Human NUF2 interacts with centromere-associated protein E and is essential for a stable spindle microtubule-kinetochore attachment.
Chromosome segregation in mitosis is orchestrated by dynamic interaction between spindle microtubules and the kinetochore, a multiprotein complex assembled onto centromeric DNA of the chromosome. Here, we show that Homo sapiens (Hs) NUF2 is required for stable kinetochore localization of centromere-associated protein E (CENP-E) in HeLa cells. HsNUF2 specifies the kinetochore association of CENP-E by interacting with its C-terminal domain. The region of HsNUF2 binding to CENP-E was mapped to its C-terminal domain by glutathione S-transferase pulldown and yeast two-hybrid assays. Suppression of synthesis of HsNUF2 by small interfering RNA abrogated the localization of CENP-E to the kinetochore, demonstrating the requirement of HsNUF2 for CENP-E kinetochore localization. In addition, depletion of HsNUF2 caused aberrant chromosome segregation. These HsNUF2-suppressed cells displayed reduced tension at kinetochores of bi-orientated chromosomes. Double knockdown of CENP-E and HsNUF2 further abolished the tension at the kinetochores. Our results indicate that HsNUF2 and CENP-E are required for organization of stable microtubule-kinetochore attachment that is essential for faithful chromosome segregation in mitosis.
Liu
Dan
D
Laboratory of Cellular Dynamics, University of Science and Technology of China and the National Laboratory for Physical Sciences, Hefei 230027, China.
Ding
Xia
X
Du
Jian
J
Cai
Xin
X
Huang
Yuejia
Y
Ward
Tarsha
T
Shaw
Andrew
A
Yang
Yong
Y
Hu
Renming
R
Jin
Changjiang
C
Yao
Xuebiao
X
D018797:Cell Cycle Proteins / Q000737:chemistry*; D002460:Cell Line; D002503:Centromere / Q000378:metabolism*; D002868:Chromosomal Proteins, Non-Histone / Q000737:chemistry*; D002875:Chromosomes / Q000648:ultrastructure; D006367:HeLa Cells; D006801:Humans; D018386:Kinetochores / Q000378:metabolism*; D008870:Microtubules / Q000378:metabolism*; D008938:Mitosis; D008954:Models, Biological; D008957:Models, Genetic; D017434:Protein Structure, Tertiary; D034741:RNA, Small Interfering / Q000378:metabolism; D020798:Two-Hybrid System Techniques
D018797:Cell Cycle Proteins; D002868:Chromosomal Proteins, Non-Histone; C490197:NUF2 protein, human; D034741:RNA, Small Interfering; C068688:centromere protein E