Molecular Investigation of Larval Development and Metamorphosis in the American Lobster, Homarus americanus.

摘要

The American lobster, Homarus americanus, is a marine decapod crustacean that occurs naturally in the coastal waters of eastern North America. High concentrations of lobster in the Gulf of Maine to the Gulf of St. Lawrence support a significant fishing industry in Atlantic Canada and the northeastern United States. A hard exoskeleton covers and protects H. americanus, which they periodically moult to facilitate growth. H. americanus has a complex life history, including a pelagic larval phase and a benthic adult phase. The larval phase is divided into three morphologically distinct stages, followed by metamorphosis to the postlarval phase. Larvae must moult to progress through each developmental stage. Previous studies have described several morphological, physiological, biochemical and behavioural changes associated with moult, growth and development of H. americanus . However, the mechanisms that regulate these changes are not fully elucidated.;This was the first study to use a 14,592 feature, spotted oligonucleotide microarray, to investigate global gene expression changes during H. americanus larval development. Public molecular databases were used to functionally annotate ∼40% of the EST sequences. A reference design was used to compare stage specific gene expression profiles of larvae and postlarvae from two year classes. A two-way ANOVA was used to determine significant differences in gene expression levels among the stages and year classes. This study found that the expression of 1,851 genes was significantly different among larval stages, although 771 genes had significant interaction between year classes and larval stages. Among the differentially expressed genes, functional annotations indicated that several genes were involved with the immune system, energy regulation and development. Ten target genes of interest were selected, based on functional annotations, for expression verification using a complementary measurement method, reverse transcriptase quantitative polymerase chain reaction (RT-qPCR). Genes with functional annotations related to immunity, energy regulation, gene expression, translation and neural or basement membrane remodeling were selected. Two Phosphoenolpyruvate carboxykinases, Argonaute 2, Ecdysone inducible protein 75 and Procollagen-lysine 2-oxoglutarate 5-dioxygenase 3, were significantly different (P < 0.05) genes of interest among stages when assayed using RT-qPCR. These genes are involved with processes that may greatly impact development such as, translation regulation, gene expression, morphological development and energy metabolism.;With increasing interest in the sustainability of the American lobster fishery, this study has provided an important foundation for future investigations of processes regarding signaling, morphological re-modeling, energy metabolism, and the immune system as they pertain to larval development. Understanding the mechanisms that regulate larval development may give insight into the effects factors such as climate change and anthropomorphic environmental impacts have on early life stages of H. americanus.