Isolation and characterization of goldfish Y box protein, a germ-cell-specific RNA-binding protein.
Cyclin B is a regulatory subunit of maturation-promoting factor. In goldfish (Carassius auratus) oocytes, cyclin B is synthesized de novo after stimulation by 17alpha,20beta-dihydroxy-4-pregnen-3-one (maturation-inducing hormone). In this study, we examined goldfish oocyte proteins bound to cyclin B mRNA. Using oligo(dT)-cellulose affinity chromatography and northwestern blotting analysis, we identified a 54-kDa cyclin B mRNA-binding protein (p54). Southwestern blotting analysis showed the binding of p54 to the Y box DNA element (CTGATTGGCCAA), suggesting that p54 is a Y box protein in goldfish. We isolated two cDNA clones, GFYP1 and GFYP2, the latter of which encodes a germ-cell-specific Y box protein. An antibody against a GFYP2 protein recognized p54, suggesting that p54 is identical or highly similar to GFYP2 protein. This is also supported by the finding that a recombinant GFYP2 expressed in bacteria bound to both the Y box DNA element and the goldfish cyclin B mRNA synthesized in vitro. These results suggest that p54 is a germ-cell-specific Y box protein and is a potential masking protein of cyclin B mRNA in goldfish oocytes.
Katsu
Y
Y
Laboratory of Reproductive Biology, National Institute for Basic Biology, Okazaki, Japan.
Yamashita
M
M
Nagahama
Y
Y
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D002846:Chromatography, Affinity; D003001:Cloning, Molecular; D019926:Cyclin B / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism; D018076:DNA, Complementary; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D029941:Fish Proteins; D015870:Gene Expression; D006054:Goldfish / Q000378:metabolism*; D008969:Molecular Sequence Data; D009865:Oocytes / Q000737:chemistry*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid
D019926:Cyclin B; D018076:DNA, Complementary; D029941:Fish Proteins; C109595:GFYP2 protein, Carassius auratus; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D011994:Recombinant Proteins