Differential gene expression during the moult cycle of Antarctic krill (Euphausia superba)

Abstract

BackgroundAll crustaceans periodically moult to renew their exoskeleton. In krill this involves partial digestion and resorption of the old exoskeleton and synthesis of new cuticle. Molecular events that underlie the moult cycle are poorly understood in calcifying crustaceans and even less so in non-calcifying organisms such as krill. To address this we constructed an Antarctic krill cDNA microarray in order to generate gene expression profiles across the moult cycle and identify possible activation pathways.ResultsA total of 26 different cuticle genes were identified that showed differential gene expression across the moult cycle. Almost all cuticle genes were up regulated during premoult and down regulated during late intermoult. There were a number of transcripts with significant sequence homology to genes potentially involved in the synthesis, breakdown and resorption of chitin. During early premoult glutamine synthetase, a gene involved in generating an amino acid used in the synthesis of glucosamine, a constituent of chitin, was up regulated more than twofold. Mannosyltransferase 1, a member of the glycosyltransferase family of enzymes that includes chitin synthase was also up regulated during early premoult. Transcripts homologous to a β-N-acetylglucosaminidase (β-NAGase) precursor were expressed at a higher level during late intermoult (prior to apolysis) than during premoult. This observation coincided with the up regulation during late intermoult, of a coatomer subunit epsilon involved in the production of vesicles that maybe used to transport the β-NAGase precursors into the exuvial cleft. Trypsin, known to activate the β-NAGase precursor, was up regulated more than fourfold during premoult. The up regulation of a predicted oligopeptide transporter during premoult may allow the transport of chitin breakdown products across the newly synthesised epi- and exocuticle layers.ConclusionWe have identified many genes differentially expressed across the moult cycle of krill that correspond with known phenotypic structural changes. This study has provided a better understanding of the processes involved in krill moulting and how they may be controlled at the gene expression level.

Seasonal and spatial influences on gene expression in Antarctic krill Euphausia superba

“…In the laboratory, the heads were taken for RNA extraction (Buchholz 1982) that were female but yet to mature fully (stage IIB or IIIA) (Makarov & Denys 1980). This step was carried out to avoid gene expression differences between sexes or moult stages from dominating the comparison (see Seear et al 2010). Altogether, 126 krill were examined, from which 37 krill suited the above criteria: 11 from Peninsula winter samples, 18 from Peninsula summer samples, and 8 from South Georgia winter samples.…”

“…This was later followed up by sequencing of a further 8 clones from the South Georgia winter cDNA library found to be differentially expressed in the microarray study. Sequencing was performed following procedures described in Seear et al (2010). Sequence chromatograms were processed using Geneious 5.3 (Drummond et al 2010) that trimmed vector, low quality sequence and 3'-polyadenylate tails.…”

“…The microarray was populated with 4792 transcripts from krill caught in the summer from 2 latitudes (55°S and 60°S) at various times of the day (Seear et al 2010), along with 960 transcripts from krill caught during the winter at Peninsula (62°S) and 960 transcripts from krill caught at South Georgia (53.5°S) during the same winter. PCR amplification of clone inserts from the winter cDNAs, quality control and microarray printing were performed as described in Seear et al (2010).…”

“…PCR amplification of clone inserts from the winter cDNAs, quality control and microarray printing were performed as described in Seear et al (2010). In total, 13 440 features were printed on the microarray, including 6712 clones from summer and winter krill cDNA libraries spotted in duplicate.…”

“…They have also revealed much about internal processes of crusta ceans such as reproduction (Nagaraju 2011), immune response (Aoki et al 2011, inducible defense (Tollrian & Leese 2010), diapause (MacRae 2010) and starvation (Lenz et al 2012). In Antarctic krill, Seear et al (2010) used cDNA microarray techniques to examine differential gene expression during successive phases of the moult cycle. Clark et al (2011) applied 454 pyro sequencing and found high levels of expression of classical 'stress proteins' such as HSP70, HSP90, ferritin and GST in Antarctic krill.…”