Cdk2 catalytic activity is essential for meiotic cell division in vivo.
Cdk2 catalytic activity is essential for meiotic cell division in vivo.
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DOI：
10.1042/bcj20160607
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发表时间：
2016-09-15
期刊：
影响因子：
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通讯作者：
Kaldis P
中科院分区：
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作者：
Chauhan S;Diril MK;Lee JH;Bisteau X;Manoharan V;Adhikari D;Ratnacaram CK;Janela B;Noffke J;Ginhoux F;Coppola V;Liu K;Tessarollo L;Kaldis P
细胞周期蛋白依赖性激酶（Cdks）通过磷酸化关键调节蛋白中的丝氨酸和苏氨酸残基来控制真核细胞周期，但是一些Cdk家族成员可能发挥激酶独立的功能，这些功能不能容易地使用基因敲除方法来评估。虽然Cdk 2缺陷型小鼠由于Cdk 1和Cdk 4的补偿活性而显示出接近正常的有丝分裂细胞增殖，但它们不能进行配子的减数分裂产生，因此是不育的。为了研究Cdk 2是否通过蛋白磷酸化或替代激酶独立机制调节减数分裂，我们产生了两种不同的敲入小鼠品系，其中Cdk 2点突变消除酶活性而不改变蛋白表达水平。突变Cdk 2D 145 N/D145 N或Cdk 2 T160 A/T160 A的纯合子小鼠在内源性启动子的控制下仅表达Cdk 2的“激酶死亡”变体，尽管表现出细胞周期调节蛋白和复合物的正常表达，但两种突变均使小鼠不育。只表达Cdk 2的“激酶死亡”变体的小鼠细胞在体外和体内均显示正常的有丝分裂细胞周期进展和增殖，表明Cdk 2激酶活性的丧失对这种细胞分裂模式几乎没有影响。与此相反，Cdk 2突变小鼠的生殖器官表现出异常的形态和功能受损与有缺陷的减数分裂细胞分裂和无法产生配子。因此，Cdk 2突变动物与完全缺乏Cdk 2蛋白的基因敲除小鼠相当。总之，我们的数据表明，Cdk 2的基本减数分裂功能取决于其激酶活性，没有它的产生的单倍体细胞被破坏，导致不育的其他健康的动物。
Cyclin-dependent kinases (Cdks) control the eukaryotic cell cycle by phosphorylating serine and threonine residues in key regulatory proteins, but some Cdk family members may exert kinase-independent functions that cannot easily be assessed using gene knockout approaches. While Cdk2-deficient mice display near-normal mitotic cell proliferation due to the compensatory activities of Cdk1 and Cdk4, they are unable to undergo meiotic generation of gametes and are consequently sterile. To investigate whether Cdk2 regulates meiosis via protein phosphorylation or by alternative kinase-independent mechanisms, we generated two different knockin mouse strains in which Cdk2 point mutations ablated enzyme activity without altering protein expression levels. Mice homozygous for the mutations Cdk2D145N/D145N or Cdk2T160A/T160A expressed only ‘kinase-dead’ variants of Cdk2 under the control of the endogenous promoter, and despite exhibiting normal expression of cell cycle regulatory proteins and complexes, both mutations rendered mice sterile. Mouse cells that expressed only ‘kinase-dead’ variants of Cdk2 displayed normal mitotic cell cycle progression and proliferation both in vitro and in vivo, indicating that loss of Cdk2 kinase activity exerted little effect on this mode of cell division. In contrast, the reproductive organs of Cdk2 mutant mice exhibited abnormal morphology and impaired function associated with defective meiotic cell division and inability to produce gametes. Cdk2 mutant animals were therefore comparable to gene knockout mice, which completely lack the Cdk2 protein. Together, our data indicate that the essential meiotic functions of Cdk2 depend on its kinase activity, without which the generation of haploid cells is disrupted, resulting in sterility of otherwise healthy animals.