Molecular characterization and developmental expression of vitellogenin in the oriental river prawn Macrobrachium nipponense and the effects of RNA interference and eyestalk ablation on ovarian maturation
Introduction
The oriental river prawn Macrobrachium nipponense (Crustacea; Decapoda; Palaemonidae) is an important commercial prawn species that is widely distributed in freshwater areas of China and other Asian countries. The total fishing production reaches 230,248 t per year in China (Bureau of Fishery, 2011), with an annual production value of more than 100 million RMB. As the scale of production expanded, “sexual precocity” began to appear. This term refers to early male and female gonad development, which leads to excessive propagation and overpopulation; in the M. nipponense population this occurs especially in autumn. Precocity results in the coexistence of multiple generations, intensive breeding density, and lack of oxygen, which can lead to short life span and low market value of the product. Thus, this phenomenon is restricting the sustainable development of M. nipponense. Understanding the reproductive process and the mechanisms that regulate ovarian maturation in M. nipponense is crucial to improving production of this commercially important species.
Vitellogenin (Vg), which is the precursor of vitellin (Vn), is synthesized by female shrimp during gonad maturation. In the mature female prawn, gonad maturity depends on the rapid synthesis and accumulation of Vg in the oocytes during the breeding season (Wilder et al., 2010). In many oviparous vertebrate and invertebrate animals, Vn provides the substrate and energy for embryonic and ovarian development (e.g., carbohydrates, amino acids, lipids, vitamins, phosphorus, sulfur, and trace elements) (Matozzo et al., 2008). The complete cDNA sequence encoding Vg has been cloned for many species of decapod crustacean. In addition, the molecular characteristics and the regulatory mechanism of Vg have been widely studied (Tsutsui et al., 2000, Yang et al., 2000, Raviv et al., 2006, Okumura et al., 2007, Jia et al., 2013). However, molecular studies of M. nipponense Vg (Mn-Vg) are needed to better understand the mechanisms involved in the reproductive process of this species.
In crustaceans, the site and the process of Vg synthesis are still controversial. Mainly have two kinds: extra-ovarian sources, namely by the organ beyond ovary synthesis precursor, Vg was considered to be taken into the developing oocytes from the hemolymph by the vitellogenin receptor (VgR) via receptor-mediated endocytosis. The mechanisms for endocytotic internalization of Vg have been well studied in certain oviparous vertebrates (Schneider, 1992) and insects (Sappington and Raikhel, 1998), but such studies in crustaceans are limited. Vg synthesis also may be endogenous (i.e., auto-synthesis), whereby the oocyte itself produces Vg with participation from relevant organelles. For example, the ovary was found to be the site of Vg synthesis in Penaeus semisulcatus (Browdy et al., 1990) and Callinectes sapidus (Lee and Watson, 1995). However, the Vg gene was uniquely expressed in the hepatopancreas of Macrobrachium rosenbergii (Yang et al., 2000) and Pandalus hypsinotus (Tsutsui et al., 2004). Further studies indicated that both the ovary and hepatopancreas were the Vg synthesis sites in Marsupenaeus japonicus (Okumura et al., 2007), Fenneropenaeus merguiensis (Phiriyangkul et al., 2007), Litopenaeus vannamei (Raviv et al., 2006), Penaeus japonicus (Tsutsui et al., 2000), Metapenaeus ensis and Penaeus monodon (Tiu et al., 2006a, Tiu et al., 2006b).
In decapods, vitellogenesis is hormonally regulated, and it can be inhibited by the occurrence of hormones in the neurosecretory cells of the X-organ/sinus gland. For example, gonad-inhibiting hormone (GIH), which is synthesized in the X-organ/sinus complex, is thought to play an inhibitory role for the initiation of vitellogenesis in the ovary (Kleijn et al., 1994, De Kleijn et al., 1998, Gu et al., 2002). Adiyodi and Adiyodi (1970) reported that eyestalk ablation removed the inhibition of neuropeptides and accelerated the accumulation of Vg. In addition, Jayasankar et al. (2002) found that eyestalk ablation increased Vg synthesis in the hepatopancreas of the giant freshwater prawn M. rosenbergii. In P. japonicus, Vg mRNA transcripts were measured both in the hepatopancreas and the ovary in normal and eyestalk-ablated adult shrimp. An obvious increment of mRNA levels was revealed in the ovary, whereas mRNA levels were negligible in the hepatopancreas (Tsutsui et al., 2005). Overall, existing data indicate that the mechanisms for hormonal regulation of Vg synthesis vary among crustaceans. Thus, molecular characterization and functional studies of Vg are critical to understand the reproductive mechanisms in M. nipponense. Such information can be used to improve aquaculture production in practice.
In this study, we cloned the cDNA encoding the Vg gene from M. nipponense (Mn-Vg) and conducted structural and phylogenetic analyses. The expression profiles of different tissues and development stages (embryo and larvae) were determined using quantitative real-time PCR (qPCR). qPCR was also used to evaluate the effects of eyestalk ablation to gain a better understanding of the hormonal regulation mechanism involved in Vg synthesis. RNAi technology was firstly applied to investigate the expression pattern of Vg in ovary cycles. The results of this study should be helpful for developing methods to cope with the problem of sexual precocity in the aquaculture setting.
Section snippets
Experiment animal
Adult healthy M. nipponense were obtained from Tai lake in Wuxi, China (120°13′44″E, 31°28′22″N). The body weight of the female/male prawns ranged from 1.26 to 4.25 g. Individuals, feed with paludina twice per day, were acclimatized in a recirculating water aquarium system filled with aerated freshwater (25–28 °C) before tissues and embryos were collected. A variety of tissues including: ovary, heart, hepatopancreas, muscle, hemocytes, gill, eyestalk, gut and brain were dissected out from mature
Molecular cloning and structural analysis of the Mn-Vg gene
The full-length Mn-Vg gene is 7804 base pairs (bps) long and includes a 5′-terminal untranslated region (UTR) of 34 bp, a 7611 bp open reading frame (ORF) encoding 2536 amino acid (aa) residues, and a 159 bp 3′-terminal UTR (excluding the poly(A) + tail). The Mn-Vg cDNA sequence was submitted to GenBank under the accession number KJ768657. Fig. 1 shows the sketch map of the deduced amino acid sequences, and the specific sequence information for Mn-Vg is provided in Fig. 1s. Mn-Vg has an estimated
Discussion
In this study, we identified the complete Mn-Vg transcript sequence, which is approximately 8 kb in size with 2536 aas encoded by its ORF. The deduced amino acid sequences revealed the common characteristic sequence of insect Vg (Chen et al., 1997, Sappington and Raikhel, 1998, Sappington et al., 2002), with considerable conservation, particularly in the N-terminus. The vitellogenin N-terminal and VYWD domains are widely found in insects and vertebrates (Baker, 1988). The GLLG motif within the
Acknowledgments
The project was supported by the Freshwater Fisheries Research Center, China Central Governmental Research Institutional Basic Special Research Project from the Public Welfare Fund (2013JBFM15), the National Natural Science Foundation of China (Grant No. 31272654), the National Science & Technology Supporting Program of the 12th Five-year Plan of China (Grant No. 2012BAD26B04), Jiangsu Provincial Natural Science Foundation for Young Scholars of China (Grant No. BK2012091), the Science &
References (59)
- et al.
The vitellogenin cDNA of Cherax quadricarinatus encodes a lipoprotein with calcium binding ability, and its expression is induced following the removal of the androgenic gland in a sexually plastic system
Gen. Comp. Endocrinol.
(2002) - et al.
Characterization of an additional molt inhibiting hormone-like neuropeptide from the shrimp Metapenaeus ensis
Peptides
(2002) Characterization of two vitellogenin cDNAs from a Pandalus shrimp (Pandalopsis japonica): expression in hepatopancreas is down-regulated by endosulfan exposure
Comp. Biochem. Physiol. B Biochem. Mol. Biol.
(2010)- et al.
Characterization and expression profile of Vitellogenin gene from Scylla paramamosain
Gene
(2013) Vitellogenesis in both sexes of gonochoristic mud shrimp, Upogebia major (Crustacea): analyses of vitellogenin gene expression and vitellogenin processing
Comp. Biochem. Physiol. B Biochem. Mol. Biol.
(2008)- et al.
Substrate specificity determinants for casein kinase II as deduced from studies with synthetic peptides
J. Biol. Chem.
(1987) - et al.
Analysis of relative gene expression data using real-time quantitative PCR and the 2 (− Delta Delta C (T)). Method
Methods
(2001) - et al.
Vitellogenin as a biomarker of exposure to estrogenic compounds in aquatic invertebrates: a review
Environ. Int.
(2008) - et al.
Phosphorylation of phosvitin by casein kinase-2 provides the evidence that phosphoserines can replace carboxylic amino acids as specificity determinants
Biochim. Biophys. Acta
(1988) - et al.
Characterization of vitellogenin from rainbow trout (Oncorhynchus mykiss)
Gene
(1996)
Vitellogenin gene expression and hemolymph vitellogenin during vitellogenesis, final maturation, and oviposition in female kuruma prawn, Marsupenaeus japonicus
Comp. Biochem. Physiol. A Mol. Integr. Physiol.
Vitellogenesis in the shrimp, Penaeus vannamei: in vitro studies of the isolated hepatopancreas and ovary
Comp. Biochem. Physiol. B
Complete sequence of Litopenaeus vannamei (Crustacea: Decapoda) vitellogenin cDNA and its expression in endocrinologically induced sub-adult females
Gen. Comp. Endocrinol.
Molecular characteristics of insect vitellogenins and vitellogenin receptors
Insect Biochem. Mol. Biol.
Molecular diversity and evolution of the large lipid transfer protein superfamily
J. Lipid Res.
Equal contribution of hepatopancreas and ovary to the production of vitellogenin (PmVg1) transcripts in the tiger shrimp, Penaeus monodon
Aquaculture
Organization of the shrimp vitellogenin gene: evidence of multiple genes and tissue specific expression by the ovary and hepatopancreas
Gene
Evolution and expression of vitellogenin genes
Trends Genet.
Gene discovery from an ovary cDNA library of oriental river prawn Macrobrachium nipponense by ESTs annotation
J. Comp. Biochem. Physiol. D
Vitellogenin gene of the silkworm, Bombyx mori: structure and sex-dependent expression
FEBS Lett.
Endocrine control of reproduction in decapod Crustacea
Biol. Rev.
Is vitellogenin an ancestor of apolipoprotein B-100 of human lipoprotein and human lipoprotein lipase
Biochemistry
Vitellin synthesis in relation to oogenesis in in vitro-incubated ovaries of Penaeus semisulcatus (Crustaceas, Decapoda, Penaeidae)
Exp. Zool.
Fisheries economic statistics
China Fishery Yearbook
Extensive sequence conservation among insect, nematode, and vertebrate vitellogenins reveals ancient common ancestry
Mol. Evol.
The morphological and histological observation of embryonic development in the oriental river prawn Macrobrachium nipponense
J. Shanghai Ocean Univ.
Expression of the crustacean hyperglycaemic hormones and the Gonad inhibiting hormone during the reproductive cycle of the female American lobster Homarus americanus
J. Endocrinol.
Is there extraovarian synthesis of vitellogenin in penaeid shrimp
Biol. Bull.
Vitellogenin expression in queen ovaries and in larvae of both sexes of Apis mellifera
Arch. Insect Biochem. Physiol.
Cited by (70)
Molecular and functional characterization of ribosome protein S24 in ovarian development of Macrobrachium nipponense
2024, International Journal of Biological MacromoleculesDual roles of CYP302A1 in regulating ovarian maturation and molting in Macrobrachium nipponense
2023, Journal of Steroid Biochemistry and Molecular BiologyInsight into crustacean cathepsins: Structure-evolutionary relationships and functional roles in physiological processes
2023, Fish and Shellfish ImmunologyEffects of four chemosterilants on Bactrocera tau
2022, Ecotoxicology and Environmental SafetyCitation Excerpt :One of the traditional explanations is that the compounds affect the metabolism of nucleic acids, thereby affecting the synthesis of proteins and inhibiting the development of the ovaries or testes (Smith et al., 1964; Kandul et al., 2019). Additionally, Vg affects insect reproduction (Bai et al., 2015; Wu et al., 2021). The broken ring of ovarian development affects the passage of Vg from the hemolymph into the oocyte to form yolk proteins, which leads to the accumulation of hemolymph proteins, especially Vg (Mellaert et al., 1983; Li et al., 2010).