Molecular mechanism underlying cAMP-induced amplification of IGF mitogenic activity through PI3-kinase binding protein, PI3KAP/XB130.

cAMP 通过 PI3 激酶结合蛋白 PI3KAP/XB130 诱导 IGF 有丝分裂活性放大的分子机制。

基本信息

批准号：
21580345
负责人：
HAKUNO Fumihiko
金额：
$ 3.16万
依托单位：
The University of Tokyo
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2009
资助国家：
日本
起止时间：
2009 至 2011
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-21580345/
关键词：
IGF-I PI 3-kinase PI3KAP 細胞増殖 PITKAP PI3-kinase

项目摘要

We previously demonstrated that long-term pretreatment of rat FRTL-5 thyroid cells with TSH or cAMP-generating reagents potentiated IGF-I-dependent DNA synthesis. Under this condition, cAMP treatment increased tyrosine phosphorylation of a 125 kDa protein(p125) and its association with a p85 regulatory subunit of phosphatidylinositol 3-kinase(p85 PI3K), which were suggested to be important for potentiation of DNA synthesis. This study was undertaken to identify p125 and to elucidate its roles in potentiation of DNA synthesis induced by IGF-I. Immunoprecipitated phosphotyrosyl p125 in cAMP-stimulated FRTL-5 cells, was shown by MALDI-TOF MS analysis, to be a rat orthologue of human XB130, which we named phosphatidylinositol 3-kinase-associated protein(PI3KAP). cAMP treatment elevated PI3KAP/XB130 mRNA and protein levels as well as tyrosine phosphorylation and PI3KAP/XB130 interaction with p85 PI3K, leading to increased PI3K activities. Importantly, PI3KAP/XB130 knockdown in FRTL-5 cells attenuated cAMP-dependent potentiation of IGF-I-induced DNA synthesis. Addition of Src family kinase inhibitors, PP1 or PP2, during cAMP treatment abolished tyrosine phosphorylation of PI3KAP/XB130 and its interaction with p85 PI3K. In addition, c-Src was associated with PI3KAP/XB130 and was activated in response to cAMP. Together, these data indicate that cAMP-dependent induction of PI3KAP/XB130 and its association with PI3K are required for enhancement of IGF mitogenic activities.

我们先前证明，用TSH或营地生成试剂对大鼠FRTL-5甲状腺细胞进行长期预处理增强了IGF-I依赖性DNA合成。在这种情况下，cAMP处理增加了125 kDa蛋白（P125）的酪氨酸磷酸化及其与磷脂酰肌醇3-激酶（P85 PI3K）的P85调节亚基的关联，这被认为对增强DNA合成的增强很重要。这项研究是为了鉴定P125并阐明其在IGF-I诱导的DNA合成增强中的作用。通过MALDI-TOF MS分析显示，在cAMP刺激的FRTL-5细胞中，免疫沉淀的磷酸酪糖基P125是人XB130的大鼠直系同源物，我们将其命名为磷脂酰肌醇3-激酶与磷脂型蛋白质酶相关蛋白（PI3KAP）。 CAMP处理升高PI3KAP/XB130 mRNA和蛋白质水平以及酪氨酸磷酸化和PI3KAP/XB130与P85 PI3K的相互作用，从而导致PI3K活性增加。重要的是，FRTL-5细胞中的PI3KAP/XB130敲低抑制了IGF-I诱导的DNA合成的CAMP依赖性增强。在营地治疗期间，添加了SRC家族激酶抑制剂PP1或PP2，废除了PI3KAP/XB130的酪氨酸磷酸化及其与P85 PI3K的相互作用。此外，C-SRC与PI3KAP/XB130相关，并因cAMP而被激活。总之，这些数据表明PI3KAP/XB130的cAMP依赖性诱导及其与PI3K的关联是增强IGF有丝分裂活性所必需的。