Muramoto Masaya, Hanawa Nozomi, Okumura Misako, Chihara Takahiro, Miura Masayuki, Shinoda Natsuki
Department of Genetics, Graduate School of Pharmaceutical Sciences, The University of Tokyo, Tokyo, Japan.
Program of Biomedical Science, Graduate School of Integrated Sciences for Life, Hiroshima University, Higashi-Hiroshima, Hiroshima, Japan.
Elife. 2025 Jun 17;13:RP99650. doi: 10.7554/eLife.99650.
The nervous system undergoes functional modification independent of cell turnover. Caspase participates in reversible neuronal modulation via non-lethal activation. However, the mechanism that enables non-lethal activation remains unclear. Here, we analyzed proximal proteins of executioner caspase in the adult brain using TurboID. We discovered that executioner caspase Drice is, as an inactive proform, proximal to cell membrane proteins, including a specific splicing isoform of cell adhesion molecule Fasciclin 3 (Fas3), Fas3G. To investigate whether sequestration of executioner caspase to plasma membrane of axons is the mechanism for non-lethal activation, we developed a Gal4-Manipulated Area-Specific CaspaseTracker/CasExpress system for sensitive monitoring of caspase activity near the plasma membrane. We demonstrated that overexpression promotes caspase activation in olfactory receptor neurons without killing them, by inducing expression of initiator caspase Dronc, which also comes close to Fas3G. Physiologically, overexpression-facilitated non-lethal caspase activation suppresses innate olfactory attraction behavior. Our findings suggest that subcellularly restricted caspase activation, defined by caspase-proximal proteins, is the mechanism for non-lethal activation, opening the methodological development of reversible modification of neuronal function via regulating caspase-proximal proteins.
神经系统会经历独立于细胞更替的功能修饰。半胱天冬酶通过非致死性激活参与可逆的神经元调节。然而，实现非致死性激活的机制仍不清楚。在此，我们使用TurboID分析了成年大脑中执行性半胱天冬酶的近端蛋白。我们发现，执行性半胱天冬酶Drice以无活性的前体形式靠近细胞膜蛋白，包括细胞粘附分子Fasciclin 3（Fas3）的一种特定剪接异构体Fas3G。为了研究将执行性半胱天冬酶隔离到轴突质膜是否是非致死性激活的机制，我们开发了一种Gal4操纵的区域特异性半胱天冬酶追踪器/半胱天冬酶表达系统，用于灵敏监测质膜附近的半胱天冬酶活性。我们证明，过表达通过诱导起始半胱天冬酶Dronc的表达促进嗅觉受体神经元中的半胱天冬酶激活，而Dronc也靠近Fas3G，且不会杀死这些神经元。在生理上，过表达促进的非致死性半胱天冬酶激活会抑制先天性嗅觉吸引行为。我们的研究结果表明，由半胱天冬酶近端蛋白定义的亚细胞限制性半胱天冬酶激活是非致死性激活的机制，开启了通过调节半胱天冬酶近端蛋白对神经元功能进行可逆修饰的方法学发展。
