Abstract
Overexpression of AURKA is a major hallmark of epithelial cancers. It encodes the multifunctional serine/threonine kinase aurora A, which is activated at metaphase and is required for cell cycle progression; assessing its activation in living cells is mandatory for next-generation drug design. We describe here a Förster's resonance energy transfer (FRET) biosensor detecting the conformational changes of aurora kinase A induced by its autophosphorylation on Thr288. The biosensor functionally replaces the endogenous kinase in cells and allows the activation of the kinase to be followed throughout the cell cycle. Inhibiting the catalytic activity of the kinase prevents the conformational changes of the biosensor. Using this approach, we discover that aurora kinase A activates during G1 to regulate the stability of microtubules in cooperation with TPX2 and CEP192. These results demonstrate that the aurora kinase A biosensor is a powerful tool to identify new regulatory pathways controlling aurora kinase A activation.
Highlights
Overexpression of AURKA is a major hallmark of epithelial cancers
We provide a functional validation of the AURKA biosensor by showing that it replaces the endogenous kinase in living cells
We investigated whether this conformational change could be tracked in space and time by Forster’s resonance energy transfer (FRET) microscopy
Summary
Overexpression of AURKA is a major hallmark of epithelial cancers. It encodes the multifunctional serine/threonine kinase aurora A, which is activated at metaphase and is required for cell cycle progression; assessing its activation in living cells is mandatory for next-generation drug design. Inhibiting the catalytic activity of the kinase prevents the conformational changes of the biosensor Using this approach, we discover that aurora kinase A activates during G1 to regulate the stability of microtubules in cooperation with TPX2 and CEP192. The fidelity of centrosomal functions is controlled by the interplay of several molecular actors, including centrosome-residing and non-residing proteins that cooperate in promoting spindle assembly and stability These proteins include mitotic kinases in charge of cell cycle progression[2] such as the serine/threonine kinase AURKA. This protein regulates the duplication and the maturation of the centrosomes, the correct timing for mitotic entry, the assembly of the mitotic spindle and cytokinesis[3]. AURKA is required to regulate the stability of the microtubule network in an interplay with TPX2 and the centrosomal protein of 192 kDa CEP192
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