Integrating endogenous TurboID and data-independent acquisition mass spectrometry for in vivo proximity labeling

ABSTRACT: Proximity labeling has emerged as a powerful approach for identifying protein-protein interaction networks within living systems, particularly those involving weak or transient associations. Here, we present a comprehensive revised proximity labelling workflow, integrating TurboID labelling of endogenously expressed fusion proteins and data-independent acquisition (DIA) mass spectrometry (MS). We benchmark this pipeline with a study of five conserved Caenorhabditis elegans proteins -NEKL-2, NEKL-3, MLT-2, MLT-3, and MLT-4- that form two NEKL-MLT kinase-scaffold subcomplexes involved in membrane trafficking and actin regulation. Profiling of NEKL-MLT interactomes across 23 experiments validated our approach through the identification of known NEKL-MLT binding partners and conserved nekl-mlt

SUBMITTER: David, S Fay 

PROVIDER: S-SCDT-10_1038-S44318-025-00660-5 | biostudies-other |

REPOSITORIES: biostudies-other