeit im Fokus stehende Alternsforschung hinaus nach der genetischen Basis speziesspezifischer, phänotypischer Merkmale zu suchen. Die Offenlegung des Quellcodes in GitHub und die Bereitstellung einer installationslos, direkt nach dem Herunterladen einsatzfähigen Software ermöglicht eine breite Anwendung sowie eine kollektive Weiterentwicklung des Programmpakets zum Nutzen der wissenschaftlichen Gemeinschaft (Manuskript I). Konkret untersuchte ich mittels PosiGene zum einen die Verkürzung von Lebensspannen auf evolutionären Zweigen der Prachtgrundkärpflinge (Nothobranchius). Diese zählen mit Lebenserwartungen von z.T. nur sechs Monaten zu den kurzlebigsten Wirbeltierarten überhaupt. Zum anderen untersuchte ich die Verlängerung von Lebensspannen auf Zweigen der Nagetierfamilie der Sandgräber (Bathyergidae), deren Vertreter mit bis zu 30 Jahren um ein Vielfaches älter werden als die meisten anderen Nagetiere. ..

(Anti-)parallel evolution of lifespan

During the evolution vertebrates, a range of lifespans of more than 2000-fold emerged [1]. The identification of the genes that are responsible for this variety could shed new light on the molecular understanding of the aging process that currently derives almost exclusively from single-gene mutations in highly inbred and short-lived laboratory species, mostly invertebrates

Outgroups and Positive Selection: The Nothobranchius furzeri Case

Applications of positive selection analysis increase with the number of species for which genome/transcriptome sequences become available. Using the recently sequenced turquoise killifish (Nothobranchius furzeri) genome as an example, we compare two different approaches based on different outgroup selection. The combination of these two methods allows the origin of positively selected sites in aging-related genes of the N. furzeri genome to be determined

An Analysis of Methylome Evolution in Primates

Although the investigation of the epigenome becomes increasingly important, still little is known about the long-term evolution of epigenetic marks and systematic investigation strategies are still lacking. Here, we systematically demonstrate the transfer of classic phylogenetic methods such as maximum likelihood based on substitution models, parsimony, and distance-based to interval-scaled epigenetic data. Using a great apes blood data set, we demonstrate that DNA methylation is evolutionarily conserved at the level of individual CpGs in promotors, enhancers, and genic regions. Our analysis also reveals that this epigenomic conservation is significantly correlated with its transcription factor binding density. Binding sites for transcription factors involved in neuron differentiation and components of AP-1 evolve at a significantly higher rate at methylation than at the nucleotide level. Moreover, our models suggest an accelerated epigenomic evolution at binding sites of BRCA1, chromobox homolog protein 2, and factors of the polycomb repressor 2 complex in humans. For most genomic regions, the methylation-based reconstruction of phylogenetic trees is at par with sequence-based reconstruction. Most strikingly, phylogenetic reconstruction using methylation rates in enhancer regions was ineffective independently of the chosen model. We identify a set of phylogenetically uninformative CpG sites enriched in enhancers controlling immune-related genes

Parallel evolution of genes controlling mitonuclear balance in short‐lived annual fishes

The current molecular understanding of the aging process derives almost exclusively from the study of random or targeted single‐gene mutations in highly inbred laboratory species, mostly invertebrates. Little information is available as to the genetic mechanisms responsible for natural lifespan variation and the evolution of lifespan, especially in vertebrates. Here, we investigated the pattern of positive selection in annual (i.e., short‐lived) and nonannual (i.e., longer‐lived) African killifishes to identify a genomic substrate for evolution of annual life history (and reduced lifespan). We identified genes under positive selection in all steps of mitochondrial biogenesis: mitochondrial (mt) DNA replication, transcription from mt promoters, processing and stabilization of mt RNAs, mt translation, assembly of respiratory chain complexes, and electron transport chain. Signs of paralleled evolution (i.e., evolution in more than one branch of Nothobranchius phylogeny) are observed in four out of five steps. Moreover, some genes under positive selection in Nothobranchius are under positive selection also in long‐lived mammals such as bats and mole‐rats. Complexes of the respiratory chain are formed in a coordinates multistep process where nuclearly and mitochondrially encoded components are assembled and inserted into the inner mitochondrial membrane. The coordination of this process is named mitonuclear balance, and experimental manipulations of mitonuclear balance can increase longevity of laboratory species. Our data strongly indicate that these genes are also casually linked to evolution lifespan in vertebrates

PosiGene: automated and easy-to-use pipeline for genome-wide detection of positively selected genes

Many comparative genomics studies aim to find the genetic basis of species-specific phenotypic traits. A prevailing strategy is to search genome-wide for genes that evolved under positive selection based on the non-synonymous to synonymous substitution ratio. However, incongruent results largely due to high false positive rates indicate the need for standardization of quality criteria and software tools. Main challenges are the ortholog and isoform assignment, the high sensitivity of the statistical models to alignment errors and the imperative to parallelize large parts of the software. We developed the software tool PosiGene that (i) detects positively selected genes (PSGs) on genome-scale, (ii) allows analysis of specific evolutionary branches, (iii) can be used in arbitrary species contexts and (iv) offers visualization of the results for further manual validation and biological interpretation. We exemplify PosiGene's performance using simulated and real data. In the simulated data approach, we determined a false positive rate <1%. With real data, we found that 68.4% of the PSGs detected by PosiGene, were shared by at least one previous study that used the same set of species. PosiGene is a user-friendly, reliable tool for reproducible genome-wide identification of PSGs and freely available at https://github.com/gengit/PosiGene

Redox-dependent translocation of the heat shock transcription factor AtHSFA8 from the cytosol to the nucleus in Arabidopsis thaliana

Giesguth M, Sahm A, Simon S, Dietz K-J. Redox-dependent translocation of the heat shock transcription factor AtHSFA8 from the cytosol to the nucleus in Arabidopsis thaliana. FEBS letters. 2015;589(6):718-725.The hypothesis is tested that some heat stress transcription factors (HSFs) are activated after formation of inter- or intramolecular disulfide bonds. Based on in silico analyses we identified conserved cysteinyl residues in AtHSFA8 that might function as redox sensors in plants. AtHSFA8 represents a redox-sensitive transcription factor since upon treatment of protoplasts with H2O2 YFP-labeled HSFA8 was translocated to the nucleus in a time-dependent manner. Site-directed mutagenesis of the conserved residues Cys24 and Cys269 blocked translocation of HSFA8 to the nucleus. The findings concur with a model where HSFA8 functions as redox sensing transcription factor within the stress-responsive transcriptional network

Dissecting the DNA binding landscape and gene regulatory network of p63 and p53

The transcription factor p53 is the best-known tumor suppressor, but its sibling p63 is a master regulator of epidermis development and a key oncogenic driver in squamous cell carcinomas (SCC). Despite multiple gene expression studies becoming available, the limited overlap of reported p63-dependent genes has made it difficult to decipher the p63 gene regulatory network. Particularly, analyses of p63 response elements differed substantially among the studies. To address this intricate data situation, we provide an integrated resource that enables assessing the p63-dependent regulation of any human gene of interest. We use a novel iterative de novo motif search approach in conjunction with extensive ChIP-seq data to achieve a precise global distinction between p53 and p63 binding sites, recognition motifs, and potential co-factors. We integrate these data with enhancer:gene associations to predict p63 target genes and identify those that are commonly de-regulated in SCC representing candidates for prognosis and therapeutic interventions

ConCysFind: a pipeline tool to predict conserved amino acids of protein sequences across the plant kingdom

Moore M, Wesemann C, Gossmann N, et al. ConCysFind: a pipeline tool to predict conserved amino acids of protein sequences across the plant kingdom. BMC Bioinformatics. 2020;21(1): 490.Background Post-translational modifications (PTM) of amino acid (AA) side chains in peptides control protein structure and functionality. PTMs depend on the specific AA characteristics. The reactivity of cysteine thiol-based PTMs are unique among all proteinaceous AA. This pipeline aims to ease the identification of conserved AA of polypeptides or protein families based on the phylogenetic occurrence in the plant kingdom. The tool is customizable to include any species. The degree of AA conservation is taken as indicator for structural and functional significance, especially for PTM-based regulation. Further, this pipeline tool gives insight into the evolution of these potentially regulatory important peptides. Results The web-based or stand-alone pipeline tool Conserved Cysteine Finder (ConCysFind) was developed to identify conserved AA such as cysteine, tryptophan, serine, threonine, tyrosin and methionine. ConCysFind evaluates multiple alignments considering the proteome of 21 plant species. This exemplar study focused on Cys as evolutionarily conserved target for multiple redox PTM. Phylogenetic trees and tables with the compressed results of the scoring algorithm are generated for each Cys in the query polypeptide. Analysis of 33 translation elongation and release factors alongside of known redox proteins from Arabidopsis thaliana for conserved Cys residues confirmed the suitability of the tool for identifying conserved and functional PTM sites. Exemplarily, the redox sensitivity of cysteines in the eukaryotic release factor 1-1 (eRF1-1) was experimentally validated. Conclusion ConCysFind is a valuable tool for prediction of new potential protein PTM targets in a broad spectrum of species, based on conserved AA throughout the plant kingdom. The identified targets were successfully verified through protein biochemical assays. The pipeline is universally applicable to other phylogenetic branches by customization of the database