Importin-beta and the small guanosine triphosphatase Ran mediate chromosome loading of the human chromokinesin Kid.
Nucleocytoplasmic transport factors mediate various cellular processes, including nuclear transport, spindle assembly, and nuclear envelope/pore formation. In this paper, we identify the chromokinesin human kinesin-like DNA binding protein (hKid) as an import cargo of the importin-alpha/beta transport pathway and determine its nuclear localization signals (NLSs). Upon the loss of its functional NLSs, hKid exhibited reduced interactions with the mitotic chromosomes of living cells. In digitonin-permeabilized mitotic cells, hKid was bound only to the spindle and not to the chromosomes themselves. Surprisingly, hKid bound to importin-alpha/beta was efficiently targeted to mitotic chromosomes. The addition of Ran-guanosine diphosphate and an energy source, which generates Ran-guanosine triphosphate (GTP) locally at mitotic chromosomes, enhanced the importin-beta-mediated chromosome loading of hKid. Our results indicate that the association of importin-beta and -alpha with hKid triggers the initial targeting of hKid to mitotic chromosomes and that local Ran-GTP-mediated cargo release promotes the accumulation of hKid on chromosomes. Thus, this study demonstrates a novel nucleocytoplasmic transport factor-mediated mechanism for targeting proteins to mitotic chromosomes.
Tahara
Kiyoshi
K
Cellular Dynamics Laboratory, Discovery Research Institute, Institute of Physical and Chemical Research, Wako, Saitama, 351-0198, Japan.
Takagi
Masatoshi
M
Ohsugi
Miho
M
Sone
Takefumi
T
Nishiumi
Fumiko
F
Maeshima
Kazuhiro
K
Horiuchi
Yasuomi
Y
Tokai-Nishizumi
Noriko
N
Imamoto
Fumio
F
Yamamoto
Tadashi
T
Kose
Shingo
S
Imamoto
Naoko
N
D021581:Active Transport, Cell Nucleus / Q000235:genetics; D002467:Cell Nucleus / Q000235:genetics / Q000378:metabolism; D023902:Chromosome Pairing / Q000235:genetics*; D002875:Chromosomes / Q000235:genetics / Q000378:metabolism*; D003593:Cytoplasm / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D006160:Guanosine Triphosphate / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D016547:Kinesins / Q000235:genetics / Q000378:metabolism*; D008938:Mitosis / Q000235:genetics*; D009685:Nuclear Envelope / Q000235:genetics / Q000378:metabolism; D019913:Nuclear Localization Signals / Q000235:genetics; D010766:Phosphorylation; D021381:Protein Transport / Q000235:genetics; D008941:Spindle Apparatus / Q000235:genetics; D028901:alpha Karyopherins / Q000235:genetics; D028961:beta Karyopherins / Q000235:genetics / Q000502:physiology*; D020931:ran GTP-Binding Protein / Q000235:genetics*
D004268:DNA-Binding Proteins; C098764:KIF22 protein, human; D019913:Nuclear Localization Signals; D028901:alpha Karyopherins; D028961:beta Karyopherins; C087978:karyopherin alpha 2; D006160:Guanosine Triphosphate; D016547:Kinesins; D020931:ran GTP-Binding Protein