Blue light-induced autophosphorylation of phototropin is a primary step for signaling
Blue light-induced autophosphorylation of phototropin is a primary step for signaling
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DOI：
10.1073/pnas.0709189105
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发表时间：
2008-04-08
影响因子：
11.1
通讯作者：
Shimazaki, Ken-ichiro
中科院分区：
文献类型：
--
作者：
Inoue, Shin-ichiro;Kinoshita, Toshinori;Shimazaki, Ken-ichiro
趋光蛋白是植物蓝光受体的自磷酸化蛋白激酶。它们调节各种蓝光反应，包括向光性、叶绿体运动、下胚轴生长抑制、叶片展平和气孔开放。然而，自磷酸化的生理作用仍然未知。在这里，我们确定了丝氨酸或苏氨酸的磷酸化位点的N端，铰链1区，激酶结构域，和C端的拟南芥phototropin 1（phot 1）通过液相色谱串联质谱在体内。我们取代这些Ser或Thr残基与Ala在phot 1和分析其功能，通过检查phot 1介导的气孔开放，向光性，叶绿体积累，叶片扁平化后的phot 1 phot 2双突变体的转化。在这些位点中，我们发现在激酶结构域的激活环中的Ser-851的自身磷酸化是上述反应所必需的，而除了激活环中的那些之外的其他Ser和Thr的磷酸化不是必需的。环路中的Ser-849可能在反应中起额外的作用。免疫学分析表明，Ser-851被蓝光以一种能量密度依赖的方式迅速磷酸化，并在黑暗中逐渐去磷酸化。我们的结论是，自磷酸化的Ser-851是一个主要的步骤，介导光化学反应和生理事件之间的信号。
Phototropins are autophosphorylating protein kinases of plant-specific blue light receptors. They regulate various blue light responses, including phototropism, chloroplast movements, hypocotyl growth inhibition, leaf flattening, and stomatal opening. However, the physiological role of autophosphorylation remains unknown. Here, we identified phosphorylation sites of Ser or Thr in the N terminus, Hinge1 region, kinase domain, and C terminus in Arabidopsis phototropin1 (phot1) by liquid chromatography-tandem mass spectrometry in vivo. We substituted these Ser or Thr residues with Ala in phot1 and analyzed their functions by inspecting the phot1-mediated responses of stomatal opening, phototropism, chloroplast accumulation, and leaf flattening after the transformation of the phot1 phot2 double mutant. Among these sites, we found that auto phosphorylation of Ser-851 in the activation loop of the kinase domain was required for the responses mentioned above, whereas the phosphorylation of the other Ser and Thr, except those in the activation loop, was not. Ser-849 in the loop may have an additional role in the responses. Immunological analysis revealed that Ser-851 was phosphorylated rapidly by blue light in a fluence-dependent manner and dephosphorylated gradually upon darkness. We conclude that autophosphorylation of Ser-851 is a primary step that mediates signaling between photochemical reaction and physiological events.