人cAMP依赖型蛋白激酶B抑制剂(PKIB)是一种新的耐热蛋白.实验证明，PKIB对PKA催化亚单位具有抑制作用.为研究PKIB在细胞衰老中的功能，以年轻和年老人胚肺二倍体成纤维细胞(2BS细胞株)为实验对象,通过实时PCR发现,在年老细胞中，PKIB表达高于年轻细胞;用PKA活化剂处理年轻和年老细胞,结果显示,在年老细胞中PKA活性变化较年轻细胞小.通过免疫共沉淀实验证实,在年老细胞中，PKIB与PKA催化亚单位结合较年轻细胞中多；进一步通过在年轻细胞中过表达PKIB，检测细胞PKA活性，发现PKA活性明显降低，进一步证实了在人胚肺二倍体成纤维细胞中PKIB对PKA活性的抑制作用；利用Western实验结果证实,PKA催化亚单位在年轻细胞中的表达高于年老细胞.以上结果证明,在2BS细胞中，PKA活性受PKIB的抑制；这种抑制作用与细胞的衰老有一定的关联作用.
Abstract
cAMP-dependent protein kinase β inhibitors (PKIB) are discovered as a new class of heatresistant endogenous proteins in human that inhibit the kinase activity. To investigate the function of PKIB in senescent cells, long-cultured 2BS diploid fibroblast from human fetal lung was used as the experimental subject and compared with the cells with less passages. The PKIB expression was higher in senescent cells as determined by real time-PCR and lead to only limited activation of cAMP-dependent protein kinase after treated with PKA-activator Forskolin. The ability of PKIB binding to PKA catalytic subunit was reduced comparing to young cells as shown by immunoprecipitation. The PKIB overexpression in the young cells significantly reduced the cAMPdependent protein kinase activity, while the amount of PKA catalytic subunit was similar in both old and young cells as confirmed by Western blotting. These result suggested that the inhibition of cAMP-dependent protein kinase might be associated with cell senescence.
关键词
人cAMP依赖型蛋白激酶B抑制剂(PKIB) /
细胞衰老 /
PKA /
人胚肺二倍体成纤维细胞
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Key words
protein kinase inhibitorB (PKIB)； cell senescence； cAMP-dependent protein kinase； human embryonic lung diploid fibroblasts
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中图分类号：
Q225
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