E1A modulates phosphorylation of p130 and p107 by differentially regulating the activity of G1/S cyclin/CDK complexes.
E1A modulates phosphorylation of p130 and p107 by differentially regulating the activity of G1/S cyclin/CDK complexes.
复制标题
E1A 通过差异调节 G1/S 细胞周期蛋白/CDK 复合物的活性来调节 p130 和 p107 的磷酸化。
DOI：
10.1038/sj.onc.1204644
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发表时间：
2001
期刊：
影响因子：
--
通讯作者：
Grana,X
中科院分区：
文献类型：
--
作者：
Parreno,M;Garriga,J;Limon,A;Albrecht,JH;Grana,X
我们以前已经表明，腺病毒12 S E1 A蛋白调节p130和p107的磷酸化状态，而视网膜母细胞瘤蛋白的细胞周期依赖性磷酸化没有明显的变化。在这里，我们报告的机制，E1 A修改差异口袋蛋白的磷酸化状态。在瞬时表达E1 A的人U-2 OS骨肉瘤细胞中，单独或组合的D型细胞周期蛋白的异位表达，但不包括细胞周期蛋白E和/或A，完全挽救了E1 A介导的p130过度磷酸化形成3的阻滞。然而，细胞周期蛋白E和A，单独或一起，诱导过度磷酸化的p130的物种与中间的流动性。磷酸肽图谱表明E1 A抑制CDK可磷酸化位点的磷酸化。其中一个位点是Ser-1044。在表达E1 A的细胞中，用p16和显性负性CDK 2阻断内源性和外源性细胞周期蛋白的活性的效果进一步表明，p130被D型细胞周期蛋白和细胞周期蛋白E/CDK复合物磷酸化，并且E1 A通过独立的机制调节这些G1/S CDK的活性。E1 A在MC 3 T3-E1细胞中的稳定表达导致D型细胞周期蛋白的下调，以及细胞周期蛋白E和A的上调。这伴随着CDK 2激酶活性的增加。这些细胞中D型细胞周期蛋白的下调与阻断p130过度磷酸化形成3和p107过度磷酸化相关。这被D型细胞周期蛋白拯救，但不被细胞周期蛋白E拯救。此外，我们表明，上调细胞周期蛋白E和A至少部分依赖于一个完整的口袋蛋白/E2 F途径，但下调D型细胞周期蛋白不是。此外，我们提供的证据表明，虽然缺乏一个功能性pRB途径也导致了对p130的过度磷酸化形成3块，这是不足以诱导组成型表达的p130形式2b。
We have previously shown that the adenoviral 12S E1A protein modulates the phosphorylation status of p130 and p107 without apparent changes in the cell cycle dependent phosphorylation of the retinoblastoma protein. Here we report on the mechanisms by which E1A modifies differentially the phosphorylation status of pocket proteins. In human U-2 OS osteosarcoma cells transiently expressing E1A, ectopic expression of D-type cyclins alone or combined, but not cyclins E and/or A, fully rescues E1A-mediated block in hyperphosphorylation of p130 to form 3. However, cyclins E and A, individually or together, induce hyperphosphorylation of p130 to species with intermediate mobility. Phosphopeptide maps indicate that E1A inhibits phosphorylation of sites phosphorylatable by CDKs. One of these sites is Ser-1044. The effects of blocking the activities of endogenous and exogenous cyclins with p16 and dominant negative CDK2 in E1A expressing cells further indicate that p130 is phosphorylated by both D-type cyclin and cyclin E/CDK complexes and that E1A modulates the activity of these G1/S CDKs by independent mechanisms. Stable expression of E1A in MC3T3-E1 cells leads to downregulation of D-type cyclins, and upregulation of cyclins E and A. This is accompanied by increased CDK2 kinase activity. Downregulation of D-type cyclins in these cells correlates with a block on both p130 hyperphosphorylation to form 3 and hyperphosphorylation of p107. This is rescued by D-type cyclins but not by cyclin E. In addition, we show that the upregulation of cyclins E and A is at least partially dependent on an intact pocket protein/E2F pathway, but downregulation of D-type cyclins is not. Moreover, we provide evidence that while the lack of a functional pRB pathway also results in a block on hyperphosphorylation of p130 to form 3, this is not sufficient to induce constitutive expression of p130 form 2b.