Pim1 kinase promotes angiogenesis through phosphorylation of endothelial nitric oxide synthase at Ser-633
Pim1 kinase promotes angiogenesis through phosphorylation of endothelial nitric oxide synthase at Ser-633
复制标题
Pim1 激酶通过内皮一氧化氮合酶 Ser-633 的磷酸化促进血管生成。
DOI：
10.1093/cvr/cvv250
复制
发表时间：
2016-01-01
影响因子：
10.8
通讯作者：
Sun, Jianxin
中科院分区：
文献类型：
--
作者：
Chen, Ming;Yi, Bing;Sun, Jianxin
旨在
翻译后修饰，如磷酸化，在调节内皮型一氧化氮合酶（eNOS）的激活中起着至关重要的作用。在本研究中，我们的目的是确定是否eNOS可以磷酸化和调节的一种新的丝氨酸/苏氨酸蛋白激酶Pim 1在血管内皮细胞（EC）。
方法和结果
使用免疫沉淀和蛋白激酶测定，我们证明了Pim 1特异性地与eNOS相互作用，这导致eNOS在Ser-633处显著磷酸化并增加一氧化氮（NO）的产生。有趣的是，响应于VEGF刺激，eNOS在Ser-633处的磷酸化表现出两个不同的阶段：在0和60 min之间发生的瞬时磷酸化和在2和24 h之间发生的持续磷酸化，其分别由蛋白激酶A（PKA）和Pim 1介导。通过药理学抑制剂SMI-4a或显性负性形式的Pim 1抑制Pim 1显著减弱VEGF诱导的管形成，而Pim 1过表达以NO依赖性方式显著增加EC管形成和迁移。重要的是，在高糖处理的EC和db/db糖尿病小鼠的主动脉中，Ser-633处的Pim 1表达和eNOS磷酸化显著降低。Pim 1表达增加改善糖尿病小鼠血管新生受损，通过离体主动脉环测定确定。
结论
我们的研究结果表明，Pim 1作为一种新的激酶，负责eNOS在Ser-633的磷酸化，并增强糖尿病小鼠主动脉环的EC发芽，这表明Pim 1可能作为一种新的治疗靶点，用于血管重建策略。
AIMS
Posttranslational modification, such as phosphorylation, plays an essential role in regulating activation of endothelial NO synthase (eNOS). In the present study, we aim to determine whether eNOS could be phosphorylated and regulated by a novel serine/threonine-protein kinase Pim1 in vascular endothelial cells (ECs).
METHODS AND RESULTS
Using immunoprecipitation and protein kinase assays, we demonstrated that Pim1 specifically interacts with eNOS, which leads to a marked phosphorylation of eNOS at Ser-633 and increased production of nitric oxide (NO). Intriguingly, in response to VEGF stimulation, eNOS phosphorylation at Ser-633 exhibits two distinct phases: transient phosphorylation occurring between 0 and 60 min and sustained phosphorylation occurring between 2 and 24 h, which are mediated by the protein kinase A (PKA) and Pim1, respectively. Inhibiting Pim1 by either pharmacological inhibitor SMI-4a or the dominant-negative form of Pim1 markedly attenuates VEGF-induced tube formation, while Pim1 overexpression significantly increases EC tube formation and migration in an NO-dependent manner. Importantly, Pim1 expression and eNOS phosphorylation at Ser-633 were substantially decreased in high glucose-treated ECs and in the aorta of db/db diabetic mice. Increased Pim1 expression ameliorates impaired vascular angiogenesis in diabetic mice, as determined by an ex vivo aortic ring assay.
CONCLUSION
Our findings demonstrate Pim1 as a novel kinase that is responsible for the phosphorylation of eNOS at Ser-633 and enhances EC sprouting of aortic rings from diabetic mice, suggesting that Pim1 could potentially serve as a novel therapeutic target for revascularization strategies.