Proximity proteomics reveals a co-evolved LRRK2-regulatory network linked to centrosomes

Abstract Leucine-rich repeat kinase 2 (LRRK2) not only plays a vital role in familial forms of Parkinson’s disease (PD) but also represents a risk factor for idiopathic PD. Its multi-domain architecture enables fine-tuned regulation of its biological function by orchestrating intra- and intermolecular interactions. Here, we present BioID proximity proteomes of LRRK2 that reveal new interactors, which we further characterize using a novel evolutionary and structural bioinformatics pipeline. Co-evolutionary analysis of the protein-protein interaction network identifies a structural and functional module enriched in cytoskeletal components associated with the centrosome and microtubules. In addition, structural modelling of binary interactions using AlphaFold-Multimer reveals distinct groups of interactors that engage LRRK2 in a manner dependent on specific conformations and epitopes. Furthermore, we identify distinct changes in the LRRK2 proximity proteome that are induced by the type I kinase inhibitor MLi-2 or by co-expression of the LRRK2 upstream effector RAB29. Depending on its activity state and conformation, these protein-protein interactions link LRRK2 to defined cellular sub-compartments, including centriolar satellites and vesicular sub-compartments. Summary This study defines a comprehensive, LRRK2 interaction network by combining proximity proteomics with evolutionary and structural analyses, revealing how LRRK2 conformation and activity state dictate distinct protein-protein interactions. These findings link LRRK2 to conserved centrosomal, cytoskeletal, and vesicular pathways, providing mechanistic insight into its role in Parkinson’s disease. Key results Centrosome-linked LRRK2 network: BioID combined with co-evolutionary analysis identifies a highly conserved module enriched in centrosomal and microtubule-associated proteins, including CYLD as a top co-evolved interactor. Conformation-specific interactions: AlphaFold-Multimer modelling reveals distinct classes of LRRK2 interactors that bind different domains and conformations (“locked” vs. “unlocked”), corresponding to specific cellular functions. State-dependent rewiring of the interactome: The type I inhibitor MLi-2 selectively redirects LRRK2 to centriolar satellite proteins, whereas RAB29 co-expression shifts LRRK2 interactions toward vesicular and lysosomal pathways.