Application of transcriptomics in non-model crustacean species: Understanding of crustacean neurohormonal pathway

Tuan Viet Nguyen

doi:10.25907/00662

The objective of this Thesis research was to advance our understanding of the decapod crustaceans neurohormonal pathways using RNA-sequencing technology and bioinformatics, with a focus on neuropeptides and their receptors. Neuropeptides serve as important components of the neuroendocrine signalling system across Bilateria, regulating many complex developmental processes such as reproduction, sexual maturation, feeding, foraging and growth. Neuropeptides act as ligands, mainly operating through activating G protein-coupled receptors (GPCRs) found on the plasma membrane of target cells. Several studies conducted within the framework of this Thesis delineated the gene expression pattern of neuropeptides and their receptors in transcriptomes of three different decapod crustacean species, and with a focus on their involvement in sexual maturation. These species include the Australian red claw crayfish (Cherax quadricarinatus), the Norway lobster (Nephrops norvegicus) and the Tiger prawn (Penaeus monodon), representing three distinct clades of the decapod order, from two separate infraorders. While the number of known extant decapod crustacean species is finite (~15,000 species), they present a bewildering diversity in life cycles, habitats and morphologies. The three studied species each hold commercial and ecological significance, lending them to explore transcriptomic resources with the aim to cross compare between them and see how the neuroendocrine networks relate to different species-specific adaptation, while identifying the common denominators among all three. Due to the heavy utilization of next generation sequencing technology in this research (i.e. Illumina RNA-seq), the first published chapter of this Thesis outlines what I deem as optimal guidelines for RNA-seq based research in non-model decapod crustaceans. Following that, the second published chapter did characterize the neuropeptide genes expressed in the eyestalk ganglia of C. quadricarinatus and predicted which neuropeptides are involved in sexual maturation. N. norvegicus is commercially the most important crustacean species in Europe, yet very little molecular tools have been established. This gap was addressed by assembling and characterizing a comprehensive reference transcriptome based on 5 tissues of both male and female individuals. Three published chapters were forthcoming from this database through a comparison between (i) males and females, (ii) immature and mature females, and (iii) the identification of a repertoire of neuropeptides and GPCRs supported by spatial-temporal expression. Taken together, these three publications encompass a comprehensive molecular toolkit to better understand neuroendocrine pathways in N. norvegicus and through comparison with other species, also across decapods. Finally, in the P. monodon, an analysis of the transcriptomic changes between pre-vitellogenic and vitellogenic females across 4 tissues was performed, with insights into the neuropeptidome and GPCR repertoire. The use of a hybrid clustering with phylogenetic approach was helpful in addressing several deficiencies in using just a phylogenetic method alone. In conclusion, the findings presented in this Thesis have broadened the knowledgebase of neuropeptides and their GPCRs in decapods, expanding the molecular toolbox for functional studies that may assist in crustacean restocking and maturation/reproduction regulation.

Application of transcriptomics in non-model crustacean species: Understanding of crustacean neurohormonal pathway

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