Dynamic behavior of Nuf2-Hec1 complex that localizes to the centrosome and centromere and is essential for mitotic progression in vertebrate cells.
Nuf2 and Hec1 are evolutionarily conserved centromere proteins. To clarify the functions of these proteins in vertebrate cells, we characterized them in chicken DT40 cells. We generated GFP fusion constructs of Nuf2 and Hec1 to examine in detail the localization of these proteins during the cell cycle. We found that Nuf2 is associated with Hec1 throughout the cell cycle and that this complex is localized to the centrosomes during G1 and S phases and then moves through the nuclear membrane to the centromere in G2 phase. During mitosis, this complex is localized to the centromere. We also created conditional loss-of-function mutants of Nuf2 and Hec1. In both mutants, the cell cycle arrested at prometaphase, suggesting that the Nuf2-Hec1 complex is essential for mitotic progression. The inner centromere proteins CENP-A, -C, and -H and checkpoint protein BubR1 were localized to chromosomes in the mutant cells arrested at prometaphase, but Mad2 localization was abolished. Furthermore, photobleaching experiments revealed that the Nuf2-Hec1 complex is stably associated with the centromere and that interaction of this complex with the centrosome is dynamic.
Hori
Tetsuya
T
PRESTO, The Japan Science and Technology Corporation, National Institute of Genetics and The Graduate University for Advanced Studies, Mishima, Shizuoka 411-8540, Japan.
Haraguchi
Tokuko
T
Hiraoka
Yasushi
Y
Kimura
Hiroshi
H
Fukagawa
Tatsuo
T
D000818:Animals; D001324:Autoantigens / Q000235:genetics / Q000378:metabolism; D002135:Calcium-Binding Proteins / Q000235:genetics / Q000378:metabolism; D018797:Cell Cycle Proteins; D002478:Cells, Cultured; D002503:Centromere / Q000235:genetics / Q000378:metabolism*; D000076247:Centromere Protein A; D018385:Centrosome / Q000378:metabolism*; D002645:Chickens; D002868:Chromosomal Proteins, Non-Histone / Q000235:genetics / Q000378:metabolism; D002875:Chromosomes / Q000235:genetics / Q000378:metabolism; D003001:Cloning, Molecular; D003598:Cytoskeletal Proteins; D036681:Fluorescence Recovery After Photobleaching; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism*; D016193:G1 Phase / Q000235:genetics / Q000502:physiology; D016195:G2 Phase / Q000235:genetics / Q000502:physiology; D007150:Immunohistochemistry; D017404:In Situ Hybridization, Fluorescence; D064246:Mad2 Proteins; D008677:Metaphase / Q000235:genetics / Q000502:physiology; D009154:Mutation; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases; D012097:Repressor Proteins; D016196:S Phase / Q000235:genetics / Q000502:physiology
D001324:Autoantigens; C000619700:CENPA protein, human; C121933:CENPH protein, human; D002135:Calcium-Binding Proteins; D018797:Cell Cycle Proteins; D000076247:Centromere Protein A; D002868:Chromosomal Proteins, Non-Histone; D003598:Cytoskeletal Proteins; D005656:Fungal Proteins; C577697:MAD2L1 protein, human; D064246:Mad2 Proteins; C120529:NDC80 protein, human; C490197:NUF2 protein, human; D009687:Nuclear Proteins; D012097:Repressor Proteins; C060905:centromere protein C; D011494:Protein Kinases; C584277:BUB1 protein, human; C090464:Bub1 spindle checkpoint protein; D017346:Protein Serine-Threonine Kinases