TPX2-LIKE PROTEIN3 Is the Primary Activator of α-Aurora Kinases and Is Essential for Embryogenesis

Abstract

Aurora kinases are key regulators of mitosis. Multicellular eukaryotes generally possess two functionally diverged types of Aurora kinases. In plants, including Arabidopsis (Arabidopsis thaliana), these are termed aand b-Auroras. As the functional specification of Aurora kinases is determined by their specific interaction partners, we initiated interactomics analyses using both Arabidopsis a-Aurora kinases (AUR1 and AUR2). Proteomics results revealed that TPX2-LIKE PROTEINS2 and 3 (TPXL2/3) prominently associated with a-Auroras, as did the conserved TPX2 to a lower degree. Like TPX2, TPXL2 and TPXL3 strongly activated the AUR1 kinase but exhibited cell-cycle-dependent localization differences on microtubule arrays. The separate functions of TPX2 and TPXL2/3 were also suggested by their different influences on AUR1 localization upon ectopic expressions. Furthermore, genetic analyses showed that TPXL3, but not TPX2 and TPXL2, acts nonredundantly to enable proper embryo development. In contrast to vertebrates, plants have an expanded TPX2 family and these family members have both redundant and unique functions. Moreover, as neither TPXL2 nor TPXL3 contains the C-terminal Kinesin-5 binding domain present in the canonical TPX2, the targeting and activity of this kinesin must be organized differently in plants.

Functional Divergence of Microtubule-Associated TPX2 Family Members in Arabidopsis thaliana

“…Taken together, our data confirmed the validity of our hypothesis that TPXL genes might have evolved different functions during plant development. Similar expression patterns for TPX2, TPXL2, TPXL3, and TPXL5 point to possible functional redundancy [18]. Moreover, these four proteins are expressed in a similar pattern to Aurora1 and Aurora2 in agreement with the fact that they are physiological activators of the kinases.…”

“…It should also be noted that compared to animals, plants usually encode two genes of α-Aurora kinases [5]. Although we do not have any data about the specificity of complex formation between different α-Auroras and TPXL proteins during development we have already shown the interaction of AUR1 and AUR2 with TPXL2 and TPXL3 [18]. On the other hand, unlike animals, plants show a high level of endopolyploidization, which is highly specific for different tissues and different stages of development.…”

“…Similarly, Aurora kinase binding domain of canonical Arabidopsis TPX2 can activate Aurora1 in vitro [15]. Recently we showed that Aurora kinase binding domains of TPXL2 and TPLX3 also can activate Aurora1 in vitro [18]. To characterize the Aurora binding domain of plant group A TPXL proteins, a multiple sequence alignment was performed with orthologues of Arabidopsis TPX2.…”

“…The Aurora kinase binding domains of canonical Arabidopsis TPX2, TPXL2, and TPXL3 were shown to activate Aurora1 in vitro and increase its phosphorylation activity towards histone H3 as a physiological substrate [15,18]. To address the functionality of the Aurora kinase binding domains of Arabidopsis TPXL proteins, we performed in vitro kinase assays.…”

“…In contrast to analyzed genomes of animals, where only one TPX2 gene was found, Arabidopsis encodes 20 genes with sequence homology to some of the domains of canonical TPX2. The first members of the TPX2-LIKE PROTEIN (TPXL) gene family in Arabidopsis, TPXL2 and TPXL3, were functionally characterized only recently [18]. In plants, outside Arabidopsis, a TPXL gene family containing 12 members was described in Eucalyptus grandis [19].…”