The binding and X-ray crystallographic data presented here show that the ankyrin repeats of G9a and GLP are capable of binding methylated histone tails. This is the first example of such a function by an ankyrin repeat domain. The specificity for mono- and dimethyllysine of the G9a and GLP ankyrin repeats is comparable to that of other methyl binding protein modules, such as the chromodomain, tudor domain and PHD finger10-14
, which can preferentially bind lysines methylated to specific degrees at specific histone tail residues. The organization of the methyllysine binding hydrophobic cage structure with one acidic residue in the ankyrin repeat domains of G9a and GLP is also similar to those found in the H4K20me2 binding double tudor domain of 53bp1 and, to a lesser extent, in the H3K4me3,2 binding PHD finger of BPTF (Supplementary Fig. 3
The histone code hypothesis suggests that specific epigenetic marks on histones can be translated into distinct biological outcomes through effectors that are recruited to these marks and subsequently act on the local chromatin structure or transcriptional machinery15,16
. Several histone-methylating complexes contain components to both synthesize and bind a specific histone mark, such as Suv39h–HP1 (for H3K9me3)17
and LSD1–BHC80 (for H3K4me0)18
, where the components that make (or remove) and recognize a specific histone mark are separate proteins. In these examples, both components are necessary for the overall function of the complex. G9a and GLP are the first example of histone-methylating proteins that contain modules, within the same polypeptide, for both making (via the SET domain) and recognizing (via the ankyrin repeats) a given methyl mark. The presence of a G9a–GLP heterodimer2
, however, may allow one enzyme to create the modification whereas the other enzyme binds this modification. In addition, the opposite approximately two-fold preference for either H3K9me1 or H3K9me2 by GLP and G9a, respectively, may better enable the G9a–GLP heterodimer to bind both of the H3K9me1 and H3K9me2 marks.
Recruitment of complexes containing G9a–GLP to regulated genes may be accomplished through direct interaction with sequence-specific DNA binding proteins or through the interaction with coactivator or co-repressor proteins. The studies presented here provide an alternative method of recruitment of G9a–GLP complexes through direct ankyrin repeat–mediated interaction with K9 methylation marks on histone tails. Although it is tempting to speculate that, similarly to the paired protein examples given above, G9a and GLP contribute these dual histone-methylating and methyl histone binding roles to the coactivator and co-repressor functions of complexes, further studies will be needed to elucidate the functional significance of this newly identified mode of histone tail recognition.