SplitTurboID mapping of dimeric protein phosphatase complex interactomes

Unlike its counterpart Ser/Thr kinases, the predominant Ser/Thr protein phosphatase 1 (PP1) is a promiscuous enzyme that gains its subcellular localization and substrate specificity from a large panel of regulatory proteins with which it associates. Mapping the interactomes of catalytic and regulatory subunits on their own provides evidence for the targeting of PP1 to a wide range of cellular pathways, but does not provide a conditional analysis of individual holophosphatase complexes. To achieve this, we adapted a splitTurboID fragment complementation approach for the proximal interactome mapping of distinct catalytic-regulatory subunit pairs. We demonstrated the reliability and high spatial resolution of this technique, using an analysis pipeline that highlights unique and enriched near neighbors above shared common contaminants. In addition to confirming that a newly identified regulatory subunit has both PP1- and non-PP1-related associations, we further demonstrate the general applicability of this approach to other types of protein-protein pairs.