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My thesis entitled ‘Characterization of cryptic components of the ancestral vertebrate genome’ aims at reconstructing the changes in DNA that parallel the evolution of vertebrates. The central question is which changes on the genomic level accompany, and maybe even account for, the emergence of phenotypic novelties. This approach is also key to a deeper understanding of the evolution of the human genome. Vertebrates are distinguished from invertebrates by numerous characteristics. Vertebrates are phenotypically characterized, for example, by a complex tripartite brain with integrative centers such as the telencephalon and an embryonic neural crest that contributes to elaborate craniofacial features that enable a predatory lifestyle. On the genomic level, vertebrates are distinguished from other chordates by two rounds of whole genome duplication (2R-WGD) that occurred in the last common ancestor of vertebrates around 525 million years ago. Initially, the ancestral vertebrate possessed four copies of each gene. Some of this redundant genetic material was subsequently deleted from the genome, or genes accumulated mutations and eventually became nonfunctional pseudogenes. A differential gene loss between vertebrate lineages might partly explain the phenotypic differences across vertebrates. My approach focuses on key developmental gene families (Bmp, Pax, Hox and ENC gene families) whose members are not present in all vertebrate lineages.A subproject of this thesis focused on the famous Hox gene family that specifies positional identity along the primary body axis in the early embryo across metazoans. The Hox14 gene was hitherto identified only in a handful of basal vertebrates (shark, lamprey and coelacanth), and I revealed the existence of a Hox14 gene in the Australian lungfish. In addition, I showed that its expression in lungfish, similar to shark and lamprey, is decoupled from the typical ‘Hox-code’.Another subproject involved the Pax6 gene that is considered to be the ‘master control gene’ for eye development throughout bilaterians. My research revealed that its sister gene Pax4, that was hitherto only identified in mammals, also exists in the genomes of teleosts, the coelacanth and some reptiles (turtles and crocodiles). Interestingly, I identified a previously unknown gene, Pax10, that is most likely the third gene of the original gene quartet, including Pax4 and -6, derived from the 2R-WGD. A comparative study including phylogenetic, syntenic and expression analyses of Pax4, -6 and -10 genes in diverse vertebrates shed light on the asymmetric evolution of the Pax4/6/10 class of genes. Based on these results I reconstructed a likely evolutionary scenario that describes the secondary modifications in this gene family.The ectodermal neural cortex (ENC) gene family, whose members are implicated in neurogenesis, is part of the kelch repeat superfamily. My analyses revealed that most vertebrates possess three distinct ENC genes derived from the 2R-WGD suggesting the loss of the forth subtype early in vertebrate evolution. Only eutherians secondarily lost ENC3. A comparison of the ENC1 expression patterns I obtained in shark with ENC1 expression profiles in tetrapods suggests a high level of conservation of developmental roles of this gene. Compared with many other gene families including key developmental regulators, the ENC gene family is unique in that conventional molecular phylogenetic inferences could not identify any obvious invertebrate ortholog. This suggests that the ENC gene family might have been too rapidly evolving to provide sufficient phylogenetic signals marking orthology to their invertebrate counterparts. Such gene families that experienced saltatory evolution likely remain unexplored, and might also have contributed to phenotypic evolution of vertebrates.One aspect of my thesis focused on a recently identified sister gene of the key developmental genes Bmp2 and -4, designated Bmp16. This gene greatly differs from its well-investigated sister genes in two aspects. Firstly, the absence of Bmp16 in many vertebrate lineages (mammals, amphibians and archosaurs) is in stark contrast to the universal presence of Bmp2 and -4 in vertebrate genomes. Secondly, gene expression analyses of Bmp16 in teleosts (zebrafish), chondrichthyans (sharks) and reptiles (anoles) revealed a high degree of evolutionary plasticity that has never been documented for any Bmp2 or -4 gene. By using morpholino-induced knockdown techniques, I investigated to what extent sister genes are capable of compensating for the loss of a functional Bmp16 gene. This approach might allude to why this gene independently got lost at least three times during vertebrate evolution.My thesis reveals recurrent patterns of gene family evolution in vertebrates. My detailed studies of selected gene families describe the dynamics that shaped the gene repertoires of extant vertebrates and thus contributed to phenotypic evolution leading to the biodiversity of vertebrates. |
