Mesoderm is a crucial germ layer that contributes to the complexity of bilaterian animals. In chordates, mesodermal cells are first specified into dorsal notochord to support the body and bilateral somites that later differentiate into muscles. These two mesodermal structures are defining features of chordates and are believed to contribute to a better motility for their tadpole-type larvae comparing to non-chordate larvae that propel by cilia. Studies in various chordate species have demonstrated that fibroblast growth factor (FGF) signaling is essential for notochord development through the activation of brachyury at notochord in vertebrates and ascidian. FGF signaling also regulates somite and muscle development in vertebrates but not the formation of trunk muscle in ascidian. In non-chordate deuterostome such as sea urchin embryos, FGF signaling is required for muscle development, but not the expression of brachyury. The functional differences of FGF signaling in controlling muscle development lead to ambiguity in the ancient role of FGF signaling in deuterostomes, and how FGF signaling had evolved to a new function in controlling notochord development through brachyury in chordates.
In order to understand the role of FGF signaling during deuterostome evolution, we investigated functions of FGF signaling in mesoderm development during embryogenesis and metamorphosis in a non-chordate marine animal Ptychodera flava, an indirect-developing hemichordate that possess larval morphology similar to echinoderms and adult body features that resemble chordates. We have identified five FGF ligands and three FGF receptors in P. flava. Phylogenetic analyses revealed that hemichordates possess a conserved FGF8/17/18 in addition to several putative hemichordate-specific FGFs. Further functional studies showed that the mesodermal cell fate is specified at the early gastrula stage, and then theses cells are differentiated stepwise into the hydroporic canal, the pharyngeal muscle, and the muscle string; notably, formation of the last two muscular structures are regulated by FGF signaling. Moreover, the transcription levels of FGF ligands and receptors were significantly increased during metamorphosis, and augmentation of FGF signaling accelerated the process, suggesting its' role in facilitating the transformation from cilia-driven swimming larvae into muscle-driven worms. These results support the ancestral role of FGF signaling in muscle development in deuterostomes. Further studies are in progress for elucidating how the novel role of FGF signaling in notochord development had evolved from its ancestral role in the lineage leading to chordates.Acknowledgment………………………………………….....…………...……………………i
Abstract…………………………………………………………….....…...……………………..ii
List of Tables Figures……………………………………………....……………………….iv
List of Supplementary tables and Figures ………………….....………………....v
Part 1. Evolutionary developmental biology (Evo-Devo), chordate evolution, and hemichordates…………………….…………………….....……….…..1
Part 2. FGF signaling repertoire of the indirect developing hemichordate Ptychodera flava……………………………………………….......…12
Abstract………………………………………………………………........……………………13
Introduction…………………………………………………………………......……………..14
Materials and Methods………………………………………….……………………….…17
Results and Discussion…..……………………………………..………………………...19
Conclusion……………………………………………………………….......…………………31
Part 3. Reiterative use of FGF signaling in mesoderm development during embryogenesis and metamorphosis in the hemichordate Ptychodera flava………….......……....……....……....……....……....…….........…39
Abstract……………………………........………………………………………………………40
Introduction………………………………......……………………………..…………………41
Materials and Methods……………………………….……………………………………44
Results……………………………………………........….……………………………….……47
Discussion…………………………………………………………………………………68
Conclusion…………………………………………………………………………………70
Part 4. Discussion and conclusion……..……………..…..…………………………76
Discussion ……………...………………………....……………………………………77
Conclusion ……………...…………………………......………………………………79
Part 5. The regulation of Pfbra and future prospect………….….………….81
Preliminary results ……...……………………………….............………………82
Future prospects ……...…………………………...…………………............…85
Appendix……...………………………………………………………………………….........86
Results of trials for optimizing the microinjection technology……87
Microinjection workflow in the hemichordate P. flava……......…...88
References…………………………………………………………………………………....…91
Resume……………………………………………………………………………………........10
