CTCF protein recruits centromeric protein CENP‐E to the centromeric regions through unusual CTCF binding sites

Abstract

Centromere‐associated protein E (CENP‐E) is an integral component of the kinetochore, and has been shown to be essential for chromosome congression during mitosis. However, the molecular mechanism by which CENP‐E is recruited to the kinetochore is poorly understood. In the present study, we report that CENP‐E interacts directly with CTCF, the insulator protein that is involved in large scale organization of chromatin and in insulation. We demonstrate that α‐satellite repeats containing both CENP‐B boxes­­­ and CTCF binding sites in HeLa cells that are arrested in G2/M or Mitosis are co‐occupied by CTCF and CENP‐E. CENP‐E localization appears to depend on CTCF, because deletion of CTCF by siRNA disrupts binding of CENP‐E to the centromeric regions. The CTCF binding sites in centromeric regions are unusual in that most of them carry only the ‘M2′ sequence motif, a 9‐bp long DNA fragment that was originally thought to appear together with the core CTCF binding motif (also called ‘M1′ motif). Furthermore, we provide evidence that the ‘M2′ motif engages only the C‐terminal zinc fingers of CTCF, and binds CTCF quite strongly. Based on these findings, we suggest that CTCF helps recruit CENP‐E to the kinetochore and that the CTCF‐CENP‐E complex may represent a novel structural feature of centromeres.