Structural insights into the intramolecular interactions of centromere protein CENP-I.

In mitosis, the accurate segregation of sister chromosomes relies on kinetochore, a multiple subunits complex assembled on centromere of each sister chromosome. As a core component of inner kinetochore, CENP-I plays important functions to mediate kinetochore assembly and supports the faithful chromosome segregation. The structures of the N-terminus and C-terminus of CENP-I homologs in complex with CENP-H/K have been reported, respectively. Unfortunately, the intramolecular interactions of CENP-I are poorly understood, and how CENP-I interacts with CENP-M remains unknown. Here, we verified a unique helix α11, which forms the intramolecular interactions with N-terminal HEAT repeats in fungal CENP-I. Deletion of the helix α11 exposed the hydrophobic surface and resulted in the in vitro protein aggregation of N-terminal HEAT repeats of fungal CENP-I. The corresponding helix and its intramolecular interaction are highly conserved in human CENP-I. Deletion of the corresponding helix in human CENP-I dramatically reduced the functional activity to interact with CENP-H and CENP-M. Mutations of the conserved residues on the helix in human CENP-I significantly weakened the binding to CENP-M, but not CENP-H, in HeLa cells. Therefore, our findings for the first time unveiled a conserved helix of CENP-I, which is important for the intramolecular interaction and function, and would be helpful for understanding the structure basis of how CENP-I mediates the kinetochore assembly during cell cycle and mitosis.