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      Transcriptomic analysis of the differentiating ovary of the protogynous ricefield eel Monopterus albus

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          Abstract

          Background

          The ricefield eel is a protogynous hermaphroditic Synbranchiform species that changes sex naturally from female to male, which offers an interesting model for studying gonadal (particularly ovarian) differentiation in vertebrates. In the present study, transcriptome sequencing of the gonad of ricefield eel larvae was performed to explore the molecular mechanisms underlying the ovarian differentiation and development.

          Results

          A total of 301,267,988 clean reads were generated from cDNA libraries of gonadal tissues of ricefield eel larvae at 6, 9, 12, and 20 days post hatching (dph), which contained undifferentiated gonads, differentiating ovaries, ovaries with oogonia, and ovaries with meiotic oocytes, respectively. De-novo assembly of all the clean reads generated a total of 265,896 unigenes with a mean size of 720 bp and a N50 of 1107 bp. RT-qPCR analysis of the developmental expression of 13 gonadal development-related functional genes indicated that RNA-seq data are reliable. Transcriptome data suggest that high expression of female development-related genes and low expression of male development-related genes in the early gonads of ricefield eel larvae participate in the cascade of sex differentiation leading to the final female phenotype. The contrasting expression patterns of genes involved in retinoid acid (RA) synthesis and degradation might result in peak production of RA at 12 dph in the gonad of ricefield eel larvae, and induce molecular events responsible for the initiation of meiosis before the meiotic signs could be observed at 20 dph. In addition, only stra6 but not stra8 could be identified in gonadal transcriptome data of ricefield eel larvae, and the expression pattern of stra6 paralleled those of genes involved in RA synthesis, suggesting that stra6 may be a downstream target of RA and play a role in RA metabolism and/or meiotic initiation in the gonad of ricefield eel larvae.

          Conclusions

          The present study depicted the first large-scale RNA sequencing of the gonad of ricefield eel larvae, and identified many important functional genes, GO terms and KEGG pathways involved in gonadal development and germ cell meiosis. Results of the present study will facilitate future study on the ovarian differentiation of ricefield eels and other teleosts as well.

          Electronic supplementary material

          The online version of this article (doi:10.1186/s12864-017-3953-6) contains supplementary material, which is available to authorized users.

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          Most cited references75

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          TIGR Gene Indices clustering tools (TGICL): a software system for fast clustering of large EST datasets.

          TGICL is a pipeline for analysis of large Expressed Sequence Tags (EST) and mRNA databases in which the sequences are first clustered based on pairwise sequence similarity, and then assembled by individual clusters (optionally with quality values) to produce longer, more complete consensus sequences. The system can run on multi-CPU architectures including SMP and PVM.
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            DMY is a Y-specific DM-domain gene required for male development in the medaka fish.

            Although the sex-determining gene Sry has been identified in mammals, no comparable genes have been found in non-mammalian vertebrates. Here, we used recombinant breakpoint analysis to restrict the sex-determining region in medaka fish (Oryzias latipes) to a 530-kilobase (kb) stretch of the Y chromosome. Deletion analysis of the Y chromosome of a congenic XY female further shortened the region to 250 kb. Shotgun sequencing of this region predicted 27 genes. Three of these genes were expressed during sexual differentiation. However, only the DM-related PG17 was Y specific; we thus named it DMY. Two naturally occurring mutations establish DMY's critical role in male development. The first heritable mutant--a single insertion in exon 3 and the subsequent truncation of DMY--resulted in all XY female offspring. Similarly, the second XY mutant female showed reduced DMY expression with a high proportion of XY female offspring. During normal development, DMY is expressed only in somatic cells of XY gonads. These findings strongly suggest that the sex-specific DMY is required for testicular development and is a prime candidate for the medaka sex-determining gene.
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              R-spondins function as ligands of the orphan receptors LGR4 and LGR5 to regulate Wnt/beta-catenin signaling.

              The Wnt/β-catenin signaling system plays essential roles in embryonic development and in the self-renewal and maintenance of adult stem cells. R-spondins (RSPOs) are a group of secreted proteins that enhance Wnt/β-catenin signaling and have pleiotropic functions in development and stem cell growth. LGR5, an orphan receptor of the G protein-coupled receptor (GPCR) superfamily, is specifically expressed in stem cells of the intestinal crypt and hair follicle. Knockout of LGR5 in the mouse results in neonatal lethality. LGR4, a receptor closely related to LGR5, also has essential roles in development, as its knockout leads to reduced viability and retarded growth. Overexpression of both receptors has been reported in several types of cancer. Here we demonstrate that LGR4 and LGR5 bind the R-spondins with high affinity and mediate the potentiation of Wnt/β-catenin signaling by enhancing Wnt-induced LRP6 phosphorylation. Interestingly, neither receptor is coupled to heterotrimeric G proteins or to β-arrestin when stimulated by the R-spondins, indicating a unique mechanism of action. The findings provide a basis for stem cell-specific effects of Wnt/β-catenin signaling and for the broad range of functions LGR4, LGR5, and the R-spondins have in normal and malignant growth.
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                Author and article information

                Contributors
                282894295@qq.com
                yangw6@mail2.sysu.edu.cn
                yizezhang007@163.com
                zhihe@sicau.edu.cn
                86-20-84113327 , lsszwm@mail.sysu.edu.cn
                86-20-8411-0828 , zhlih@mail.sysu.edu.cn
                Journal
                BMC Genomics
                BMC Genomics
                BMC Genomics
                BioMed Central (London )
                1471-2164
                3 August 2017
                3 August 2017
                2017
                : 18
                : 573
                Affiliations
                [1 ]ISNI 0000 0001 2360 039X, GRID grid.12981.33, Department of Biology, , School of Life Sciences, Sun Yat-sen University, ; Guangzhou, 510275 People’s Republic of China
                [2 ]ISNI 0000 0001 2360 039X, GRID grid.12981.33, Institute of Aquatic Economic Animals and Guangdong Province Key Laboratory for Aquatic Economic Animals, , Sun Yat-Sen University, ; Guangzhou, 510275 P. R. China
                [3 ]ISNI 0000 0001 0185 3134, GRID grid.80510.3c, , College of Animal Sciences and Technology, Sichuan Agricultural University, ; Ya’an, 625014 P. R. China
                Article
                3953
                10.1186/s12864-017-3953-6
                5541746
                28768496
                4e324e26-053a-4cb3-8c5e-3579a31170fa
                © The Author(s). 2017

                Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License ( http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver ( http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.

                History
                : 26 December 2016
                : 23 July 2017
                Funding
                Funded by: National Natural Science Foundation of China (CN)
                Award ID: 31172088
                Award Recipient :
                Funded by: National Natural Science Foundation of China (CN)
                Award ID: 31572604
                Award Recipient :
                Funded by: National Natural Science Foundation of China (CN)
                Award ID: 31372513
                Award Recipient :
                Categories
                Research Article
                Custom metadata
                © The Author(s) 2017

                Genetics
                monopterus albus,ovary,transcriptome,differentiation,meiosis
                Genetics
                monopterus albus, ovary, transcriptome, differentiation, meiosis

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