Deurloo M H S, Eide S, Turlova E, Li Q, Spijker S, Sun H-S, Groffen A J A, Feng Z-P
Department of Physiology, University of Toronto, Toronto, Canada.
Department Molecular and Cellular Neurobiology, Neurogenomics and Cognition Research, VU University of Amsterdam, Amsterdam, The Netherlands.
Cell Biosci. 2024 Jan 21;14(1):13. doi: 10.1186/s13578-024-01193-w.
Rasal1 is a Ras GTPase-activating protein which contains C2 domains necessary for dynamic membrane association following intracellular calcium elevation. Membrane-bound Rasal1 inactivates Ras signaling through its RasGAP activity, and through such mechanisms has been implicated in regulating various cellular functions in the context of tumors. Although highly expressed in the brain, the contribution of Rasal1 to neuronal development and function has yet to be explored.
We examined the contributions of Rasal1 to neuronal development in primary culture of hippocampal neurons through modulation of Rasal1 expression using molecular tools. Fixed and live cell imaging demonstrate diffuse expression of Rasal1 throughout the cell soma, dendrites and axon which localizes to the neuronal plasma membrane in response to intracellular calcium fluctuation. Pull-down and co-immunoprecipitation demonstrate direct interaction of Rasal1 with PKC, tubulin, and CaMKII. Consequently, Rasal1 is found to stabilize microtubules, through post-translational modification of tubulin, and accordingly inhibit dendritic outgrowth and branching. Through imaging, molecular, and electrophysiological techniques Rasal1 is shown to promote NMDA-mediated synaptic activity and CaMKII phosphorylation.
Rasal1 functions in two separate roles in neuronal development; calcium regulated neurite outgrowth and the promotion of NMDA receptor-mediated postsynaptic events which may be mediated both by interaction with direct binding partners or calcium-dependent regulation of down-stream pathways. Importantly, the outlined molecular mechanisms of Rasal1 may contribute notably to normal neuronal development and synapse formation.
Rasal1是一种Ras GTP酶激活蛋白，含有细胞内钙升高后动态膜结合所必需的C2结构域。膜结合的Rasal1通过其RasGAP活性使Ras信号失活，并通过这种机制参与调节肿瘤背景下的各种细胞功能。尽管Rasal1在脑中高度表达，但其对神经元发育和功能的贡献尚未得到探索。
我们通过使用分子工具调节Rasal1的表达，研究了Rasal1在海马神经元原代培养中对神经元发育的贡献。固定和活细胞成像显示，Rasal1在整个细胞体、树突和轴突中呈弥漫性表达，在细胞内钙波动时定位于神经元质膜。下拉和共免疫沉淀显示Rasal1与PKC、微管蛋白和CaMKII直接相互作用。因此，发现Rasal1通过对微管蛋白的翻译后修饰来稳定微管，并相应地抑制树突的生长和分支。通过成像、分子和电生理技术表明，Rasal1可促进NMDA介导的突触活动和CaMKII磷酸化。
Rasal1在神经元发育中发挥两种不同的作用；钙调节神经突生长以及促进NMDA受体介导的突触后事件，这可能通过与直接结合伙伴的相互作用或下游途径的钙依赖性调节来介导。重要的是，Rasal1概述的分子机制可能对正常神经元发育和突触形成有显著贡献。
