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

Joanna Boruc,Bo Liu,Nienke Besbrugge,Dominique Eeckhout,Eva Dvořák Tomaštíková,Xingguang Deng,Matthias Van Durme,Tong-Reen Connie Tan,Dmitri Demidov,Moritz K Nowack,Tom Beeckman,Honghui Lin,Michaël Vandorpe,Daniël Van Damme,Geert De Jaeger,Evelien Mylle

doi:10.1104/pp.18.01515

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 α- and β-Auroras. As the functional specification of Aurora kinases is determined by their specific interaction partners, we initiated interactomics analyses using both Arabidopsis α-Aurora kinases (AUR1 and AUR2). Proteomics results revealed that TPX2-LIKE PROTEINS2 and 3 (TPXL2/3) prominently associated with α-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.

Highlights

Aurora kinases are key regulators of mitosis
We conclude that in contrast to TPX2, which we could only identify using the CGSrhino and CGSyellow tags coupled with the more sensitive LTQ detection, these TPX-Like proteins (TPXLs) were found in our experiments using the GSTEV tags combined with MALDI detection and can be considered as bona-fide interactors of both AUR1 and AUR2
Our results showed that in Arabidopsis, representing dicotyledonous plants, the expansion of the TPX2 protein family has led to the selection of TPXL3 as an essential MTassociated proteins (MAPs) while the canonical TPX2 has become dispensable

Summary

Introduction

Aurora kinases are key regulators of mitosis. Multicellular eukaryotes generally possess two functionally diverged types of Aurora kinases. AUR B, on the other hand, predominantly interacts with the chromosomal passenger complex (containing the inner centromere protein INCEN-P, Survivin, and Borealin) This complex associates with the centromeres in early stages of mitosis and translocates to MTs in the central spindle from anaphase through cytokinesis (for review, see Carmena and Earnshaw, 2003). Upon NEBD, a high RanGTP concentration, and high levels of free TPX2, are maintained around the chromosomes due to the association of the RanGEF (Ras-related nuclear protein guanine nucleotide exchange factor) Regulator of chromosome condensation with chromatin These RanGTP and TPX2 gradients create a positional cue that determines the site of TPX2-mediated MT nucleation (for review, see Neumayer et al, 2014). Arabidopsis TPX2 has MT nucleation capacity in vitro, and overexpression of TPX2 causes ectopic intranuclear MT nucleation in vivo that is independent of Aurora (Vos et al, 2008; Petrovská et al, 2013), suggesting that TPX2 is essential for bipolar spindle formation in plants, like in animals

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