No abstract

As an important transcription factor, SOX2 involves in embryogenesis, maintenance of stem cells and proliferation of primordial germ cell (PGC). However, little was known about its function in mature gonads. Herein, we investigated the SOX2 gene profiles in testis of scallop, Chlamys farreri. The level of C. farreri SOX2 (Cf-SOX2) mRNA increased gradually along with gonadal development and reached the peak at mature stage, and was located in all germ cells, including spermatogonia, spermatocytes, spermatids and spermatozoa. Knockdown of Cf-SOX2 using RNAi leaded to a mass of germ cells lost, and only a few spermatogonia retained in the nearly empty testicular acini after 21 days. TUNEL assay showed that apoptosis occurred in spermatocytes. Furthermore, transcriptome profiles of the testes were compared between Cf-SOX2 knockdown and normal scallops, 131,340 unigenes were obtained and 2,067 differential expression genes (DEGs) were identified. GO and KEGG analysis showed that most DEGs were related to cell apoptosis (casp2, casp3, casp8), cell proliferation (samd9, crebzf, iqsec1) and spermatogenesis (htt, tusc3, zmynd10, nipbl, mfge8), and enriched in p53, TNF and apoptosis pathways. Our study revealed Cf-SOX2 is essential in spermatogenesis and testis development of C. farreri and provided important clues for better understanding of SOX2 regulatory mechanisms in bivalve testis.

The larval segment formation and secondary loss in echiurans is a special phenomenon, which is considered to be one of the important characteristics in the evolutionary relationship between the Echiura and Annelida. To better understand the molecular mechanism of this phenomenon, we revealed the larval transcriptome profile of the echiuran worm Urechis unicinctus using RNA-Seq technology. Twelve cDNA libraries of U. unicinctus larvae, late-trochophore (LT), early-segmentation larva (ES), segmentation larva (SL), and worm-shaped larva (WL) were constructed. Totally 243,381 unigenes were assembled with an average length of 1125 bp and N50 of 1836 bp, and 149,488 unigenes (61.42%) were annotated. We obtained 70,517 differentially expressed genes (DEGs) by pairwise comparison of the larval transcriptome data at different developmental stages and clustered them into 20 gene expression profiles using STEM software. Based on the typical profiles during the larval segment formation and secondary loss, eight signaling pathways were enriched, and five of which, mTOR, PI3K-AKT, TGF-β, MAPK, and Dorso-ventral axis formation signaling pathway, were proposed for the first time to be involved in the segment formation. Furthermore, we identified 119 unigenes related to the segment formation of annelids, arthropods, and chordates, in which 101 genes were identified in Drosophila and annelids. The function of most segment polarity gene homologs (hedgehog, wingless, engrailed, etc.) was conserved in echiurans, annelids, and arthropods based on their expression profiles, while the gap and pair-rule gene homologs were not. Finally, we verified that strong positive signals of Hedgehog were indeed located on the boundary of larval segments using immunofluorescence. Data in this study provide molecular evidence for the understanding of larval segment development in echiurans and may serve as a blueprint for segmented ancestors in future research.

In many bilaterians, Hox genes are generally clustered along the chromosomes and expressed in spatial and temporal order. In vertebrates, the expression of Hox genes follows a whole-cluster spatio-temporal collinearity (WSTC) pattern, whereas in some invertebrates the expression of Hox genes exhibits a subcluster-level spatio-temporal collinearity pattern. In bilaterians, the diversity of collinearity patterns and the cause of collinearity differences in Hox gene expression remain poorly understood. Here, we investigate genomic organization and expression pattern of Hox genes in the echiuran worm Urechis unicinctus (Annelida, Echiura). Urechis unicinctus has a split cluster with four subclusters divided by non-Hox genes: first subcluster ( Hox1 and Hox2 ), second subcluster ( Hox3 ), third subcluster ( Hox4 , Hox5 , Lox5 , Antp and Lox4 ), fourth subcluster ( Lox2 and Post2 ). The expression of U. unicinctus Hox genes shows a subcluster-based whole-cluster spatio-temporal collinearity (S-WSTC) pattern: the anterior-most genes in each subcluster are activated in a spatially and temporally colinear manner (reminiscent of WSTC), with the subsequent genes in each subcluster then being very similar to their respective anterior-most subcluster gene. Combining genomic organization and expression profiles of Hox genes in different invertebrate lineages, we propose that the spatio-temporal collinearity of invertebrate Hox is subcluster-based.

Background In marine invertebrate life cycles, which often consist of planktonic larval and benthonic adult stages, settlement of the free-swimming larva to the sea floor in response to environmental cues is a key life cycle transition. Settlement is regulated by a specialized sensory–neurosecretory system, the larval apical organ. The neuroendocrine mechanisms through which the apical organ transduces environmental cues into behavioral responses during settlement are not fully understood yet. Results In this study, a total of 54 neuropeptide precursors (pNPs) were identified in the Urechis unicinctus larva and adult transcriptome databases using local BLAST and NpSearch prediction, of which 10 pNPs belonging to the ancient eumetazoa, 24 pNPs belonging to the ancient bilaterian, 3 pNPs belonging to the ancient protostome, 9 pNPs exclusive in lophotrochozoa, 3 pNPs exclusive in annelid, and 5 pNPs only found in U. unicinctus. Furthermore, four pNPs (MIP, FRWamide, FxFamide and FILamide) which may be associated with the settlement and metamorphosis of U. unicinctus larvae were analysed by qRT-PCR. Whole-mount in situ hybridization results showed that all the four pNPs were expressed in the region of the apical organ of the larva, and the positive signals were also detected in the ciliary band and abdomen chaetae. We speculated that these pNPs may regulate the movement of larval cilia and chaeta by sensing external attachment signals. Conclusions This study represents the first comprehensive identification of neuropeptides in Echiura, and would contribute to a complete understanding on the roles of various neuropeptides in larval settlement of most marine benthonic invertebrates.

No abstract

The intertidal zone is a transitional area of the land-sea continuum, in which physical and chemical properties vary during the tidal cycle and highly toxic sulfides are rich in sediments due to the dynamic regimes. As a typical species thriving in this habitat, Urechis unicinctus presents strong sulfide tolerance and is expected to be a model species for sulfide stress research. Heat shock proteins (HSPs) consist of a large group of highly conserved molecular chaperones, which play important roles in stress responses. In this study, we systematically analyzed the composition and expression of HSPs in U. unicinctus. A total of eighty-six HSP genes from seven families were identified, in which two families, including sHSP and HSP70, showed moderate expansion, and this variation may be related to the benthic habitat of the intertidal zone. Furthermore, expression analysis revealed that almost all the HSP genes in U. unicinctus were significantly induced under sulfide stress, suggesting that they may be involved in sulfide stress response. Weighted gene co-expression network analysis (WGCNA) showed that 12 HSPs, including 5 sHSP and 4 HSP70 family genes, were highly correlated with the sulfide stress response which was distributed in steelblue and green modules. Our data indicate that HSPs, especially sHSP and HSP70 families, may play significant roles in response to sulfide stress in U. unicinctus. This systematic analysis provides valuable information for further understanding of the function of the HSP gene family for sulfide adaptation in U. unicinctus and contributes a better understanding of the species adaptation strategies of marine benthos in the intertidal zone.

Background: In marine invertebrate life cycles, which often consist of planktonic larval and benthonic adult stages, settlement of the free-swimming larva to the sea floor in response to environmental cues is a key life cycle transition. Settlement is regulated by a specialized sensory–neurosecretory system, the larval apical organ. The neuroendocrine mechanisms through which the apical organ transduces environmental cues into behavioral responses during settlement are not fully understood yet.Results: In this study, a total of 54 neuropeptide precursors (pNPs) were identified in the Urechis unicinctus larva and adult transcriptome databases using local BLAST and de novo prediction, of which 10 pNPs belonging to the ancient eumetazoa, 23 pNPs belonging to the ancient bilaterian, 3 pNPs belonging to the ancient protostome, 10 pNPs exclusive in lophotrochozoa, 3 pNPs exclusive in annelid, and 5 pNPs only found in U. unicinctus. Furthermore, four pNPs (MIP, FRWamide, FxFamide and FILamide) which may be associated with the settlement and metamorphosis of U. unicinctus larvae were verified successfully by qRT-PCR. Whole-mount in situ hybridization results showed that all the four pNPs were expressed in the region of the apical organ of the larva, and the positive signals were also detected in the ciliary band and abdomen chaetae. We speculated that these pNPs may regulate the movement of larval cilia and chaeta by sensing external attachment signals.Conclusions: This study represents the first comprehensive identification of neuropeptides in Echiura, and would contribute to a complete understanding on the roles of various neuropeptides in larval settlement of most marine benthonic invertebrates.