Membrane-binding and activation mechanism of PTEN
Membrane-binding and activation mechanism of PTEN
复制标题
DOI：
10.1073/pnas.0932835100
复制
发表时间：
2003-06-24
影响因子：
11.1
通讯作者：
Cho, WW
中科院分区：
文献类型：
--
作者：
Das, S;Dixon, JE;Cho, WW
PTEN是一种肿瘤抑制因子，其通过催化磷酸肌醇的3'磷酸的去除来逆转磷酸肌醇3-激酶的作用。尽管PTEN在细胞信号传导和调节中起着关键作用，但其膜募集和激活的机制仍然知之甚少。PTEN由N-末端磷酸酶结构域、C2结构域和C-末端尾区组成，所述C-末端尾区含有PSD-95/Dlg/ZO-1同源性（PDZ）结构域结合序列和多个磷酸化位点。我们在体外表面等离子体共振测量使用固定囊泡表明，磷酸酶结构域和C2域，但不是C-末端尾，参与静电膜结合的PTEN。此外，在PTEN的C-末端尾部的磷酸化模拟突变导致其膜亲和力降低约80倍，主要是通过减缓膜结合步骤。转染到HEK 293 T和HeLa细胞中的PTEN的亚细胞定位研究表明，靶向PTEN到质膜与快速降解偶联，并且磷酸酶结构域和C2结构域对于其膜募集都是必要的和足够的。结果还表明，磷酸化调节PTEN的靶向质膜不是通过阻断PDZ结构域结合位点，而是通过干扰静电膜结合的PTEN。在这些结果的基础上，我们提出了cl膜结合和激活机制的PTEN，其中的C-末端区域的磷酸化/去磷酸化作为一个静电开关，控制膜易位的蛋白质。
PTEN is a tumor suppressor that reverses the action of phosphoinositide 3-kinase by catalyzing the removal of the 3' phosphate of phosphoinositides. Despite the critical role of PTEN in cell signaling and regulation, the mechanisms of its membrane recruitment and activation is still poorly understood. PTEN is composed of an N-terminal phosphatase domain, a C2 domain, and a C-terminal tail region that contains the PSD-95/Dlg/ZO-1 homology (PDZ) domain-binding sequence and multiple phosphorylation sites. Our in vitro surface plasmon resonance measurements using immobilized vesicles showed that both the phosphatase domain and the C2 domain, but not the C-terminal tail, are involved in electrostatic membrane binding of PTEN. Furthermore, the phosphorylation-mimicking mutation on the C-terminal tail of PTEN caused an approximate to80-fold reduction in its membrane affinity, mainly by slowing the membrane-association step. Subcellular localization studies of PTEN transfected into HEK293T and HeLa cells indicated that targeting of PTEN to the plasma membrane is coupled with rapid degradation and that the phosphatase domain and the C2 domain are both necessary and sufficient for its membrane recruitment. Results also indicated that the phosphorylation regulates the targeting of PTEN to the plasma membrane not by blocking the PDZ domain-binding site but by interfering with electrostatic membrane binding of PTEN. On the basis of these results, we propose cl membrane-binding and activation mechanism for PTEN, in which the phosphorylation/dephosphorylation of the C-terminal region serves as an electrostatic switch that controls the membrane translocation of the protein.