Structure-function analysis of the EF-hand protein centrin-2 for its intracellular localization and nucleotide excision repair

• Published in: Nucleic acids research Vol. 41; no. 14; pp. 6917 - 6929
• Main Authors: Nishi, Ryotaro, Sakai, Wataru, Tone, Daisuke, Hanaoka, Fumio, Sugasawa, Kaoru
• Format: Journal Article
• Language: English
• Published: England Oxford University Press 01.08.2013
• Subjects: Calcium-Binding Proteins - analysis; Calcium-Binding Proteins - chemistry; Calcium-Binding Proteins - metabolism; Cell Cycle Proteins - analysis; Cell Cycle Proteins - chemistry; Cell Cycle Proteins - metabolism; Cell Line; Cell Nucleus - chemistry; DNA Damage; DNA Repair; DNA-Binding Proteins - metabolism; Genome Integrity, Repair and; Humans; Protein Structure, Tertiary; Structure-Activity Relationship; Ultraviolet Rays; Xeroderma Pigmentosum Group A Protein - metabolism
• ISSN: 0305-1048; 1362-4962
• DOI: 10.1093/nar/gkt434

Centrin-2 is an evolutionarily conserved, calmodulin-related protein, which is involved in multiple cellular functions including centrosome regulation and nucleotide excision repair (NER) of DNA. Particularly to exert the latter function, complex formation with the XPC protein, the pivotal NER damage recognition factor, is crucial. Here, we show that the C-terminal half of centrin-2, containing two calcium-binding EF-hand motifs, is necessary and sufficient for both its localization to the centrosome and interaction with XPC. In XPC-deficient cells, nuclear localization of overexpressed centrin-2 largely depends on co-overexpression of XPC, and mutational analyses of the C-terminal domain suggest that XPC and the major binding partner in the centrosome share a common binding surface on the centrin-2 molecule. On the other hand, the N-terminal domain of centrin-2 also contains two EF-hand motifs but shows only low-binding affinity for calcium ions. Although the N-terminal domain is dispensable for enhancement of the DNA damage recognition activity of XPC, it contributes to augmenting rather weak physical interaction between XPC and XPA, another key factor involved in NER. These results suggest that centrin-2 may have evolved to bridge two protein factors, one with high affinity and the other with low affinity, thereby allowing delicate regulation of various biological processes.