A combinatorial domain screening platform reveals epigenetic effector interactions for transcriptional perturbation

Epigenetic regulation involves the coordinated interplay of diverse proteins. To systematically explore these combinations, we present COMBINE (combinatorial interaction exploration), a high-throughput platform that tests over 50,000 pairs of epigenetic effector domains up to 2,094 amino acids in length for their ability to modulate endogenous human gene transcription. COMBINE reveals diverse synergistic interactions between epigenetic protein domains, including a potent KRAB-L3MBTL3 fusion that increases the effective targeting window, enhances gene silencing in dose-limited conditions, and enables robust dual-directional CRISPR perturbation. Inducible screening shows DNA methylation modifiers are essential for epigenetic memory, with distinct combinations driving long-term repression and activation. This systematic analysis of pairwise domain interactions advances our understanding of epigenetic crosstalks and the development of next-generation epigenome editing tools. More broadly, COMBINE offers a generalizable platform to functionally characterize combinatorial biological processes at scale.