Y Nagahama

Publications (3)6.75 Total impact

• Article: Molecular characterization and quantification of the gonadotropin receptors FSH-R and LH-R from Atlantic cod (Gadus morhua).

  C Mittelholzer, E Andersson, G L Taranger, D Consten, T Hirai, B Senthilkumaran, Y Nagahama, B Norberg
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  ABSTRACT: In order to elucidate regulatory mechanisms during puberty final oocyte maturation and spawning, full-length sequences coding for the receptors for follicle-stimulating hormone (FSH-R) and luteinizing hormone (LH-R) were isolated from female Atlantic cod (Gadus morhua) by a RACE-PCR based strategy. The nucleotide and amino acid sequences showed high homologies with the corresponding sequences of other fish species but contained some distinct differences. Conserved features important for functionality, such as a long N-terminal extracellular domain (ECD), seven transmembrane domains and a short C-terminal intracellular domain, were identified in both predicted proteins. Partial genomic sequences for these genes were also determined, allowing the design of mRNA-specific quantitative PCR assays. Due to suspected alternative splicing during expression of these genes, additional real-time PCR assays detecting variants containing the membrane-anchoring domain were established. Besides the expected expression of FSH-R and LH-R mRNA in the gonads similarly strong signals for LH-R were also obtained in male gill, and in female and male brain. When relative expression was analysed at different stages of sexual maturation, levels for FSH-R increased moderately during gonadal growth whereas those of LH-R showed a high peak at spawning.
  General and Comparative Endocrinology 11/2008; 160(1):47-58. · 3.27 Impact Factor
• Article: 20 beta-hydroxysteroid dehydrogenase and CYP19A1 are differentially expressed during maturation in Atlantic cod (Gadus morhua).

  C Mittelholzer, E Andersson, D Consten, T Hirai, Y Nagahama, B Norberg
  [show abstract] [hide abstract]
  ABSTRACT: In order to better quantify the molecular mechanisms regulating final oocyte maturation and spawning, complete coding sequences with partially or fully untranslated regions for the steroidogenic enzymes, cytochrome P450 aromatase and 20 beta-hydroxysteroid dehydrogenase, were cloned from ovaries of Atlantic cod (Gadus morhua). The nucleotide and amino acid sequences showed high homologies with the corresponding sequences of other fish species, and conserved features important for functionality were identified in both predicted proteins. The sequences of the corresponding genomic loci were also determined, allowing the design of mRNA-specific quantitative PCR assays. As a reference gene for the real-time RT-PCR assays, eukaryotic elongation factor 1 alpha was chosen, and the mRNA as well as the genomic sequence was determined. In addition, a real-time quantitative PCR assay for the 18S rRNA was adapted to be used in cod. Analysis of immature and maturing female cod from July to January respectively showed that the enzyme genes showed the expected quantitative changes associated with physiological regulation. However, mRNA for eukaryotic elongation factor 1 alpha, and to a lesser extent even 18S rRNA, showed variable expression in these samples as well. To find accurate standards for real-time PCR in such a dynamic organ as the cod ovary is not an easy task, and several possible solutions are discussed.
  Journal of Molecular Endocrinology 11/2007; 39(4):319-28. · 3.48 Impact Factor
• Article: Molecular characterization and quantification of the gonadotropin receptors FSH-R and LH-R from Atlantic cod (Gadus morhua)

  C. Mittelholzer, E. Andersson, G.L. Taranger, D. Consten, T. Hirai, B. Senthilkumaran, Y. Nagahama, B. Norberg
  [show abstract] [hide abstract]
  ABSTRACT: In order to elucidate regulatory mechanisms during puberty final oocyte maturation and spawning, full-length sequences coding for the receptors for follicle-stimulating hormone (FSH-R) and luteinizing hormone (LH-R) were isolated from female Atlantic cod (Gadus morhua) by a RACE-PCR based strategy. The nucleotide and amino acid sequences showed high homologies with the corresponding sequences of other fish species but contained some distinct differences. Conserved features important for functionality, such as a long N-terminal extracellular domain (ECD), seven transmembrane domains and a short C-terminal intracellular domain, were identified in both predicted proteins. Partial genomic sequences for these genes were also determined, allowing the design of mRNA-specific quantitative PCR assays. Due to suspected alternative splicing during expression of these genes, additional real-time PCR assays detecting variants containing the membrane-anchoring domain were established. Besides the expected expression of FSH-R and LH-R mRNA in the gonads similarly strong signals for LH-R were also obtained in male gill, and in female and male brain. When relative expression was analysed at different stages of sexual maturation, levels for FSH-R increased moderately during gonadal growth whereas those of LH-R showed a high peak at spawning.
  General and Comparative Endocrinology.