Localization of Human TACC3 to Mitotic Spindles Is Mediated by Phosphorylation on Ser558 by Aurora A: A Novel Pharmacodynamic Method for Measuring Aurora A Activity

Abstract

Aurora A is a serine/threonine protein kinase essential for normal mitotic progression. Aberrant increased expression of Aurora A, which occurs frequently in human cancers, results in abnormal mitoses leading to chromosome instability and possibly tumorigenesis. Consequently, Aurora A has received considerable attention as a potential target for anticancer therapeutic intervention. Aurora A coordinates several essential mitotic activities through phosphorylation of a variety of proteins, including TACC3, which modulates microtubule stabilization of the mitotic spindle. Recent studies identified a conserved serine in Xenopus (Ser 626 ) and Drosophila (Ser 863 ) TACC3 orthologues that is phosphorylated by Aurora A. We show that this conserved serine on human TACC3 (Ser 558 ) is also phosphorylated by Aurora A. Moreover, phosphorylation of TACC3 by Aurora A in human cells is essential for its proper localization to centrosomes and proximal mitotic spindles. Inhibition of Aurora A with the selective small molecule inhibitor MLN8054 in cultured human tumor cells resulted in mislocalization of TACC3 away from mitotic spindles in a concentration-dependent manner. Furthermore, oral administration of MLN8054 to nude mice bearing HCT-116 human tumor xenografts caused a dose-dependent mislocalization of TACC3 away from spindle poles that correlated with tumor growth inhibition. As TACC3 localization to mitotic spindles depends on Aurora A-mediated phosphorylation, quantifying TACC3 mislocalization represents a novel pharmacodynamic approach for measuring Aurora A activity in cancer patients treated with inhibitors of Aurora A kinase. [Cancer Res 2007;67(11):5362-70]

Mitotic drivers—inhibitors of the Aurora B Kinase

“…Consistent with established roles for Aurora A, MLN8054 induces abnormal spindles, often with unseparated centrosomes, and delays progression through mitosis [31]. This is accompanied by reduced phosphorylation and localization of the Aurora A substrate TACC3 [32]. These observations are entirely consistent with data from model systems, indicating that MLN8054 has potential, not only as a new chemical-biology tool for probing Aurora A function, but also as a new antimitotic agent.…”

The centrosomal protein TACC3 controls paclitaxel sensitivity by modulating a premature senescence program

“…Interestingly, while the relative distribution of ERK1/2 Model summarizing the role of the Aurora A-TACC3 axis in regulating commitment to premature senescence. Aurora A kinase phosphorylates TACC3 and thereby regulates its mitotic localization (LeRoy et al, 2007). Interestingly, (i) small molecule-mediated inhibition of Aurora A by MLN8054 or Aurora A depletion (Huck et al), (ii) knockdown of TACC3 (this study), and (iii) reduction of the centromeric protein CENP-A (Maehara et al), all trigger a comparable premature senescence response preventing propagation of defective mitoses.…”

“…Whereas TACC3-deficient murine fibroblasts with functional post-mitotic checkpoints enter a reversible G 1 and G 2 arrest, checkpointcompromised cells rapidly succumb to mitotic cell death and polyploidization (Schneider et al, 2007(Schneider et al, , 2008. Given that centrosomal localization of human TACC3 requires its phosphorylation by Aurora A kinase at Ser558 (LeRoy et al, 2007) and because pharmacological inhibition of Aurora A kinase induces premature senescence (Huck et al, 2010), we explored here (i) whether senescence is the prevailing outcome of TACC3 reduction in G 1 checkpoint-proficient transformed and non-transformed breast epithelial cells and (ii) to which extent their chemosensitivity against PTX can be modulated by targeting TACC3 expression.…”

Aurora A kinase and its substrate TACC3 are required for central spindle assembly

“…To determine whether TACC3 phosphorylation by AurA is required for central spindle formation, we performed rescue experiments by expressing either Flag-TACC3 WT or Flag-TACC3 S558A, a phosphorylation mutant on the AurA site, [16] in TACC3-silenced cells (supplementary Fig S3C online) and analysed the cells by immunofluorescence quantifying the central spindle MT fluorescence intensities (Fig 3D,E). TACC3 silencing promoted a substantial reduction of central spindle MT density.…”