The attachment and metamorphosis is a critical process in the life of many marine invertebrates, through which a swimming larva undergoes drastic anatomic and physiological changes to develop into a juvenile. The lack of comprehensive genetic information on may marine invertebrate restricts us from understanding the detailed mechanism of this critical process at the molecular level....[ Read more ]

The attachment and metamorphosis is a critical process in the life of many marine invertebrates, through which a swimming larva undergoes drastic anatomic and physiological changes to develop into a juvenile. The lack of comprehensive genetic information on may marine invertebrate restricts us from understanding the detailed mechanism of this critical process at the molecular level.

In this thesis work, I first used suppressive subtractive hybridization technology studied the competency, attachment and metamorphosis of the marine fouling polychaete H. elegans. In the competency screen, twenty-one genes which were up regulated during the competency of H. elegans were isolated. Among these genes, a novel putative novel p38 Mitogen activate protein kinase was characterized. With bioassay, the possible involvement of the reactive oxygen species in controlling larval settlement was also demonstrated. In the following attachment and metamorphosis SSH screen, the genes differentially expressed between competent larvae and metamorphosing juvenile were isolated. Ninety-seven genes were isolated from the forward hybridization library, while 121 genes were isolated from the reversed hybridization library. Following-up real-time PCR, in situ hybridization, and bio-assay revealed the putative roles of some of the isolated genes in the attachment and metamorphosis.

Furthermore, we applied transcriptome profiling on two marine fouling species, the bryozoan Bugula neritina and the polychaete Hydroides elegans by using the titanium 454 GS FLX platform. From the B. neritina, we generated a total 48 Mbp of transcriptome data from the cDNA pool of individuals undergoing metamorphosis. After de novo assembling, 122,650 of the 131,450 high quality reads formed 6,392 contigs of average length of 538 bases. The remaining 8,800 reads remained as singletons. Among the 15,192 unigenes, 13,863 open reading frames (ORF) were predicted, and 6,917 ORFs were annotated with function. Using the annotated transcriptome as a reference and combined with LC-MS based proteome analysis, we further identified and quantified 882 proteins from B. neritina. For the H. elegans transcriptome profiling, the cDNA samples from four developmental stages, including pre-competent larvae, competent larvae, male and female adults, were sequenced by using the 454 pyrosequencing platform. After de novo assembly, 136,490 ORFs were predicted, 38,259 of them could be matched to known genes on NCBI database whereas no homolog was found for the other 98,231 ORFs. The pyrosequencing result of H. elegans revealed the existence of putative spliced-leader in H. elegans. In addition, a large number of genes and pathways in H. elegans that may control development were also found. This result for the first time demonstrated the utility that next generation sequencing technology for generating high throughput transcriptome data to advance the study of molecular mechanism regulating attachment and metamorphosis in non-model species.