THESIS
2015
xxvi, 279 pages : illustrations (some color) ; 30 cm. + 1 computer disc (color ; 4 3/4 in.)
Abstract
The barnacle Amphibalanus amphitrite is a model biofouling organism. Recently, the p38
MAPK and NO/cGMP pathways were identified to participate in larval settlement of A.
amphitrite. However, the underlying molecular mechanisms are still unknown. This thesis aims
to clarify their mechanisms. Firstly, several MAPK and MKK homologues were expressed and
the following in vitro kinase assay revealed that MKK3 directly activated p38 MAPK. Secondly,
settlement bioassays showed that the NF-κB pathway was required for the larval settlement
process, and kinase assays revealed that NF-κB p65 could be phosphorylated at Ser529 by p38
MAPK activation in cyprids, suggesting that NF-κB p65 might be a down-stream regulator of
the p38 MAPK pathway. Thirdly, proteomic analysis showed that p38 MAPK...[
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The barnacle Amphibalanus amphitrite is a model biofouling organism. Recently, the p38
MAPK and NO/cGMP pathways were identified to participate in larval settlement of A.
amphitrite. However, the underlying molecular mechanisms are still unknown. This thesis aims
to clarify their mechanisms. Firstly, several MAPK and MKK homologues were expressed and
the following in vitro kinase assay revealed that MKK3 directly activated p38 MAPK. Secondly,
settlement bioassays showed that the NF-κB pathway was required for the larval settlement
process, and kinase assays revealed that NF-κB p65 could be phosphorylated at Ser529 by p38
MAPK activation in cyprids, suggesting that NF-κB p65 might be a down-stream regulator of
the p38 MAPK pathway. Thirdly, proteomic analysis showed that p38 MAPK might regulate
multiple functions in A. amphitrite cyprids, including the energy supply, metamorphosis, neural
system and cement glands; and NO might inhibit larval settlement in A. amphitrite by
modulating the process of muscle locomotion and molting. From the proteomic analysis, this
thesis selected several genes, which were regulated by the p38 MAPK and/or NO/cGMP
pathway, for further functional characterization. Among these candidate genes, CUB-serine
protease was localized to the neural system and might take a role in signal detection and
transduction. cAMP-dependent kinase (PKA) was localized to cement glands and body wall of
cyprids, suggesting its possible functions in cementation and molting. Arginine kinase was
localized to muscle tissues in both nauplius and cypris larvae, which might provide burst energy
supply during muscle contraction. A pheromone for conspecific settlement, namely settlement-inducing proteins complex (SIPC), was also regulated by p38 MAPK and NO. In this thesis, a
novel function of SIPC in biomineralization and a novel releasing mechanism of SIPC in
cyprids were described. At last, a new method to transfect siRNA into barnacle larvae was
developed.
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