BEHAVIOR OF THE COMPONENTS OF MATURATION-PROMOTING FACTOR, CDC2 KINASE AND CYCLIN-B, DURING OOCYTE MATURATION OF GOLDFISH
BEHAVIOR OF THE COMPONENTS OF MATURATION-PROMOTING FACTOR, CDC2 KINASE AND CYCLIN-B, DURING OOCYTE MATURATION OF GOLDFISH
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DOI：
10.1006/dbio.1993.1289
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发表时间：
1993-11-01
影响因子：
2.7
通讯作者：
NAGAHAMA, Y
中科院分区：
文献类型：
--
作者：
KATSU, Y;YAMASHITA, M;NAGAHAMA, Y
我们利用针对金鱼cdc2激酶c端序列和大肠杆菌产生的全长金鱼细胞周期蛋白B的单克隆抗体，检测了在金鱼卵母细胞成熟过程中，成熟促进因子（MPF）的两种成分cdc2激酶和细胞周期蛋白B的变化。除了35-kDa形式外，在发生生发囊泡破裂（GVBD）的卵母细胞中检测到34-kDa活性cdc2激酶。Cyclin B在未成熟卵母细胞中不存在，仅在GVBD出现之前出现，与34 kda活性cdc2激酶的出现完全一致。用p13suclbeads和抗cyclin B抗体进行沉淀，发现cyclin B一出现就与cdc2激酶形成复合物。将1 ng cyclin B引入未成熟卵母细胞或卵母细胞提取物后诱导MPF激活。这与成熟卵母细胞中发现的细胞周期蛋白B的数量相对应（卵母细胞中的浓度为2μg/ml）。这些结果表明，鱼类卵母细胞的MPF激活是由卵母细胞成熟过程中预先存在的cdc2激酶和新合成的细胞周期蛋白B形成的复合物诱导的，这与xenopusand海星的情况不同，后者的未成熟卵母细胞中已经存在cdc2激酶-细胞周期蛋白B复合物。与非洲爪蟾不同，未受精的成熟金鱼卵母细胞中蛋白质合成的抑制导致cdc2激酶活性/细胞周期蛋白B蛋白水平的降低，并导致从减数分裂中期到减数分裂后期的进展。这一结果表明，金鱼卵母细胞维持MPF活性的机制与非洲爪蟾不同。
We examined the changes that occurred in the two components of maturation-promoting factor (MPF), cdc2 kinase and cyclin B, during oocyte maturation in goldfish, using monoclonal antibodies against the C-terminal sequence of goldfish cdc2 kinase andEscherichia coli-produced full-length goldfish cyclin B. Immature oocytes contained a 35-kDa inactive cdc2 kinase. In addition to the 35-kDa form, a 34-kDa active cdc2 kinase was detected in oocytes undergoing germinal vesicle breakdown (GVBD). Cyclin B was absent in immature oocytes and appeared just before GVBD, coinciding exactly with the appearance of the 34-kDa active cdc2 kinase. Precipitation with p13suclbeads and anti-cyclin B antibody revealed that cyclin B formed a complex with cdc2 kinase as soon as it appeared. MPF activation was induced by 1 ng cyclin B after introduction into immature oocytes or oocyte extracts. This corresponds to the amount of cyclin B found in mature oocytes (the concentration in the oocyte is 2μg/ml). These results suggest that MPF activation in fish oocytes is induced by complex formation with preexisting cdc2 kinase and newly synthesized cyclin B during oocyte maturation, a situation differing from that inXenopusand starfish, in which the cdc2 kinase-cyclin B complex is already present in immature oocytes. Unlike that inXenopus, an inhibition of protein synthesis in unfertilized mature goldfish oocytes caused a decrease in the cdc2 kinase activity/cyclin B protein level and led to a progression from meiotic metaphase to meiotic anaphase. This result indicates that the mechanisms of maintaining MPF activity in mature goldfish oocytes differ from those inXenopus.