Identification and molecular cloning of germinal vesicle lamin B3 in goldfish (Carassius auratus) oocytes

Akihiko Yamaguchi, Masakane Yamashita, Michiyasu Yoshikuni, Yoshitaka Nagahama

Research output: Contribution to journalArticle

11 Citations (Scopus)

Abstract

A bulk isolation method was developed to collect a large number of germinal vesicles (GV) from postvitellogenic oocytes of goldfish (Carassius auratus). Using this method, we obtained GV lamina which are resistant to high salt and nonionic detergent. 2D PAGE revealed that the goldfish GV lamina contained several spots with similar molecular masses (67 kDa) and slightly different neutral isoelectro-focusing values (pI 5.8-6.2). After trypsin digestion and extraction of a major spot (pI 6.1), the peptide was subjected to RP-HPLC and sequenced. A homology search identified this spot as a nuclear lamin. A cDNA encoding goldfish GV lamin was isolated by RT-PCR using degenerate primers designed from the GV lamin tryptic peptide sequence. The goldfish GV lamin cDNA encodes a predicted molecular mass of 67 455 Da with a pI of 5.84. Phylogenetic analysis indicates that the amino-acid sequence is most similar to Xenopus oocyte-specific GV lamin B3, but differs from somatic lamins (A, B1 or B2). In contrast to somatic lamins, neither goldfish nor Xenopus GV lamin contain conserved phosphorylation sites for nuclear transport, except the nuclear localization sequence. Therefore, we conclude that the goldfish oocyte GV is mainly comprised of GV-type lamin (the homolog of Xenopus lamin B3).

Original languageEnglish
Pages (from-to)932-939
Number of pages8
JournalEuropean Journal of Biochemistry
Volume268
Issue number4
DOIs
Publication statusPublished - May 1 2001
Externally publishedYes

Fingerprint

Lamins
Goldfish
Cloning
Molecular Cloning
Oocytes
Xenopus
Molecular mass
Complementary DNA
Lamin Type A
Peptides
Phosphorylation
Cell Nucleus Active Transport
Protein Sequence Analysis
Electrophoresis, Gel, Two-Dimensional
lamin B2
Detergents
Trypsin
Digestion
Salts
High Pressure Liquid Chromatography

All Science Journal Classification (ASJC) codes

  • Biochemistry

Cite this

Identification and molecular cloning of germinal vesicle lamin B3 in goldfish (Carassius auratus) oocytes. / Yamaguchi, Akihiko; Yamashita, Masakane; Yoshikuni, Michiyasu; Nagahama, Yoshitaka.

In: European Journal of Biochemistry, Vol. 268, No. 4, 01.05.2001, p. 932-939.

Research output: Contribution to journalArticle

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