Mitsui Norihiro, Inatome Ryoko, Takahashi Shusuke, Goshima Yoshio, Yamamura Hirohei, Yanagi Shigeru
Division of Proteomics, Department of Genome Sciences, Kobe University Graduate School of Medicine, Chuo-ku, Kobe, 650-0017, Japan.
EMBO J. 2002 Jul 1;21(13):3274-85. doi: 10.1093/emboj/cdf328.
Collapsin response mediator proteins (CRMPs)/TOAD64/Ulips/DRPs and CRAM have emerged as strong candidates for a role in semaphorin signaling. In this study we identified Fes/Fps (Fes) tyrosine kinase in the CRMP-CRAM complex and investigated whether Fes was involved in semaphorin3A (Sema3A) signaling. In COS-7 cells, the interaction between Fes and plexinA1 (PlexA1) and the tyrosine phosphorylation of PlexA1 by Fes were observed; however, these events were significantly attenuated by co-expression of neuropilin-1 (NP-1). Even with NP-1 co-expression, Sema3A was able to enhance the association of Fes with PlexA1 and Fes-mediated tyrosine phosphorylation of PlexA1, CRAM and CRMP2. Co-expression of Fes with PlexA1 exhibited COS-7 cell contraction activity, indicating that Fes can convert inactive PlexA1 to its active form, whereas combination of Fes/NP-1/PlexA1 or Fes kinase-negative mutants/PlexA1 did not alter cell morphology. Finally, Sema3A-induced growth cone collapse of dorsal root ganglion neurons was suppressed by expression of Fes kinase-negative mutants. Taken together, our findings suggest that Fes links Sema3A signals to CRMP-CRAM, and that NP-1 negatively regulates PlexA1 activation by Fes in resting condition.
塌陷反应中介蛋白（CRMPs）/TOAD64/Ulips/DRPs和CRAM已成为在信号素信号传导中发挥作用的有力候选者。在本研究中，我们在CRMP-CRAM复合物中鉴定出Fes/Fps（Fes）酪氨酸激酶，并研究Fes是否参与信号素3A（Sema3A）信号传导。在COS-7细胞中，观察到Fes与丛状蛋白A1（PlexA1）之间的相互作用以及Fes对PlexA1的酪氨酸磷酸化作用；然而，通过共表达神经纤毛蛋白-1（NP-1），这些事件显著减弱。即使共表达NP-1，Sema3A仍能够增强Fes与PlexA1的结合以及Fes介导的PlexA1、CRAM和CRMP2的酪氨酸磷酸化。Fes与PlexA1共表达表现出COS-7细胞收缩活性，表明Fes可以将无活性的PlexA1转化为其活性形式，而Fes/NP-1/PlexA1或Fes激酶阴性突变体/PlexA1的组合并未改变细胞形态。最后，Fes激酶阴性突变体的表达抑制了Sema3A诱导的背根神经节神经元生长锥塌陷。综上所述，我们的研究结果表明，Fes将Sema3A信号与CRMP-CRAM联系起来，并且在静息状态下NP-1负向调节Fes对PlexA1的激活。
