Structure of human methylosome–Lsm10/11 complex. (A) Schematic drawing of the human methylosome–Lsm10/11 complex. The molecules in the top layer are shown in color, and those in the bottom layer are in gray. Peptide segments from pICln and Lsm11, as well as the adenosine portion of the SAM cofactor, are shown in the sphere model. (B) Cryo-EM density for the Lsm11 peptide in the active site of PRMT5. The fitted atomic model is shown in the stick model. (C) Detailed interactions between the Lsm11 peptide and the active site of PRMT5. The structure of histone H4 peptide bound to the methylosome is shown in the overlay (in gray). (D) Electrostatic surface of the active site region of PRMT5. The Lsm11 peptide is shown in the stick model. The structure figures were produced with PyMOL (www.pymol.org). (E,F) WT Lsm11 encompassing the first 168 amino acids amino-terminally fused to MBP (MBP-168N) and indicated Lsm11 mutants were incubated with a mouse cytoplasmic extract. Proteins bound to Lsm11 were immobilized on amylose beads via MBP. The beads were split evenly into two halves and analyzed by western blotting (panel E) for the presence of methylosome components or resuspended in a buffer containing 3H SAM to test methylation of Lsm11, as detected using fluorography (panel F).
