Hedgehog-stimulated phosphorylation of the kinesin-related protein Costal2 is mediated by the serine/threonine kinase fused
Hedgehog-stimulated phosphorylation of the kinesin-related protein Costal2 is mediated by the serine/threonine kinase fused
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DOI：
10.1074/jbc.m110730200
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发表时间：
2002-07-05
影响因子：
4.8
通讯作者：
Bishop, JM
中科院分区：
文献类型：
--
作者：
Nybakken, KE;Turck, CW;Bishop, JM
Hedgehog（Hh）信号分子是果蝇中许多组织发育所必需的。在细胞内，Hh信号转导利用由融合（Fu）、Costal 2（Cos 2）和Cubitisinterruptus（Ci）蛋白组成的大蛋白复合物，但是这些蛋白之间的功能相互作用仍然在很大程度上未被表征。使用杆状病毒系统，我们表明，丝氨酸/苏氨酸激酶Fu磷酸化的驱动蛋白样蛋白Cos 2时，与Cos 2共表达。Cos 2和激酶失活形式的Fu的共表达消除了大部分Cos 2磷酸化。然后，我们表明，主要Fu诱导的磷酸化位点的Cos 2是丝氨酸572，而丝氨酸931磷酸化程度较低。丝氨酸572突变为丙氨酸消除了大多数，但不是全部，特定的磷酸肽的Cos 2时，共表达与Fu。我们还表明，由杆状病毒共表达与激酶死亡Fu产生的Cos 2的磷酸化模式几乎与从未刺激的S2细胞中分离的Cos 2的磷酸化模式相同。最后，由杆状病毒共感染野生型Fu产生的Cos 2的磷酸化模式与从Hh刺激的S2细胞分离的Cos 2的磷酸化模式几乎相同，表明丝氨酸572和931的磷酸化是真正的Hh信号传导事件。本研究阐明了Fu和Cos 2在Hh信号转导中的独特功能，并确定了驱动蛋白样分子的第二个已知磷酸化位点。
The Hedgehog (Hh) signaling molecule is required for the development of numerous tissues in Drosophila. Within the cell, Hh signal transduction utilizes a large protein complex consisting of the Fused (Fu), Costal2 (Cos2), and Cubitis interruptus (Ci) proteins, but the functional interactions between these proteins are still largely uncharacterized. Using a baculovirus system, we demonstrate that the serine/threonine kinase Fu phosphorylates the kinesin-like protein Cos2 when coexpressed with Cos2. Coexpression of Cos2 and a kinase-inactive version of Fu eliminates the majority of Cos2 phosphorylation. We then show that the primary Fu-induced phosphorylation site of Cos2 is serine 572, whereas serine 931 is phosphorylated to a lesser extent. Mutation of serine 572 to alanine eliminates most, but not all, specific phosphopeptides of Cos2 when coexpressed with Fu. We also demonstrate that the phosphorylation pattern of Cos2 produced by baculovirus coexpression with kinase-dead Fu is almost identical to the phosphorylation pattern of Cos2 isolated from unstimulated S2 cells. Finally, the phosphorylation pattern of Cos2 produced by baculovirus coinfection with wildtype Fu is almost identical to that of Cos2 isolated from S2 cells stimulated by Hh, indicating that phosphorylation of serines 572 and 931 is a genuine Hh signaling event. This study clarifies the unique functions of Fu and Cos2 in Hh signal transduction and identifies only the second known phosphorylation site of a kinesin-like molecule.