CTCF DNA binding domain undergoes dynamic and selective protein–protein interactions

CTCF is a predominant insulator protein required for three-dimensional chro-matin organization. However, the roles of its insulation of enhancers in a 3D nu-clear organization have not been fully explained. Here, we found that the CTCF DNA-binding domain (DBD) forms dynamic self-interacting clusters. Strikingly, CTCF DBD clusters were found to incorporate other insulator proteins but are not coenriched with transcriptional activators in the nucleus. This property is not observed in other domains of CTCF or the DBDs of other transcription fac-tors. Moreover, endogenous CTCF shows a phenotype consistent with the DBD by forming small protein clusters and interacting with CTCF motif arrays that have fewer transcriptional activators bound. Our results reveal an interesting phenomenon in which CTCF DBD interacts with insulator proteins and selectively localizes to nuclear positions with lower concentrations of transcriptional activa-tors, providing insights into the insulation function of CTCF.