Phosphorylation and activation of PINOID by the phospholipid signaling kinase 3-phosphoinositide-dependent protein kinase 1 (PDK1) in Arabidopsis.
Phosphorylation and activation of PINOID by the phospholipid signaling kinase 3-phosphoinositide-dependent protein kinase 1 (PDK1) in Arabidopsis.
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DOI：
10.1073/pnas.0510283103
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发表时间：
2006-04
影响因子：
11.1
通讯作者：
H. Zegzouti;R. Anthony;N. Jahchan;L. Bögre;Sioux K. Christensen
中科院分区：
文献类型：
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作者：
H. Zegzouti;R. Anthony;N. Jahchan;L. Bögre;Sioux K. Christensen
丝氨酸-苏氨酸蛋白激酶PINFORMED（PID）的活性与生长素转运促进剂的膜相关PINFORMED（PIN）家族的不对称定位有关。然而，PID调节PIN蛋白分布的方式尚不清楚。我们已经使用重组PID蛋白在体外解剖PID活性的调节。我们证明，PID磷酸化外源性底物的能力，需要分子内PID自磷酸化。PID样哺乳动物AGC激酶在由磷脂相关激酶3-磷酸肌醇依赖性蛋白激酶1（PDK 1）启动的磷酸化级联中起作用，PDK 1结合PDK 1底物中发现的C-末端疏水PDK 1相互作用片段（PIF）结构域。我们发现拟南芥PDK 1与PID相互作用，并且PDK 1的转磷酸化增加PID自身磷酸化。我们发现，PID激活环丝氨酸是PDK 1依赖的PID磷酸化所必需的。这种激活是快速的，需要PIF结构域。花和幼苗的细胞提取物以组织特异性方式显著增加PID磷酸化。其中PIF结构域突变的PID蛋白变体未能被幼苗提取物激活。PID免疫沉淀拟南芥细胞中PDK 1的表达被抑制的RNAi表现出显着减少髓鞘碱性蛋白底物的转磷酸化。这些结果表明，AtPDK 1是一个有效的PID活性增强剂，并提供证据表明，磷脂信号可能在控制极性生长素运输的信号转导过程中发挥作用。
Activity of the serine-threonine protein kinase PINOID (PID) has been implicated in the asymmetrical localization of the membrane-associated PINFORMED (PIN) family of auxin transport facilitators. However, the means by which PID regulates PIN protein distribution is unknown. We have used recombinant PID protein to dissect the regulation of PID activity in vitro. We demonstrate that intramolecular PID autophosphorylation is required for the ability of PID to phosphorylate an exogenous substrate. PID-like mammalian AGC kinases act in a phosphorylation cascade initiated by the phospholipid-associated kinase, 3-phosphoinositide-dependent protein kinase 1 (PDK1), which binds to the C-terminal hydrophobic PDK1-interacting fragment (PIF) domain found in PDK1 substrates. We find that Arabidopsis PDK1 interacts with PID, and that transphosphorylation by PDK1 increases PID autophosphorylation. We show that a PID activation loop serine is required for PDK1-dependent PID phosphorylation. This activation is rapid and requires the PIF domain. Cell extracts from flowers and seedling shoots dramatically increase PID phosphorylation in a tissue-specific manner. A PID protein variant in which the PIF domain was mutated failed to be activated by the seedling shoot extracts. PID immunoprecipitated from Arabidopsis cells in which PDK1 expression was inhibited by RNAi showed a dramatic reduction in transphosphorylation of myelin basic protein substrate. These results indicate that AtPDK1 is a potent enhancer of PID activity and provide evidence that phospholipid signaling may play a role in the signaling processes controlling polar auxin transport.