Transient Receptor Potential Canonical 5 Channels Activate Ca2+/Calmodulin Kinase Iγ to Promote Axon Formation in Hippocampal Neurons
Transient Receptor Potential Canonical 5 Channels Activate Ca2+/Calmodulin Kinase Iγ to Promote Axon Formation in Hippocampal Neurons
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DOI：
10.1523/jneurosci.1544-09.2009
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发表时间：
2009-08-05
影响因子：
5.3
通讯作者：
Wayman, Gary A.
中科院分区：
文献类型：
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作者：
Davare, Monika A.;Fortin, Dale A.;Wayman, Gary A.
神经元的功能依赖于树突和轴突的区室化极化。一个神经突相对于其他神经突的快速和选择性生长形成轴突在启动神经元极性中是至关重要的。轴突发生部分受最佳细胞内钙离子浓度的调节。我们的调查涉及轴突形成的Ca 2+信号通路，使用培养的海马神经元证明了Ca 2 +/钙调蛋白激酶激酶（CaMKK）及其下游靶Ca 2 +/钙调蛋白激酶I（CaMKI）的作用。组成型活性CaMKI的表达诱导形成多个轴突，而阻断CaMKK或CaMKI活性与药理学，显性负，或短发夹RNA（shRNA）的方法显着抑制轴突的形成。CaMKK信号通过CaMKI的γ-同种型作为对CaMKI γ的shRNA，而不是其他CaMKI同种型，抑制轴突形成。此外，过度表达野生型CaMKI γ，但不是一个突变体不能膜协会，加速轴突形成的速度。存在于发育中的轴突生长锥中的瞬时受体电位典型5（TRPC 5）通道的药理学或小干扰RNA抑制抑制了CaMKI γ的CaMKK介导的活化以及轴突形成。我们证明，使用生化分级分离和免疫细胞化学，CaMKI γ和TRPC 5共定位的脂筏。这些结果与其中通过TRPC 5通道的高度局部化的钙内流激活CaMKK和CaMKI γ的模型一致，所述CaMKK和CaMKI γ随后促进轴突形成。
Functionality of neurons is dependent on their compartmentalized polarization of dendrites and an axon. The rapid and selective outgrowth of one neurite, relative to the others, to form the axon is critical in initiating neuronal polarity. Axonogenesis is regulated in part by an optimal intracellular calcium concentration. Our investigation of Ca2+-signaling pathways involved in axon formation using cultured hippocampal neurons demonstrates a role for Ca2+/calmodulin kinase kinase (CaMKK) and its downstream target Ca2+/calmodulin kinase I (CaMKI). Expression of constitutively active CaMKI induced formation of multiple axons, whereas blocking CaMKK or CaMKI activity with pharmacological, dominant-negative, or short hairpin RNA (shRNA) methods significantly inhibited axon formation. CaMKK signals via the gamma-isoform of CaMKI as shRNA to CaMKI gamma, but not the other CaMKI isoforms, inhibited axon formation. Furthermore, overexpression of wild-type CaMKI gamma, but not a mutant incapable of membrane association, accelerated the rate of axon formation. Pharmacological or small interfering RNA inhibition of transient receptor potential canonical 5 (TRPC5) channels, which are present in developing axonal growth cones, suppressed CaMKK-mediated activation of CaMKI gamma as well as axon formation. We demonstrate using biochemical fractionation and immunocytochemistry that CaMKI gamma and TRPC5 colocalize to lipid rafts. These results are consistent with a model in which highly localized calcium influx through the TRPC5 channels activates CaMKK and CaMKI gamma, which subsequently promote axon formation.