Mapping adipocyte interactome networks by HaloTag-enrichment–mass spectrometry

Mapping protein interaction complexes in their natural state in a cell in vivo is a holy grail of protein network analysis. We here describe the development of HaloMS, a high-throughput HaloTag-based affinity purification–mass spectrometry assay for protein interaction discovery. The approach enabled quick capture of newly expressed protein and eliminated the tedious conventional one-by-one assay. As a proof-of-principle, we used HaloMS to evaluate protein complex interactions of 17 regulatory proteins in human adipocyte. The adipocyte interactome network was validated using an in vitro pull-down assay and artificial intelligence-based prediction tools. The application of HaloMS to probe adipocyte differentiation signaling pathways allowed the identification of previously unknown transcription factor–protein complexes and revealed proteome-wide human adipocyte transcription factor networks. This study using new technology reveals hidden networks of adipocyte signaling pathways, shedding light on how signals from different pathways are integrated.