Protein arginine methyltransferase 5 (PRMT5) is a Type II arginine methyltransferase and an essential enzyme. PRMT5 symmetrically dimethylates substrate proteins by catalyzing the 2-step transfer of 2 methyl groups from 2 S-adenosyl methionine (SAM) co-factor molecules to specific arginine residues

PRMT5 is an arginine methyltransferase and a therapeutic target in MTAP null cancers. PRMT5 utilizes adaptor proteins for substrate recruitment through a previously undefined mechanism. Here, we identify an evolutionarily conserved peptide sequence shared among the three known substrate adaptors (pICln/CLNS1A, RIOK1 and COPR5) and show it is necessary and sufficient for interaction with PRMT5. We structurally resolve the interface with PRMT5 and show via genetic perturbation that it is required for methylation of adaptor-recruited substrates including the spliceosome, histones, and ribosome assembly complexes. Genetic disruption of the PRMT5-substrate adaptor interface leads to a hypomorphic decrease in growth of MTAP null tumor cells and is thus a novel site for development of therapeutic inhibitors of PRMT5.