Ageta-Ishihara Natsumi, Takemoto-Kimura Sayaka, Kondo Yayoi, Okamura Michiko, Bito Haruhiko
Department of Biomolecular Science, Faculty of Science, Toho University, Funabashi, Japan.
PRESTO, Japan Science and Technology Agency, Kawaguchi, Japan.
Front Cell Neurosci. 2023 Aug 3;17:1204302. doi: 10.3389/fncel.2023.1204302. eCollection 2023.
CLICK-III/CaMKIγ is a lipid-anchored neuronal isoform of multifunctional Ca/calmodulin-dependent protein kinases, which mediates BDNF-dependent dendritogenesis in cultured cortical neurons. We found that two distinct lipidation states of CaMKIγ, namely, prenylation and palmitoylation, controlled its association with detergent-resistant microdomains in the dendrites and were essential for its dendritogenic activity. However, the impact of each lipid modification on membrane targeting/trafficking and how it specifies functional coupling leading to polarized changes in neuronal morphology are not clear. Here, we show that prenylation induces membrane anchoring of CaMKIγ, permitting access to the Golgi apparatus, and a subsequent palmitoylation facilitates association with cholesterol-enriched lipid microdomains or lipid rafts, in particular at the Golgi. To specifically test the role of palmitoylated CaMKγ in neurite extension, we identified and took advantage of a cell system, PC12, which, unlike neurons, conveniently lacked CaMKIγ and was deficient in the activity-dependent release of a neuritogenic growth factor while possessing the ability to activate polarized rafts signaling for morphogenesis. This system allowed us to rigorously demonstrate that an activity-dependent, lipid rafts-restricted Rac activation leading to neuritogenesis could be functionally rescued by dually lipidated CaMKIγ expression, revealing that not only prenylation but also palmitoylation is essential for CaMKIγ to activate a compartmentalized STEF-Rac1 pathway. These results shed light on the significance of recruiting prenylated and palmitoylated CaMKIγ into the coalescing signalosomes at lipid rafts together with Rac1 and its specific GEF and STEF and forming a compartmentalized Ca signaling pathway that underlies activity-dependent neuritogenesis and morphogenesis during axodendritic polarization critical for brain development and circuitogenesis.
CLICK-III/CaMKIγ是多功能钙/钙调蛋白依赖性蛋白激酶的一种脂质锚定神经元亚型，它在培养的皮层神经元中介导脑源性神经营养因子（BDNF）依赖性树突形成。我们发现CaMKIγ的两种不同脂质化状态，即异戊二烯化和棕榈酰化，控制着它与树突中抗去污剂微区的结合，并且对其树突生成活性至关重要。然而，每种脂质修饰对膜靶向/运输的影响以及它如何指定功能偶联从而导致神经元形态的极化变化尚不清楚。在这里，我们表明异戊二烯化诱导CaMKIγ的膜锚定，使其能够进入高尔基体，随后的棕榈酰化促进其与富含胆固醇的脂质微区或脂筏结合，特别是在高尔基体处。为了特异性测试棕榈酰化的CaMKγ在神经突延伸中的作用，我们鉴定并利用了一种细胞系统——PC12细胞，与神经元不同，该细胞方便地缺乏CaMKIγ，并且在活性依赖性神经生长因子释放方面存在缺陷，同时具有激活极化脂筏信号以进行形态发生的能力。这个系统使我们能够严格证明，通过双重脂质化的CaMKIγ表达可以功能性地挽救导致神经突生成的活性依赖性、脂筏限制的Rac激活，这表明不仅异戊二烯化而且棕榈酰化对于CaMKIγ激活分区化的STEF-Rac1途径都是必不可少的。这些结果揭示了将异戊二烯化和棕榈酰化的CaMKIγ与Rac1及其特异性鸟嘌呤核苷酸交换因子（GEF）和STEF一起募集到脂筏中合并的信号体中，并形成分区化的钙信号通路的重要性，该信号通路是脑发育和回路形成关键的轴突-树突极化过程中活性依赖性神经突生成和形态发生的基础。
