Abstract
The essential mammalian gene TACC3 is frequently mutated and amplified in cancers and its fusion products exhibit oncogenic activity in glioblastomas. TACC3 functions in mitotic spindle assembly and chromosome segregation. In particular, phosphorylation on S558 by the mitotic kinase, Aurora-A, promotes spindle recruitment of TACC3 and triggers the formation of a complex with ch-TOG-clathrin that crosslinks and stabilises kinetochore microtubules. Here we map the Aurora-A-binding interface in TACC3 and show that TACC3 potently activates Aurora-A through a domain centered on F525. Vertebrate cells carrying homozygous F525A mutation in the endogenous TACC3 loci exhibit defects in TACC3 function, namely perturbed localization, reduced phosphorylation and weakened interaction with clathrin. The most striking feature of the F525A cells however is a marked shortening of mitosis, at least in part due to rapid spindle assembly. F525A cells do not exhibit chromosome missegregation, indicating that they undergo fast yet apparently faithful mitosis. By contrast, mutating the phosphorylation site S558 to alanine in TACC3 causes aneuploidy without a significant change in mitotic duration. Our work has therefore defined a regulatory role for the Aurora-A-TACC3 interaction beyond the act of phosphorylation at S558. We propose that the regulatory relationship between Aurora-A and TACC3 enables the transition from the microtubule-polymerase activity of TACC3-ch-TOG to the microtubule-crosslinking activity of TACC3-ch-TOG-clathrin complexes as mitosis progresses. Aurora-A-dependent control of TACC3 could determine the balance between these activities, thereby influencing not only spindle length and stability but also the speed of spindle formation with vital consequences for chromosome alignment and segregation.
Highlights
Formation of a functional mitotic spindle is a pre-requisite for equal distribution of chromosomes between two daughter cells
We identify a new binding interface between Aurora-A and TACC3 that is crucial for assembly of TACC3-clathrin complexes and proper spindle localization of TACC3
We conclude that mutual regulation between Aurora-A kinase and its substrate TACC3 constitutes a molecular switch between different spindle assembly pathways, thereby influencing the speed of spindle formation
Summary
Formation of a functional mitotic spindle is a pre-requisite for equal distribution of chromosomes between two daughter cells. When present the centrosomes act as dominant sites of spindle pole formation, but in all animal cells MT motors and MT-associated proteins play important roles in stabilizing, organising and transporting MTs generated elsewhere in the cytoplasm for incorporation into the mitotic spindle [2,3]. A range of additional phenotypes such as short bipolar and multipolar spindles, MT hyper-stabilisation, centrosome and spindle pole fragmentation or chromosome alignment and segregation defects have been reported in several model systems [7,8,9,10,11,12]. Aurora-A exerts its control over spindle assembly through phosphorylation of several substrates, including TACC3 [14,15]
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