A molecular study of larval metamorphosis in the marine bryozoan bugula neritina

The clade lophotrochozoa, the third major branch of bilaterians, is emerging as a model to study the evolutionary history of the Bilaterians. It has been suggested that differential expression of regulatory genes accounts for body plan diversification. However, the molecular mechanisms underlying the development of the majority of lophotrochozoans are largely unknown. This thesis used several different approaches to study the molecular mechanisms of larval metamorphosis of a lophotrochozoan, the marine bryozoan Bugula neritina. To begin with, I defined three metamorphic stages based on the anatomy of the developing juvenile feeding apparatus polypide, and the overall morphology of the pre-ancestrulae. In addition, I described a novel structure in B. neritina, the basal adhesion disc, which may provide mechanical support to the erect pre-ancestrulae. To identify candidate proteins regulating metamorphosis, I performed comparative proteomic analysis on various metamorphic stages using both gel-based and gel-free proteomic techniques. In the gel-based proteomic analysis, I found significant down-regulation of 60 protein spots during early metamorphosis, among which the mitochondrial processing peptidase beta subunit and severin were dramatically down-regulated. In the gel-free proteomic analysis, I identified more than 1100 proteins at each stage, among which 61 were found to be differentially expressed. Proteins involved in energy metabolism and cytoskeleton molecules were generally down-regulated, whereas those involved in transcription and translation, the extracellular matrix and calcification were strongly up-regulated during larval metamorphosis. Spatial expression patterns of differentially expressed proteins such as collagen alpha-2 (I, and IV) and the stimulator myotrophin, carbonic anhydrase II (CA II), Translationally-controlled tumour protein (TCTP), Cathepsin L, leukotriene A-4 hydrolase and temptin-like protein suggested that they may play an important role in the larval metamorphosis. To identify important genes and signaling pathways in the larval metamorphosis, I performed protein domain analysis and annotation enrichment analysis on the B. neritina transcriptome dataset. Genes mapped to Wnt signaling pathways were significantly over-represented in the transcriptome. I then profiled the temporal-spatial expression pattern of genes involved in Wnt signaling pathways. BnWnt10 was expressed spatially opposite to the Wnt antagonist BnsFRP within the blastemas, which is the presumptive polypide. BnWnt6 was expressed in the epidermis. Overall, the findings suggest that the Wnt signaling pathway may be important to the pattern formation of polypide and the development of epidermis