Platelet-derived growth factor BB induces nuclear export and proteasomal degradation of CREB via phosphatidylinositol. 3-kinase/Akt signaling in pulmonary artery smooth muscle cells
Platelet-derived growth factor BB induces nuclear export and proteasomal degradation of CREB via phosphatidylinositol. 3-kinase/Akt signaling in pulmonary artery smooth muscle cells
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DOI：
10.1128/mcb.02477-05
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发表时间：
2006-07-01
影响因子：
5.3
通讯作者：
Klemm, Dwight J.
中科院分区：
文献类型：
--
作者：
Garat, Chrystelle V.;Fankell, Dana;Klemm, Dwight J.
肺动脉高压动物肺动脉重构中的平滑肌细胞（SMCs）以及高血压个体致动脉粥样硬化的全身动脉SMC层和心肌细胞中的环AMP反应元件结合蛋白（CREB）含量降低。慢性暴露于缺氧或血小板衍生生长因子BB （PDGF-BB）可诱导培养的SMCs中CREB的丢失。在这里，我们研究了PDGF引起SMC CREB耗竭的信号通路和机制。慢性PDGF治疗增加了SMCs中CREB的泛素化，而用蛋白酶体抑制剂lactacystin治疗SMCs则阻止了CREB含量的下降。核输出抑制剂leptomycin B也可以单独或与lactacystin联合阻止SMC CREB的消耗。随后的研究表明，PDGF激活了SMCs中的细胞外信号调节激酶、Jun n端蛋白激酶和磷脂酰肌醇3 （PI3）激酶途径。抑制这些途径可阻断SMC对PDGF的增殖反应，但只有抑制pi3激酶或其效应物Akt才能阻断PDGF诱导的CREB损失。最后，将含有增强的青色荧光蛋白的嵌合蛋白与野生型CREB或在几个可识别的磷酸化位点发生突变的CREB分子连接到SMCs中。PDGF处理降低了这些嵌合蛋白的水平，除了相邻丝氨酸残基（丝氨酸103和107）的一个突变，这表明CREB的丢失依赖于这些位点的CREB磷酸化。我们得出结论，PDGF通过pi3 -激酶/Akt信号通路刺激SMCs的核输出和CREB的蛋白酶体降解。这些结果表明，除了直接磷酸化外，蛋白质水解和细胞内定位是调节SMCs中CREB含量和活性的关键机制。
Cyclic AMP response element binding protein (CREB) content is diminished in smooth muscle cells (SMCs) in remodeled pulmonary arteries from animals with pulmonary hypertension and in the SMC layers of atherogenic systemic arteries and cardiornyocytes from hypertensive individuals. Loss of CREB can be induced in cultured SMCs by chronic exposure to hypoxia or platelet-derived growth factor BB (PDGF-BB). Here we investigated the signaling pathways and mechanisms by which PDGF elicits depletion of SMC CREB. Chronic PDGF treatment increased CREB ubiquitination in SMCs, while treatment of SMCs with the proteasome inhibitor lactacystin prevented decreases in CREB content. The nuclear export inhibitor leptomycin B also prevented depletion of SMC CREB alone or in combination with lactacystin. Subsequent studies showed that PDGF activated extracellular signal-regulated kinase, Jun N-terminal protein kinase, and phosphatidylinositol 3 (PI3)-kinase pathways in SMCs. Inhibition of these pathways blocked SMC proliferation in response to PDGF, but only inhibition of PI3-kinase or its effector, Akt, blocked PDGF-induced CREB loss. Finally, chimeric proteins containing enhanced cyan fluorescent protein linked to wild-type CREB or CREB molecules with mutations in several recognized phosphorylation sites were introduced into SMCs. PDGF treatment reduced the levels of each of these chimeric proteins except for one containing mutations in adjacent serine residues (serines 103 and 107), suggesting that CREB loss was dependent on CREB phosphorylation at these sites. We conclude that PDGF stimulates nuclear export and proteasomal degradation of CREB in SMCs via PI3-kinase/Akt signaling. These results indicate that in addition to direct phosphorylation, proteolysis and intracellular localization are key mechanisms regulating CREB content and activity in SMCs.