Spindle pole regulation by a discrete Eg5-interacting domain in TPX2.
Targeting protein for Xklp2 (TPX2) activates the Ser/Thr kinase Aurora A in mitosis and targets it to the mitotic spindle [1, 2]. These effects on Aurora A are mediated by the N-terminal domain of TPX2, whereas a C-terminal fragment has been reported to affect microtubule nucleation [3]. Using the Xenopus system, we identified a novel role of TPX2 during mitosis. Injection of TPX2 or its C terminus (TPX2-CT) into blastomeres of two-cell embryos led to potent cleavage arrest. Despite cleavage arrest, TPX2-injected embryos biochemically undergo multiple rounds of DNA synthesis and mitosis, and arrested blastomeres have abnormal spindles, clustered centrosomes, and an apparent failure of cytokinesis. In Xenopus S3 cells, transfection of TPX2-FL causes spindle collapse, whereas TPX2-CT blocks pole segregation, resulting in apposing spindle poles with no evident displacement of Aurora A. Analysis of TPX2-CT deletion peptides revealed that only constructs able to interact with the class 5 kinesin-like motor protein Eg5 induce the spindle phenotypes. Importantly, injection of Eg5 into TPX2-CT-arrested blastomeres causes resumption of cleavage. These results define a discrete domain within the C terminus of TPX2 that exerts a novel Eg5-dependent function in spindle pole segregation.
Eckerdt
Frank
F
Howard Hughes Medical Institute, Department of Pharmacology, University of Colorado School of Medicine, Aurora, Colorado 80045, USA.
Eyers
Patrick A
PA
Lewellyn
Andrea L
AL
Prigent
Claude
C
Maller
James L
JL
D000818:Animals; D018797:Cell Cycle Proteins / Q000378:metabolism*; D002460:Cell Line; D004625:Embryo, Nonmammalian / Q000378:metabolism / Q000502:physiology; D047108:Embryonic Development / Q000502:physiology*; D016547:Kinesins / Q000378:metabolism*; D008869:Microtubule-Associated Proteins / Q000378:metabolism*; D008938:Mitosis / Q000502:physiology*; D009363:Neoplasm Proteins / Q000378:metabolism*; D009687:Nuclear Proteins / Q000378:metabolism*; D010750:Phosphoproteins / Q000378:metabolism*; D054730:Protein Interaction Domains and Motifs / Q000502:physiology; D008941:Spindle Apparatus / Q000378:metabolism* / Q000502:physiology; D014981:Xenopus; D029867:Xenopus Proteins / Q000378:metabolism*
D018797:Cell Cycle Proteins; C088335:KIF11 protein, Xenopus; D008869:Microtubule-Associated Proteins; D009363:Neoplasm Proteins; D009687:Nuclear Proteins; D010750:Phosphoproteins; C486551:TPX2 protein, Xenopus; D029867:Xenopus Proteins; D016547:Kinesins