A LisH-domain protein interaction map reveals a Lis1-ARIH2-dynein regulatory axis.

LisH-domain-containing proteins are involved in diverse cellular processes and disease mechanisms, yet their functional interaction landscape remains poorly characterized. Here, we employed a proteomics-based strategy to systematically map the interaction network of 27 LisH-domain-containing proteins, uncovering 1,410 high-confidence interactions-90% of which are previously unreported. This network reveals unanticipated roles for LisH proteins in cellular regulation and uncovers links to human disease, including cancer-associated interactions. Focusing on Lis1, a cytoplasmic dynein regulator, we identify the RBR-E3 ubiquitin ligase ARIH2 as a key functional interactor. We show that Lis1 promotes ARIH2 deneddylation via the COP9 signalosome (CSN), modulating its ubiquitin ligase activity. Active, neddylated ARIH2 ubiquitinates dynein intermediate chain 1 (DIC1), thus facilitating dynein function in proper organelle positioning. Our findings provide a comprehensive LisH protein interaction network and uncover a regulatory axis involving Lis1-ARIH2 that is important for dynein dependent intracellular transport.