Potential role of mitogen-activated protein kinase during meiosis resumption in bovine oocytes
Potential role of mitogen-activated protein kinase during meiosis resumption in bovine oocytes
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DOI：
10.1095/biolreprod55.6.1261
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发表时间：
1996-12-01
影响因子：
3.6
通讯作者：
VandeWoude, GF
中科院分区：
文献类型：
--
作者：
Fissore, RA;He, CL;VandeWoude, GF
在减数分裂成熟过程中，许多细胞质和核事件发生，为卵母细胞受精做准备。这些变化是由几种激酶活性的增加引起的，最明显的是促成熟因子，也称为组蛋白H1激酶。另一种激酶，丝裂原活化蛋白（MAP）激酶，也在此期间受到刺激。在这项研究中，我们研究了MAP激酶在牛卵母细胞成熟中的作用。首先，在成熟过程中的组蛋白H1和MAP激酶的激活动力学进行了评估，同时通过评估其在体外的催化活性。我们发现，它们在大约相同的时间被激活，在生殖囊泡破裂（GVBD）周围。然后，在大约15小时的成熟，H1激酶的活性暂时下降，而MAP激酶保持高通过中期II阶段。第二，MAP激酶的活化和催化活性通过蛋白质印迹和凝胶内激酶测定直接评价。我们确定，MAP激酶被激活，并表现出通过SDS-聚丙烯酰胺凝胶的流动性降低，其催化活性随着成熟的进展而增加。在我们的系统中，大部分MAP激酶活性可归因于p42（MAPK 2）。第三，探索了MAP激酶的激活途径。在非洲爪蟾卵母细胞中，MAP激酶被包括几个上游激活物的激酶级联激活;其中之一是原癌基因mos的产物。在牛卵母细胞中，注射Mos RNA引起MAP激酶的快速和最大激活，导致减数分裂和GVBD的加速恢复。这些结果被认为是由激酶活性Mos产物的过表达介导的，因为注射激酶失活的Mos RNA未能激活MAP激酶。总之，这些结果表明MAP激酶在牛卵母细胞减数分裂的启动和进行过程中的作用。这些数据还表明存在MAP激酶激活级联反应，可以由Mos蛋白启动。
During meiotic maturation, numerous cytoplasmic and nuclear events take place that prepare the oocytes for fertilization. These changes are initiated by an increase in the activity of several kinases, most notably maturation-promoting factor, also called histone H1 kinase. Another kinase, mitogen-activated protein (MAP) kinase, is also stimulated during this period. In this study, we investigated the role of MAP kinase in bovine oocyte maturation. First, the kinetics of activation of histone H1 and MAP kinases during maturation were assessed simultaneously by evaluating their catalytic activities in vitro. We found that they are activated at approximately the same time, around germinal vesicle breakdown (GVBD). Then, at approximately 15 h of maturation, the activity of H1 kinase temporarily decreases, whereas MAP kinase remains high through the metaphase II stage. Second, the activation and catalytic activity of MAP kinase was directly evaluated by Western blotting and by an in-gel kinase assay. We determined that MAP kinase becomes activated and exhibits a decreased mobility through SDS-polyacrylamide gels, and that its catalytic activity increases as maturation progresses. In our system, most of the MAP kinase activity can be attributed to p42(MAPK2). Third, the activation pathway of MAP kinase was explored. In Xenopus oocytes, MAP kinase is activated by a kinase cascade that includes several upstream activators; one of them is the product of the proto-oncogene mos. In bovine oocytes, injection of Mos RNA elicited a rapid and maximal activation of MAP kinase that resulted in accelerated resumption of meiosis and GVBD. These results were thought to be mediated by an overexpression of a kinase-active Mos product because injection of a kinase-inactive Mos RNA failed to activate MAP kinase. Together, these results suggest a role for MAP kinase during the initiation and progression of meiosis in bovine oocytes. The data also suggest the presence of an MAP kinase-activating cascade that can be initiated by the Mos protein.