Characterization of mouse embryonic stem (ES) clonal cell lines expressing G9a bearing point mutations in the cage and non-cage residues of the ankyrin repeat (ANK) domain. (A) Top panel: Diagram of full-length mG9a showing amino acid sequence numbers and specific domains: E, Glu-rich; Cys, Cys-rich ring finger-like; ANK, ankyrin repeat; SET, methyltransferase; and Pre and Post, Cys-rich Pre-SET and post-SET. Bottom panel: Schematic representation of the structural basis for ankyrin repeat (green) recognition of dimethylated histone H3 Lysine 9 (gray). G9a residue numbers are shown. H3K9me2 binds in a partial hydrophobic cage composed of residues highlighted in red. Peptide binding is further specified by the interaction of non-cage G9a residues (highlighted in purple) with H3 Ser10, Thr11, Gly12 and Gly13. (B) Immunoblots (IB) were performed on the indicated proteins (with GAPDH as loading control) using whole cell extracts derived from wild-type (TT2) mouse embryonic stem cells (mESCs), G9a-null mESCs (knock-out; KO) or distinct clonal ESC lines (indicated by #1 and #2) stably expressing wild-type G9a (+WT) or G9a harboring the indicated point mutations in the ANK domain.