A novel nucleolar protein, NIFK, interacts with the forkhead associated domain of Ki-67 antigen in mitosis.
In a previous study, we demonstrated that the forkhead associated (FHA) domain of pKi-67 interacts with the novel kinesin-like protein, Hklp2 (Sueishi, M., Takagi, M., and Yoneda, Y. (2000) J. Biol. Chem. 275, 28888-28892). In this study, we report on the identification of a putative RNA-binding protein of 293 residues as another binding partner of the FHA domain of pKi-67 (referred to as NIFK for nucleolar protein interacting with the FHA domain of pKi-67). Human NIFK (hNIFK) interacted with the FHA domain of pKi-67 (Ki-FHA) efficiently in vitro when hNIFK was derived from mitotically arrested cells. In addition, a moiety of hNIFK was co-localized with pKi-67 at the peripheral region of mitotic chromosomes. The hNIFK domain that interacts with Ki-FHA was mapped in the yeast two-hybrid system to a portion encompassed by residues 226-269. In a binding assay utilizing Xenopus egg extracts, it was found that the mitosis-specific environment and two threonine residues within this portion of hNIFK (Thr-234 and Thr-238) were crucial for the efficient interaction of hNIFK and Ki-FHA, suggesting that hNIFK interacts with Ki-FHA in a mitosis-specific and phosphorylation-dependent manner. These findings provide a new clue to our understanding of the cellular function of pKi-67.
Takagi
M
M
Department of Cell Biology and Neuroscience, Graduate School of Medicine, Osaka University, 2-2 Yamada-oka, Suita, Osaka 565-0871, Japan.
Sueishi
M
M
Saiwaki
T
T
Kametaka
A
A
Yoneda
Y
Y
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002453:Cell Cycle; D003001:Cloning, Molecular; D029721:Drosophila Proteins; D006367:HeLa Cells; D006801:Humans; D019476:Insect Proteins / Q000737:chemistry*; D047908:Intracellular Signaling Peptides and Proteins; D019394:Ki-67 Antigen / Q000737:chemistry / Q000378:metabolism*; D051379:Mice; D008840:Microfilament Proteins; D008938:Mitosis; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010449:Peptide Mapping; D010766:Phosphorylation; D011485:Protein Binding; D016601:RNA-Binding Proteins; D013912:Threonine / Q000378:metabolism
D002352:Carrier Proteins; D029721:Drosophila Proteins; D019476:Insect Proteins; D047908:Intracellular Signaling Peptides and Proteins; D019394:Ki-67 Antigen; D008840:Microfilament Proteins; C492406:NIFK protein, human; C431942:NIFK protein, mouse; D009687:Nuclear Proteins; D016601:RNA-Binding Proteins; C084628:f protein, Drosophila; D013912:Threonine