Publications by authors named "Masatoshi Nakamoto"

18 Publications

  • Page 1 of 1

RADSex: A computational workflow to study sex determination using restriction site-associated DNA sequencing data.

Mol Ecol Resour 2021 Jul 9;21(5):1715-1731. Epub 2021 Mar 9.

Physiological Chemistry, Biocenter, University of Wuerzburg, Wuerzburg, Germany.

The study of sex determination and sex chromosome organization in nonmodel species has long been technically challenging, but new sequencing methodologies now enable precise and high-throughput identification of sex-specific genomic sequences. In particular, restriction site-associated DNA sequencing (RAD-Seq) is being extensively applied to explore sex determination systems in many plant and animal species. However, software specifically designed to search for and visualize sex-biased markers using RAD-Seq data is lacking. Here, we present RADSex, a computational analysis workflow designed to study the genetic basis of sex determination using RAD-Seq data. RADSex is simple to use, requires few computational resources, makes no prior assumptions about the type of sex-determination system or structure of the sex locus, and offers convenient visualization through a dedicated R package. To demonstrate the functionality of RADSex, we re-analysed a published data set of Japanese medaka, Oryzias latipes, where we uncovered a previously unknown Y chromosome polymorphism. We then used RADSex to analyse new RAD-Seq data sets from 15 fish species spanning multiple taxonomic orders. We identified the sex determination system and sex-specific markers in six of these species, five of which had no known sex-markers prior to this study. We show that RADSex greatly facilitates the study of sex determination systems in nonmodel species thanks to its speed of analyses, low resource usage, ease of application and visualization options. Furthermore, our analysis of new data sets from 15 species provides new insights on sex determination in fish.
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http://dx.doi.org/10.1111/1755-0998.13360DOI Listing
July 2021

Rhizotaxis Modulation in Arabidopsis Is Induced by Diffusible Compounds Produced during the Cocultivation of Arabidopsis and the Endophytic Fungus Serendipita indica.

Plant Cell Physiol 2020 Apr;61(4):838-850

Graduate School of Life and Environmental Sciences, Osaka Prefecture University, 1-1 Gakuen-cho, Sakai, 599-8531 Japan.

Rhizotaxis is established under changing environmental conditions via periodic priming of lateral root (LR) initiation at the root tips and adaptive LR formation along the primary root (PR). In contrast to the adaptable LR formation in response to nutrient availability, there is little information on root development during interactions with beneficial microbes. The Arabidopsis root system is characteristically modified upon colonization by the root endophytic fungus Serendipita indica, accompanied by a marked stimulation of LR formation and the inhibition of PR growth. This root system modification has been attributed to endophyte-derived indole-3-acetic acid (IAA). However, it has yet to be clearly explained how fungal IAA affects the intrinsic LR formation process. In this study, we show that diffusible compounds (chemical signals) other than IAA are present in the coculture medium of Arabidopsis and S. indica and induce auxin-responsive DR5::GUS expression in specific sections within the pericycle layer. The DR5::GUS expression was independent of polar auxin transport and the major IAA biosynthetic pathways, implicating unidentified mechanisms responsible for the auxin response and LR formation. Detailed metabolite analysis revealed the presence of multiple compounds that induce local auxin responses and LR formation. We found that benzoic acid (BA) cooperatively acted with exogenous IAA to generate a local auxin response in the pericycle layer, suggesting that BA is one of the chemical signals involved in adaptable LR formation. Identification and characterization of the chemical signals will contribute to a greater understanding of the molecular mechanisms underlying adaptable root development and to unconventional technologies for sustainable agriculture.
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http://dx.doi.org/10.1093/pcp/pcaa008DOI Listing
April 2020

A SNP in a Steroidogenic Enzyme Is Associated with Phenotypic Sex in Seriola Fishes.

Curr Biol 2019 06 23;29(11):1901-1909.e8. Epub 2019 May 23.

Department of Marine Biosciences, Tokyo University of Marine Science and Technology, 4-5-7 Konan, Minato, Tokyo 108-8477, Japan. Electronic address:

Vertebrate sex development consists largely of two processes: "sex determination," the initial bifurcation of sexual identity, and "sex differentiation," which subsequently facilitates maleness or femaleness according to the sex determination signal. Steroid hormones promote multiple types of sexual dimorphism in eutherian mammals and avians [1-3], in which they are indispensable for proper sex differentiation. By contrast, in many poikilothermic vertebrates, steroid hormones have been proposed to be key players in sex determination as well as sex differentiation [4-8]. This hypothesis was introduced more than 50 years ago but has never been rigorously tested due to difficulties in discriminating the roles of steroids in sex determination and differentiation. We found that a missense SNP in the gene encoding the steroidogenic enzyme 17β-hydroxysteroid dehydrogenase 1 (Hsd17b1) was perfectly associated with ZZ/ZW sex determination in Seriola fishes. Biochemical analyses revealed that a glutamate residue present specifically in Z-type HSD17B1 attenuated interconversion between 17-keto and 17β-hydroxy steroids relative to the allelic product from the W chromosome, which harbors glycine at that position, by disrupting the hydrogen bond network between the steroid and the enzyme's catalytic residues. Hsd17b1 mRNA is constitutively expressed in undifferentiated and differentiating gonads of both genotypic sexes, whereas W-type mRNA is expressed only in genotypic females. Meanwhile, Cyp19a1 is predominantly expressed in differentiating ovary. We conclude that the combination of Hsd17b1 alleles determines sex by modulating endogenous estrogen levels in Seriola species. These findings strongly support the long-standing hypothesis on steroids in sex determination.
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http://dx.doi.org/10.1016/j.cub.2019.04.069DOI Listing
June 2019

Ovarian aromatase loss-of-function mutant medaka undergo ovary degeneration and partial female-to-male sex reversal after puberty.

Mol Cell Endocrinol 2018 01 13;460:104-122. Epub 2017 Jul 13.

Institution for Collaborative Relations, Ehime University, Matsuyama, Ehime 790-8577, Japan. Electronic address:

Although estrogens have been generally considered to play a critical role in ovarian differentiation in non-mammalian vertebrates, the specific functions of estrogens during ovarian differentiation remain unclear. We isolated two mutants with premature stops in the ovarian aromatase (cyp19a1) gene from an N-ethyl-N-nitrosourea-based gene-driven mutagenesis library of the medaka, Oryzias latipes. In XX mutants, gonads first differentiated into normal ovaries containing many ovarian follicles that failed to accumulate yolk. Subsequently, ovarian tissues underwent extensive degeneration, followed by the appearance of testicular tissues on the dorsal side of ovaries. In the newly formed testicular tissue, strong expression of gsdf was detected in sox9a2-positive somatic cells surrounding germline stem cells suggesting that gsdf plays an important role in testicular differentiation during estrogen-depleted female-to-male sex reversal. We conclude that endogenous estrogens synthesized after fertilization are not essential for early ovarian differentiation but are critical for the maintenance of adult ovaries.
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http://dx.doi.org/10.1016/j.mce.2017.07.013DOI Listing
January 2018

A novel C-type lectin gene is a strong candidate gene for Benedenia disease resistance in Japanese yellowtail, Seriola quinqueradiata.

Dev Comp Immunol 2017 11 10;76:361-369. Epub 2017 Jul 10.

Department of Aquatic Marine Biosciences, Tokyo University of Marine Science and Technology, Tokyo 108-8477, Japan. Electronic address:

Little is known about mechanisms of resistance to parasitic diseases in marine finfish. Benedenia disease is caused by infection by the monogenean parasite Benedenia seriolae. Previous quantitative trait locus (QTL) analyses have identified a major QTL associated with resistance to Benedenia disease in linkage group Squ2 of the Japanese yellowtail/amberjack Seriola quinqueradiata. To uncover the bioregulatory mechanism of Benedenia disease resistance, complete Illumina sequencing of BAC clones carrying genomic DNA for the QTL region in linkage group Squ2 was performed to reveal a novel C-type lectin in this region. Expression of the mRNA of this C-type lectin was detected in skin tissue parasitized by B. seriolae. Scanning for single nucleotide polymorphisms (SNPs) uncovered a SNP in the C-type lectin/C-type lectin-like domain that was significantly associated with B. seriolae infection levels. These results strongly suggest that the novel C-type lectin gene controls resistance to Benedenia disease in Japanese yellowtails.
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http://dx.doi.org/10.1016/j.dci.2017.07.010DOI Listing
November 2017

Evolution of Shh endoderm enhancers during morphological transition from ventral lungs to dorsal gas bladder.

Nat Commun 2017 02 3;8:14300. Epub 2017 Feb 3.

Mammalian Genetics Laboratory, Genetic Strains Research Center, National Institute of Genetics, Mishima, Shizuoka 411-8540, Japan.

Shh signalling plays a crucial role for endoderm development. A Shh endoderm enhancer, MACS1, is well conserved across terrestrial animals with lungs. Here, we first show that eliminating mouse MACS1 causes severe defects in laryngeal development, indicating that MACS1-directed Shh signalling is indispensable for respiratory organogenesis. Extensive phylogenetic analyses revealed that MACS1 emerged prior to the divergence of cartilaginous and bony fishes, and even euteleost fishes have a MACS1 orthologue. Meanwhile, ray-finned fishes evolved a novel conserved non-coding sequence in the neighbouring region. Transgenic assays showed that MACS1 drives reporter expression ventrally in laryngeal epithelium. This activity has been lost in the euteleost lineage, and instead, the conserved non-coding sequence of euteleosts acquired an enhancer activity to elicit dorsal epithelial expression in the posterior pharynx and oesophagus. These results implicate that evolution of these two enhancers is relevant to the morphological transition from ventral lungs to dorsal gas bladder.
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http://dx.doi.org/10.1038/ncomms14300DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5296767PMC
February 2017

Critical Involvement of Environmental Carbon Dioxide Fixation to Drive Wax Ester Fermentation in Euglena.

PLoS One 2016;11(9):e0162827. Epub 2016 Sep 26.

Graduate School of Life and Environmental Sciences, Osaka Prefecture University, Sakai, Osaka, Japan.

Accumulation profiles of wax esters in Euglena gracilis Z were studied under several environmental conditions. The highest amount of total wax esters accumulated under hypoxia in the dark, and C28 (myristyl-myristate, C14:0-C14:0) was prevalent among all conditions investigated. The wax ester production was almost completely suppressed under anoxia in the light, and supplying exogenous inorganic carbon sources restored wax ester fermentation, indicating the need for external carbon sources for the wax ester fermentation. 13C-labeling experiments revealed specific isotopic enrichment in the odd-numbered fatty acids derived from wax esters, indicating that the exogenously-supplied CO2 was incorporated into wax esters via the propionyl-CoA pathway through the reverse tricarboxylic acid (TCA) cycle. The addition of 3-mercaptopicolinic acid, a phosphoenolpyruvate carboxykinase (PEPCK) inhibitor, significantly affected the incorporation of 13C into citrate and malate as the biosynthetic intermediates of the odd-numbered fatty acids, suggesting the involvement of PEPCK reaction to drive wax ester fermentation. Additionally, the 13C-enrichment pattern of succinate suggested that the CO2 assimilation might proceed through alternative pathways in addition to the PEPCK reaction. The current results indicate that the mechanisms of anoxic CO2 assimilation are an important target to reinforce wax ester fermentation in Euglena.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0162827PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5036851PMC
September 2016

Diversification of sterol methyltransferase enzymes in plants and a role for β-sitosterol in oriented cell plate formation and polarized growth.

Plant J 2015 Dec 3;84(5):860-74. Epub 2015 Nov 3.

Graduate School of Life and Environmental Sciences, Osaka Prefecture University, 599-8531, Sakai, Japan.

Phytosterols are classified into C24-ethylsterols and C24-methylsterols according to the different C24-alkylation levels conferred by two types of sterol methyltransferases (SMTs). The first type of SMT (SMT1) is widely conserved, whereas the second type (SMT2) has diverged in charophytes and land plants. The Arabidopsis smt2 smt3 mutant is defective in the SMT2 step, leading to deficiency in C24-ethylsterols while the C24-methylsterol pathway is unchanged. smt2 smt3 plants exhibit severe dwarfism and abnormal development throughout their life cycle, with irregular cell division followed by collapsed cell files. Preprophase bands are occasionally formed in perpendicular directions in adjacent cells, and abnormal phragmoplasts with mislocalized KNOLLE syntaxin and tubulin are observed. Defects in auxin-dependent processes are exemplified by mislocalizations of the PIN2 auxin efflux carrier due to disrupted cell division and failure to distribute PIN2 asymmetrically after cytokinesis. Although endocytosis of PIN2-GFP from the plasma membrane (PM) is apparently unaffected in smt2 smt3, strong inhibition of the endocytic recycling is associated with a remarkable reduction in the level of PIN2-GFP on the PM. Aberrant localization of the cytoplasmic linker associated protein (CLASP) and microtubules is implicated in the disrupted endocytic recycling in smt2 smt3. Exogenous C24-ethylsterols partially recover lateral root development and auxin distribution in smt2 smt3 roots. These results indicate that C24-ethylsterols play a crucial role in division plane determination, directional auxin transport, and polar growth. It is proposed that the divergence of SMT2 genes together with the ability to produce C24-ethylsterols were critical events to achieve polarized growth in the plant lineage.
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http://dx.doi.org/10.1111/tpj.13043DOI Listing
December 2015

Expression of 3β-hydroxysteroid dehydrogenase (hsd3b), star and ad4bp/sf-1 during gonadal development in medaka (Oryzias latipes).

Gen Comp Endocrinol 2012 Apr 6;176(2):222-30. Epub 2012 Feb 6.

Department of Biology, Faculty of Science, Shinshu University, Matsumoto, Nagano 390-8621, Japan.

In most vertebrates, sex steroids play a critical role in gonadal development, maturation of germ cells, and development of secondary sexual characteristics. Sex steroids are synthesized in steroid-producing cells (SPCs) in the testis known as Leydig cells, as well as in thecal and granulosa cells in the ovary. In SPCs, cholesterol is sequentially catalyzed by a set of steroidogenic factors and enzymes in order to produce sex steroids. Therefore, integrated expression of the genes involved in steroidogenesis is critical for the proper production of sex steroids. In the present study, regulatory mechanisms of steroidogenic factors and enzymes were examined. We focused on hsd3b, star and ad4bp/sf-1 as well as the description of temporal and spatial expression of these genes during gonadal development in medaka (Oryzias latipes). During testicular development, hsd3b, star and ad4bp/sf-1 were co-expressed in the interstitial somatic cells subsequent to the formation of the seminiferous tubule precursor, suggesting that ad4bp/sf-1 regulated the transcription of both hsd3b and star. During ovarian development, the expression pattern of hsd3b coincided with that of cyp11a1, but not with that of aromatase. Although ad4bp/sf-1 was mainly expressed in presumptive follicular cells, it was also detected in hsd3b positive interstitial cells in the developing ovary. Contrary to our expectations, the onset of star expression occurred during a later stage of ovarian development than the expression of other steroidogenic enzymes. Thus, the regulation mechanism of star transcription appears to differ from that of the other steroidogenic enzymes in the developing ovary, but not in the developing testis.
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http://dx.doi.org/10.1016/j.ygcen.2012.01.019DOI Listing
April 2012

Expression of gonadal soma derived factor (GSDF) is spatially and temporally correlated with early testicular differentiation in medaka.

Gene Expr Patterns 2010 Sep 17;10(6):283-9. Epub 2010 Jun 17.

Laboratory of Reproductive Biology, National Institute for Basic Biology, Okazaki 444-8585, Japan.

In the teleost fish, medaka (Oryzias latipes), the sex is genetically determined at the time of fertilization. The males are heterogametic with XY chromosome composition, while females are of XX chromosome composition. The male sexual differentiation is initiated in XY embryos of medaka by the sex-determining gene Dmy. In this study, we have cloned the gonadal soma derived factor (Gsdf) from medaka and characterized its expression pattern during the initiation of morphological testicular differentiation. By real-time PCR, an XY-specific up-regulation was detected in the expression levels of Gsdf in the whole embryos of medaka at 6days post fertilization (dpf), coincident with the initiation of testicular differentiation in the XY gonads. Whole mount and section in situ hybridizations reaffirmed that Gsdf was expressed exclusively in primordial gonads of only the genetic males at 6dpf. Conversely, the expression of Gsdf was found to be very weak in the XX gonads during embryogenesis. Importantly, Gsdf and Dmy were found to be co-localized in the same somatic cells in the XY gonads. When the XY embryos were treated with estradiol-17beta, in order to reverse their phenotypic sex, a decline was observed in the expression of Gsdf in these embryos by real-time PCR.
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http://dx.doi.org/10.1016/j.gep.2010.06.005DOI Listing
September 2010

Cloning and expression of medaka cholesterol side chain cleavage cytochrome P450 during gonadal development.

Dev Growth Differ 2010 May;52(4):385-95

Department of Biology, Faculty of Science, Shinshu University, Matsumoto, Nagano, Japan.

Cholesterol side chain cleavage cytochrome P450 (P450scc, Cyp11a) is responsible for the first step in steroidogenesis, catalyzing the conversion of cholesterol to prognenolone. To investigate the differentiation of steroid-producing cells and the function of sex steroids during gonadal differentiation in the teleost fish, medaka (Oryzias latipes), we isolated the full length cDNA of medaka P450scc and analyzed the expression pattern of P450scc mRNA during gonadal development using in situ hybridization. At hatching, and just after the initiation of morphological sex differentiation, we did not detect any P450scc expression in both sexes. In male gonads, expression of P450scc was detected in the interstitial somatic cells 15 days after hatching following the formation of the seminiferous tubule precursor, and was maintained in the interstitial somatic cells throughout testicular development. In the female gonad, expression of P450scc was initially detected in interstitial somatic cells 5 days after hatching. Subsequently, the expression of P450scc was continuously detected in the interstitial somatic cells of the developing ovary. This expression pattern of P450scc differed from that of female specific steroidogenic enzyme P450arom. Both P450scc and P450arom expressing cells, only P450scc expressing cells, and only P450arom expressing cells were observed. Our results suggest that expression of steroidogenic enzymes is regulated by various mechanisms during ovarian development.
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http://dx.doi.org/10.1111/j.1440-169X.2010.01178.xDOI Listing
May 2010

Gonadal sex differentiation and expression of Sox9a2, Dmrt1, and Foxl2 in Oryzias luzonensis.

Genesis 2009 May;47(5):289-99

Department of Biology, Shinshu University, Matsumoto, Nagano, Japan.

Oryzias luzonensis is closely related to the medaka, O. latipes. The sex of both species is determined by an XX-XY system. However, the testis determining gene (DMY/Dmrt1bY) found in O. latipes does not exist in O. luzonensis. Instead, a different gene is thought to act as a testis determining gene. In this study, we focused the gonadal sex differentiation process in O. luzonensis under different testis determining gene. First, we observed the gonadal development of O. luzonensis histologically. We then analyzed the expression of Sox9a2/Sox9b, Dmrt1, and Foxl2 during early development. Our results suggest that the sexual differentiation of germ cells in O. luzonensis is initiated later than in O. latipes. However, the timing of the sexual differentiation of the supporting cell linage is similar between the species.
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http://dx.doi.org/10.1002/dvg.20498DOI Listing
May 2009

Preferential localization of neural cell recognition molecule NB-2 in developing glutamatergic neurons in the rat auditory brainstem.

J Comp Neurol 2009 Apr;513(4):349-62

Department of Bioengineering, Nagaoka University of Technology, Nagaoka, Niigata 940-2188, Japan.

NB-2 is a neuronal cell recognition molecule that is preferentially expressed in auditory pathways. Mice deficient in the NB-2 gene exhibit aberrant responses to acoustic stimuli. Here we examined the expression and localization of NB-2 in the auditory brainstem during development in the rat. NB-2 was strongly expressed in the ventral cochlear nucleus (VCN), ventral acoustic stria, lateral and medial superior olivary complex (SOC), superior paraolivary nucleus, medial nucleus of the trapezoid body (MNTB), ventrolateral lemniscus, and central nucleus of the inferior colliculus (CIC). In the VCN and CIC, NB-2 was expressed in the regions that are known to respond to high frequencies. In situ hybridization combined with immunohistochemistry suggested that NB-2 is expressed only in neurons. NB-2 was colocalized with glutamatergic elements in the neuropil and the calyces of Held but not with glycinergic or GABAergic elements. NB-2 expression in the SOC was first detected at embryonic day (E)19, reached a maximum level at postnatal day (P)7, and declined thereafter. Immunolabeling with VGLUT1 and VGLUT2, markers for mature and premature glutamatergic synapses, respectively, in combination with NB-2 immunolabeling revealed that NB-2 is expressed at glutamatergic synapses. Collectively, our findings suggest that NB-2 plays a key role in maturation of glutamatergic synapses in the brainstem during the final stages of auditory development.
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http://dx.doi.org/10.1002/cne.21972DOI Listing
April 2009

Dax1 suppresses P450arom expression in medaka ovarian follicles.

Mol Reprod Dev 2007 Oct;74(10):1239-46

Department of Environmental System Science, Graduate School of Science and Technology, Shinshu University, Matsumoto, Japan.

Dax1 is a member of an unusual orphan nuclear receptor family, and is known to regulate P450arom in mammals and is involved in sex differentiation in some vertebrates. To investigate whether Dax1 is involved in the regulation of the steroidogenic pathway for estrogen biosynthesis in medaka ovarian follicles, we isolated Dax1 cDNA from adult medaka ovaries and analyzed its expression pattern in medaka gonads. In adult ovaries, Dax1 mRNA was detected only in postvitellogenic follicles and was not detected in previtellogenic and vitellogenic follicles. In adult testis, Dax1 mRNA was not detected. We compared the expression pattern of Dax1 with that of Foxl2, Ad4BP/Sf-1, P450c17, and P450arom by in situ hybridization using adjacent sections. In contrast to Dax1 expression, these genes were co-expressed in vitellogenic follicles but were not detected in postvitellogenic follicles. Thus, in medaka ovarian follicles, Dax1 did not show any overlapping expression patterns against Foxl2, Ad4BP/Sf-1, P450c17, and P450arom. Moreover, co-transfection experiments demonstrated that Dax1 inhibits Ad4BP/Sf-1- and Foxl2-mediated P450arom expression. On the other hand, during early sex differentiation, Dax1 mRNA was not detected in both males and females. Our results suggest that Dax1 down-regulates Ad4BP/Sf-1- and Foxl2-mediated P450arom expression in medaka ovarian follicles.
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http://dx.doi.org/10.1002/mrd.20689DOI Listing
October 2007

Molecular cloning and analysis of gonadal expression of Foxl2 in the medaka, Oryzias latipes.

Biochem Biophys Res Commun 2006 May;344(1):353-61

Department of Environmental System Science, Graduate School of Science and Technology, Shinshu University, Matsumoto 390-8621, Japan.

Foxl2 is a member of the winged helix/forkhead family of transcription factors and is known to be involved in ovarian development in some vertebrates. To address the role of Foxl2 in ovarian differentiation in medaka, we isolated Foxl2 cDNA and analyzed its expression patterns during sex differentiation. Expression of Foxl2 started in somatic cells surrounding germ cells in XX gonads, just after initiation of ovarian differentiation, and was maintained in granulosa cells throughout ovarian development. In the adult ovary, Foxl2 was expressed in previtellogenic and vitellogenic follicles, but expression ceased in postvitellogenic follicles. In contrast, Foxl2 mRNA could not be detected in testes. In addition, Foxl2 and aromatase mRNAs were co-localized in some somatic cells located on the ventral side of developing XX gonads. Our results suggested that Foxl2 was not involved in ovarian determination, but was involved in differentiation of granulosa cells in medaka.
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http://dx.doi.org/10.1016/j.bbrc.2006.03.137DOI Listing
May 2006

Effects of estradiol-17beta on germ cell proliferation and DMY expression during early sexual differentiation of the medaka Oryzias latipes.

Zoolog Sci 2005 Jul;22(7):791-6

Department of Environmental System Science, Graduate School of Science and Technology, Shinshu University, Matsumoto, Japan.

Sex reversal of XY male to functional females was induced by estrogen treatment during the embryonic period in the medaka Oryzias latipes. The present study aimed to examine whether exogenous estrogen (estradiol-17beta; E(2)) affects early sex differentiation, paying particular attention to DMY expression and proliferation activity of germ cells in estrogen treated XY individuals. Our results showed that germ cell number was not affected by E(2) treatment at hatching, and that DMY expression was not suppressed under conditions of sex reversal. Therefore, male differentiation of germ cells, which is triggered by the expression of DMY in the supporting cell lineage, proceeds even in E(2) treated XY individuals until hatching, and early sex differentiation is not altered by estrogen. However, sex reversal occurred after hatching probably because of estrogen remaining in the yolk. Interestingly, DMY expression was also detected in the large follicle layer of E(2 )treated XY ovary. These results suggested that DMY regulates male determination in early embryonic stage but does not suppress female follicle development.
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http://dx.doi.org/10.2108/zsj.22.791DOI Listing
July 2005

Testicular type Sox9 is not involved in sex determination but might be in the development of testicular structures in the medaka, Oryzias latipes.

Biochem Biophys Res Commun 2005 Aug;333(3):729-36

Department of Environmental System Science, Graduate School of Science and Technology, Shinshu University, Matsumoto 390-8621, Japan.

Testicular type Sox9 is the most upstream conserved gene in the sex determining cascade among vertebrate. However, in medaka, only one Sox9 gene was identified as expressed in the ovary; no other Sox9 gene was reported expressed in the testis. We explored the medaka genome and cloned a novel testicular type Sox9 cDNA. Phylogenetic analysis revealed that both our isolated Sox9 and the already reportedly cloned medaka Sox9 belongs zebrafish Sox9a branch. Therefore, we named our gene Sox9a2. Unexpectedly, Sox9a2 mRNA was expressed in somatic cells surrounding germ cells at similar high levels in both sexes during early gonadal sex differentiation. However, at the initial stage of testicular tubules development, the expression of Sox9a2 was maintained only in XY gonads, and was remarkably reduced in XX gonads. These results suggest that Sox9a2 is not involved in early sex determination and differentiation, but is involved in the later development of testicular tubules in medaka.
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http://dx.doi.org/10.1016/j.bbrc.2005.05.158DOI Listing
August 2005

Two DM domain genes, DMY and DMRT1, involved in testicular differentiation and development in the medaka, Oryzias latipes.

Dev Dyn 2004 Nov;231(3):518-26

Laboratory of Reproductive Biology, Department of Developmental Biology, National Institute for Basic Biology, Okazaki 444-8585, Japan.

The recent discovery of the DMY gene (DM domain gene on Y chromosome and one of the DMRT1 family genes) as a key determinant of male development in the medaka (Oryzias latipes) has led to its designation as the prime candidate gene for sex-determination in this species. This study focused on the sites and pattern of expression of DMY and DMRT1 genes during gonadal differentiation of medaka to further determine their roles in testis development. DMY mRNA and protein are expressed specifically in the somatic cells surrounding primordial germ cells (PGCs) in the early gonadal primordium, before morphological sex differences are seen. However, somatic cells surrounding PGCs never express DMY during the early migratory period. Expression of DMY persists in Sertoli cell lineage cells, from PGC-supporting cells to Sertoli cells, indicating that only DMY-positive cells enclose PGCs during mitotic arrest after hatching. DMRT1 is expressed in spermatogonium-supporting cells after testicular differentiation (20-30 days after hatching), and its expression is much higher than that of DMY in mature testes. In XX sex-reversed testes, DMRT1 is expressed in the Sertoli cell lineage, similar to the expression of DMY in XY testes. These results suggest strongly that DMY regulates PGC proliferation and differentiation sex-specifically during early gonadal differentiation of XY individuals and that DMRT1 regulates spermatogonial differentiation.
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http://dx.doi.org/10.1002/dvdy.20158DOI Listing
November 2004