Molecular cloning and immunological analysis of goldfish cyclin A during oocyte maturation.
Cyclin A belongs to a family of proteins involved in the regulation of the eukaryotic cell cycle. Although cyclin A is thought to be involved in the regulation of both S and M phase, its exact role in the cell cycle, especially in the meiotic cycle (oocyte maturation), is uncertain. We isolated cyclin A cDNA clones from a goldfish oocyte cDNA library. Monoclonal antibody raised against bacterially produced goldfish cyclin A recognized a 47-kDa protein that disappeared after egg activation. Unlike goldfish cyclin B, which is absent in immature oocytes, cyclin A was already present in immature oocytes and its protein level did not change remarkably during oocyte maturation. These results differ from the finding in Xenopus, in which cyclin A is absent, but cyclin B is present, in immature oocytes. Goldfish cyclin A was associated with cdc2 kinase in mature oocytes, but not with cdk2. Recombinant cyclin A bound to and activated cdc2 in a cell-free system, but cyclin A and cdk2 binding was not observed. The kinase activity of cyclin A-cdc2 was undetectable in immature oocytes and first appeared at about the time of germinal vesicle breakdown (GVBD). In contrast to the cyclin B-cdc2 activity that corresponded to the occurrence of GVBD, cyclin A-cdc2 activity increased only slightly until GVBD was completed and increased drastically after the completion of the first meiotic division. Furthermore, microinjection of cyclin A mRNA into immature oocytes did not cause GVBD; however, microinjection of cyclin B mRNA did. These results suggest that cyclin A-cdc2 kinase and cyclin B-cdc2 kinase play different roles in controlling oocyte maturation. The roles of cyclin A in the rapid activation of cyclin B-cdc2 kinase at meiosis I and II transition and in the maintenance of high maturation-promoting factor activity in mature unfertilized eggs are discussed.
Katsu
Y
Y
Department of Molecular Biomechanics, Graduate University for Advanced Studies, Sapporo, Japan.
Yamashita
M
M
Hirai
T
T
Tokumoto
T
T
Kajiura
H
H
Nagahama
Y
Y
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal; D001483:Base Sequence; D016203:CDC2 Protein Kinase / Q000378:metabolism; D003001:Cloning, Molecular; D016213:Cyclins / Q000235:genetics* / Q000276:immunology / Q000378:metabolism; D017931:DNA Primers / Q000235:genetics; D018076:DNA, Complementary / Q000235:genetics; D004789:Enzyme Activation; D006054:Goldfish / Q000235:genetics* / Q000254:growth & development* / Q000378:metabolism; D008969:Molecular Sequence Data; D009865:Oocytes / Q000254:growth & development* / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000235:genetics / Q000494:pharmacology
D000911:Antibodies, Monoclonal; D016213:Cyclins; D017931:DNA Primers; D018076:DNA, Complementary; D012333:RNA, Messenger; D016203:CDC2 Protein Kinase