Interactome analysis of C. elegans synapses by TurboID-based proximity labeling

Proximity labeling provides a powerful in vivo tool to characterize the proteome of subcellular structures and the interactome of specific proteins. Using the highly active biotin ligase TurboID, we optimize a proximity labeling protocol for C. elegans. We use this protocol to characterise the proteomes of the worm’s gut, muscle, skin, and nervous system. We express TurboID exclusively in the pair of AFD neurons and show we can identify known and previously unknown proteins expressed selectively in AFD. We knock TurboID into the endogenous elks-1 gene, which encodes a presynaptic active zone protein. We identify many known ELKS-1 interacting proteins as well as previously uncharacterised synaptic proteins. Versatile vectors, and the inherent advantage of C. elegans for biochemistry, make proximity labeling a valuable addition to the nematode’s armory.

邻近标记（proximity labeling）技术是一种强大的体内研究工具，可用于表征亚细胞结构的蛋白质组以及特定蛋白质的相互作用组。本研究采用高活性生物素连接酶TurboID，优化了一套适用于秀丽隐杆线虫（C. elegans）的邻近标记实验方案。我们依托该方案对该线虫的肠道、肌肉、皮肤及神经系统的蛋白质组开展了表征分析。我们仅在一对AFD神经元中表达TurboID，证实可鉴定出AFD神经元特异性表达的已知蛋白质与此前未被发现的新型蛋白质。我们将TurboID敲入内源性elks-1基因（该基因编码突触前活性区蛋白），成功鉴定出多种已知的ELKS-1相互作用蛋白，以及此前未被表征的突触蛋白。多功能载体体系结合秀丽隐杆线虫在生物化学研究中的固有优势，使邻近标记技术成为该线虫研究工具库中极具价值的新增工具。