The regulatory role of miRNA in gene expression is an emerging hot new topic in the control of hypometabolism. Sea cucumber aestivation is a complicated physiological process that includes obvious hypometabolism as evidenced by a decrease in the rates of oxygen consumption and ammonia nitrogen excretion, as well as a serious degeneration of the intestine into a very tiny filament. To determine whether miRNAs play regulatory roles in this process, the present study analyzed profiles of miRNA expression in the intestine of the sea cucumber (Apostichopus japonicus), using Solexa deep sequencing technology. We identified 308 sea cucumber miRNAs, including 18 novel miRNAs specific to sea cucumber. Animals sampled during deep aestivation (DA) after at least 15 days of continuous torpor, were compared with animals from a non-aestivation (NA) state (animals that had passed through aestivation and returned to the active state). We identified 42 differentially expressed miRNAs [RPM (reads per million) >10, |FC| (|fold change|) ≥1, FDR (false discovery rate) <0.01] during aestivation, which were validated by two other miRNA profiling methods: miRNA microarray and real-time PCR. Among the most prominent miRNA species, miR-200-3p, miR-2004, miR-2010, miR-22, miR-252a, miR-252a-3p and miR-92 were significantly over-expressed during deep aestivation compared with non-aestivation animals. Preliminary analyses of their putative target genes and GO analysis suggest that these miRNAs could play important roles in global transcriptional depression and cell differentiation during aestivation. High-throughput sequencing data and microarray data have been submitted to GEO database.

Sea cucumbers (Holothuroidea; Echinodermata) have the capacity to regenerate lost tissues and organs. Although the histological and cytological aspects of intestine regeneration have been extensively studied, little is known of the genetic mechanisms involved. There has, however, been a renewed effort to develop a database of Expressed Sequence Tags (ESTs) in Apostichopus japonicus, an economically-important species that occurs in China. This is important for studies on genetic breeding, molecular markers and special physiological phenomena. We have also constructed a library of ESTs obtained from the regenerative body wall and intestine of A. japonicus. The database has increased to ∼30000 ESTs. We used RNA-Seq to determine gene expression profiles associated with intestinal regeneration in A. japonicus at 3, 7, 14 and 21 days post evisceration (dpe). This was compared to profiles obtained from a normally-functioning intestine. Approximately 5 million (M) reads were sequenced in every library. Over 2400 up-regulated genes (>10%) and over 1000 down-regulated genes (∼5%) were observed at 3 and 7dpe (log₂Ratio≥1, FDR≤0.001). Specific “Go terms” revealed that the DEGs (Differentially Expressed Genes) performed an important function at every regeneration stage. Besides some expected pathways (for example, Ribosome and Spliceosome pathway term), the “Notch signaling pathway,” the “ECM-receptor interaction” and the “Cytokine-cytokine receptor interaction” were significantly enriched. We also investigated the expression profiles of developmental genes, ECM-associated genes and Cytoskeletal genes. Twenty of the most important differentially expressed genes (DEGs) were verified by Real-time PCR, which resulted in a trend concordance of almost 100% between the two techniques. Our studies demonstrated dynamic changes in global gene expression during intestine regeneration and presented a series of candidate genes and enriched pathways that contribute to intestine regeneration in sea cucumbers. This provides a foundation for future studies on the genetics/molecular mechanisms associated with intestine regeneration.

The sea cucumber Apostichopus japonicus is a foodstuff with very high economic value in China, Japan and other countries in south-east Asia. It is at the heart of a multibillion-dollar industry and to meet demand for this product, aquaculture methods and facilities have been established. However, there are challenges associated with optimization of reproduction, feeding and growth in non-natural environments. Therefore, we need to learn more about the biology of A. japonicus, including processes such as aestivation, evisceration, regeneration and albinism. One of the major classes of molecules that regulate physiology and behaviour in animals are neuropeptides, and a few bioactive peptides have already been identified in A. japonicus. To facilitate more comprehensive investigations of neuropeptide function in A. japonicus, here we have analysed genomic and transcriptomic sequence data and proteomic data to identify neuropeptide precursors and neuropeptides in this species. We identified 44 transcripts encoding neuropeptide precursors or putative neuropeptide precursors, and in some instances neuropeptides derived from these precursors were confirmed by mass spectrometry. Furthermore, analysis of genomic sequence data enabled identification of the location of neuropeptide precursor genes on genomic scaffolds and linkage groups (chromosomes) and determination of gene structure. Many of the precursors identified contain homologs of neuropeptides that have been identified in other bilaterian animals. Precursors of neuropeptides that have thus far only been identified in echinoderms were identified, including L- and F-type SALMFamides, AN peptides and others. Precursors of several peptides that act as modulators of neuromuscular activity in A. japonicus were also identified. The discovery of a large repertoire of neuropeptide precursors and neuropeptides provides a basis for experimental studies that investigate the physiological roles of neuropeptide signaling systems in A. japonicus. Looking ahead, some of these neuropeptides may have effects that could be harnessed to enable improvements in the aquaculture of this economically important species.