Isolation and characterization of <i>Vasa</i> in the frog <i>Rana rugosa</i>

Saotome, Kazuhiro; Hayashi, Kota; Adachi, Noritaka; Nakamura, Yoriko; Nakamura, Masahisa

doi:10.1002/jez.617

Cited by 8 publications

(10 citation statements)

References 34 publications

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“…In C. encaustus , the Vasa protein was specifically detected in the germline cell cytoplasm. A similar distribution pattern of this protein has been reported in mammals (Fujiwara et al , 1994; Castrillon et al , 2000; Toyooka et al , 2000), birds (Tsunekawa et al , 2000) and frogs (Saotome et al , 2010). In O. latipes , Shinomiya et al (2000) detected Vasa transcripts specifically in germline cell cytoplasm and Aoki et al (2008) reported spatial and temporal changes of the Vasa protein during sex differentiation.…”

Section: Discussionsupporting

confidence: 83%

Gonadal morphogenesis and sex differentiation in the viviparous fish Chapalichthys encaustus (Teleostei, Cyprinodontiformes, Goodeidae)

Guerrero-Estévez

Moreno‐Mendoza

2012

Journal of Fish Biology

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This study describes the structural and ultrastructural characteristics of gonadal sex differentiation and expression of Vasa, a germline marker, in different developmental stages of embryos and newborn fry of the barred splitfin Chapalichthys encaustus, a viviparous freshwater teleost endemic to Mexico. In stage 2 embryos, the gonadal crest was established; gonadal primordia were located on the coelomic epithelium, formed by scarce germ and somatic cells. At stage 3, the undifferentiated gonad appeared suspended from the mesentery of the developing swimbladder and contained a larger number of germ and somatic cells. At stages 4 and 5, the gonads had groups of meiotic and non-meiotic germ cells surrounded by somatic cells; meiosis was evident from the presence of synaptonemal complexes. These stages constituted a transition towards differentiation. At stage 6 and at birth, the gonad was morphologically differentiated into an ovary or a testis. Ovarian differentiation was revealed by the presence of follicles containing meiotic oocytes, and testicular differentiation by the development of testicular lobules containing spermatogonia in mitotic arrest, surrounded by Sertoli cells. Nuage, electron-dense material associated with mitochondria, was observed in germ cells at all gonadal stages. The Vasa protein was detected in all of the previously described stages within the germ-cell cytoplasm. This is the first report on morphological characteristics and expression of the Vasa gene during sexual differentiation in viviparous species of the Goodeidae family. Chapalichthys encaustus may serve as a model to study processes of sexual differentiation in viviparous fishes and teleosts.

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Section: Discussionsupporting

confidence: 83%

Gonadal morphogenesis and sex differentiation in the viviparous fish Chapalichthys encaustus (Teleostei, Cyprinodontiformes, Goodeidae)

Guerrero-Estévez

Moreno‐Mendoza

2012

Journal of Fish Biology

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“…Vasa protein, which is expressed in germ cells and retained as a maternal factor, is known to be a specific germ cell marker in a variety of species such as Drosophila (Hay, January, & January, ), zebrafish (Olsen, Aasland, & Fjose, ), Xenopus (Komiya, Itoh, Ikenishi, & Furusawa, ), R. rugosa (Saotome, Hayashi, et al., ) and mice (Fujiwara et al., ). Measurement of Vasa‐positive germ cells during the early development stages of R. rugosa embryos revealed that PGCs reach the genital ridge of the tadpole between St. 24 and St. 25‐1W from the original site.…”

Section: Resultsmentioning

confidence: 99%

“…We prepared an antibody to Vasa, a protein specific to germ cells in a variety of animals that can be used as a marker (Saotome, Hayashi, Adachi, Nakamura, & Nakamura, ). For immunohistology we purchased the anti‐laminin antibody from Sigma‐Aldrich (St. Louis, MO, USA).…”

Section: Methodsmentioning

confidence: 99%

“…For immunohistology we purchased the anti‐laminin antibody from Sigma‐Aldrich (St. Louis, MO, USA). Frozen sections (10 μm) from embryos, and both wild‐type (Wt) and Nanos3 ‐KD R. rugosa tadpoles were stained for laminin and Vasa as previously described (Saotome, Hayashi, et al., ), using the Vasa antibody at a dilution of 1:1,000 followed by the secondary antibody goat anti‐rabbit IgG Alexa Fluor 488 antibody (Life Technologies, Tokyo, Japan), or the laminin antibody at 1:300 followed by VECTASTAIN ABC system (Vector Laboratories, CA, USA) or non‐immune serum. Sections were counterstained with DAPI (4′,6‐diamidino‐2‐phenylindole; Invitrogen, Tokyo, Japan).…”

Section: Methodsmentioning

confidence: 99%

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Nanos3 of the frog Rana rugosa: Molecular cloning and characterization

et al. 2018

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Nanos is expressed in the primordial germ cells (PGCs) and also the germ cells of a variety of organisms as diverse as Drosophila, medaka fish, Xenopus and mouse. In Nanos3-deficient mice, PGCs fail to incorporate into the gonad and the size of the testis and ovary is thereby dramatically reduced. To elucidate the role of Nanos in an amphibian species, we cloned Nanos3 cDNA from the testis of the R. rugosa frog. RT-PCR analysis showed strong expression of Nanos3 mRNA in the testis of adult R. rugosa frogs, but expression was not sexually dimorphic during gonadal differentiation. In Nanos3-knockdown tadpoles produced by the CRISPR/Cas9 system, the number of germ cells decreased dramatically in the gonads of both male and female tadpoles before sex determination and thereafter. This was confirmed by three dimensional imaging of wild-type and Nanos3 knockdown gonads using serial sections immunostained for Vasa, a marker specific to germ cells. Taken together, these results suggest that Nanos3 protein function is conserved between R. rugosa and mouse.

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“…This was followed by Alexa Flour 488 goat antimouse IgG (Life Technologies, A11017, Tokyo, Japan), diluted at 1:1,000. Counterstaining with DAPI or Hematoxylin and Eosin (HE) is described elsewhere (18). Frozen sections were also double-stained with AMH and laminin antibodies (SIGMA, L9393) at a dilution of 1:300.…”

Section: Immunohistologymentioning

confidence: 99%

Molecular Cloning and Characterization of Anti-Müllerian Hormone (AMH) from the Japanese Wrinkled Frog, Rana rugosa

et al. 2015

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The role of anti-Müllerian hormone (AMH) during gonad development has been studied extensively in many species of mammal, bird, reptile, and fish but remains unresolved in amphibians. In male mammalian embryos, Sox9 activates AMH expression, which initiates regression of the Müllerian ducts. However, Sox9 (Sry-related HMG box 9) is unlikely to initiate AMH in chicken, because AMH precedes Sox9 expression in this species. To clarify whether AMH is involved in testicular differentiation in amphibians, we cloned the full-length AMH cDNA from the Japanese wrinkled frog, Rana rugosa. The AMH gene, which appears to be autosomal, is exclusively expressed in the testis of adult frog among 8 different tissues examined; Sertoli cells are probably responsible for its expression. AMH expression was found in the undifferentiated gonad of both male and female tadpoles, increasing in the differentiating testis. Moreover, we observed consensus binding sites for Sox9 in the 5'-flanking region of the AMH gene. Sox9 stimulated statistically significant AMH expression in luciferase reporter assays when coexpressed in Xenopus kidney-derived A6 cells. However, Sox9 expression showed no sexual dimorphism when AMH expression was up-regulated in the developing testis. These results, taken together, suggest that AMH is probably involved in testicular differentiation in R. rugosa, although an additional, perhaps tissue-specific, transcription factor may be required for the regulation of AMH transcription.

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Isolation and characterization of Vasa in the frog Rana rugosa

Cited by 8 publications

References 34 publications

Gonadal morphogenesis and sex differentiation in the viviparous fish Chapalichthys encaustus (Teleostei, Cyprinodontiformes, Goodeidae)

Gonadal morphogenesis and sex differentiation in the viviparous fish Chapalichthys encaustus (Teleostei, Cyprinodontiformes, Goodeidae)

Nanos3 of the frog Rana rugosa: Molecular cloning and characterization

Molecular Cloning and Characterization of Anti-Müllerian Hormone (AMH) from the Japanese Wrinkled Frog, Rana rugosa

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