In vivo functional dissection of human inner kinetochore protein CENP-C

• Published in: Journal of structural biology 2002-10, Vol.140 (1), p.39-48
• Main Authors: Trazzi, Stefania, Bernardoni, Roberto, Diolaiti, Daniel, Politi, Valeria, Earnshaw, William C., Perini, Giovanni, Della Valle, Giuliano
• Format: Article
• Language: English
• Subjects: Blotting, Western; CENP-C; Centromere - chemistry; Centromere localization domain; Chromatin - metabolism; Chromosomal Proteins, Non-Histone - chemistry; Chromosomal Proteins, Non-Histone - genetics; Chromosomal Proteins, Non-Histone - physiology; Cross-Linking Reagents - pharmacology; DNA binding domain; DNA, Satellite - metabolism; Electrophoresis, Polyacrylamide Gel; Formaldehyde - pharmacology; Humans; Kinetochore; Kinetochores - metabolism; Microscopy, Fluorescence; Mutation; Precipitin Tests; Protein Structure, Tertiary; α-Satellite DNA
• ISSN: 1047-8477; 1095-8657
• DOI: 10.1016/S1047-8477(02)00506-3

CENP-C is a fundamental component of the inner kinetochore plate and contributes to the formation of functional centromeres in eukaryotic organisms. Recruitment of CENP-C to kinetochore requires other centromere proteins, particularly CENP-A, CENP-H, and CENP-I. However, how CENP-C is correctly localized at the kinetochore is not clearly determined, mainly due to the functional variety of its domains, which hints at a complex recruitment mechanism. Here, by both immunofluorescent labeling and chromatin/immunoprecipitation we could show that human CENP-C contains two distinct domains, one in the central region, between amino acids 426 and 537, and the second one in the carboxyl terminal region, between amino acids 638 and 943, which are both capable of localizing at centromeres and binding α-satellite DNA. The presence of two domains that iterate the same function despite being significantly different in their amino acid sequence and structure suggests that CENP-C may target the centromere by establishing multiple contacts with both the DNA and protein constituents of the kinetochore.