Disruption of the cyclin D/cyclin-dependent kinase/INK4/retinoblastoma protein regulatory pathway in human neuroblastoma.
Disruption of the cyclin D/cyclin-dependent kinase/INK4/retinoblastoma protein regulatory pathway in human neuroblastoma.
复制标题
DOI：
--
复制
发表时间：
1998-06
期刊：
影响因子：
11.2
通讯作者：
John Easton;T. Wei;J. Lahti;Vincent J. Kidd
中科院分区：
文献类型：
--
作者：
John Easton;T. Wei;J. Lahti;Vincent J. Kidd
p16 INK 4a（MTS 1）和p18 INK 4c基因产物在人神经母细胞瘤细胞系中是正常的并且高度表达。然而，视网膜母细胞瘤蛋白（pRb）在这些细胞中磷酸化并发挥功能。如此高水平的p16 INK 4a/p18 INK 4c通常会抑制含有功能性pRb的细胞中的细胞周期蛋白依赖性激酶（CDK）4和6的活性，从而延迟细胞周期进展和生长。这些神经母细胞瘤细胞系表达CDK 4和CDK 6 mRNA和蛋白，但在本研究中仅检测到显著的CDK 6蛋白激酶活性。此外，在具有显著激酶活性的细胞中，CDK 6不存在于p16 INK 4a免疫复合物中，尽管p16 INK 4a水平很高。其他人已经表明，CDK 4基因产物的NH 2-末端区域中的特定突变可以破坏p16 INK 4a结合，从而绕过其抑制活性。为了确定CDK 6基因突变或其他机制是否是这些细胞系中CDK 6激酶活性的原因，进行了几项补充分析。检查来自每个细胞系的CDK 6基因的可能影响p16 INK 4a结合的突变，而用CDK 6免疫复合物进行p16 INKa加回实验以评估p16 INK 4a功能。一个真正的CDK 6突变，破坏p16 INK 4a的结合，并防止抑制CDK 6蛋白激酶的活性被确定在17个神经母细胞瘤细胞系之一。其余细胞系中p16 INK 4a抑制活性的破坏机制尚不清楚，但这些结果表明，神经母细胞瘤细胞可能以新的方式绕过野生型p16 INK 4a组成型表达所造成的细胞周期阻滞。
The p16INK4a (MTS1) and pl8INK4c gene products are normal, and highly expressed, in human neuroblastoma cell lines. The retinoblastoma protein (pRb) was, nonetheless, phosphorylated and functional in these cells. Such high levels of p16INK4a/p18INK4c should normally inhibit cyclin-dependent kinase (CDK) 4 and 6 activities in cells containing functional pRb, delaying cell cycle progression and growth. These neuroblastoma cell lines express both CDK4 and CDK6 mRNA and protein, but only significant CDK6 protein kinase activity was detected in this study. In addition, CDK6 was not present in p16INK4a immune complexes in cells with significant kinase activity, although p16INK4a levels were high. Others have shown that a specific mutation in the NH2-terminal region of the CDK4 gene product can disrupt p16INK4a binding, thereby bypassing its inhibitory activity. To determine whether mutation of the CDK6 gene, or some other mechanism, is responsible for the CDK6 kinase activity in these cell lines, several complementary analyses were performed. The CDK6 gene from each cell line was examined for mutations that might affect p16INK4a binding, whereas p16INKa add-back experiments were performed with CDK6 immune complexes to assess p16INK4a function. A bona fide CDK6 mutation that disrupts p16INK4a binding and prevents inhibition of CDK6 protein kinase activity was identified in 1 of 17 neuroblastoma cell lines. The mechanism(s) responsible for disruption of p16INK4a inhibitory activity in the remaining cell lines is unknown, but these results suggest that neuroblastoma cells may bypass the cell cycle block imposed by constitutive expression of wild-type p16INK4a in novel ways.