Changing Roles of Aurora-B Kinase in Two Life Cycle Stages of Trypanosoma brucei

Abstract

Aurora-B kinase is a chromosomal passenger protein essential for chromosome segregation and cytokinesis. In the procyclic form of Trypanosoma brucei, depletion of an aurora-B kinase homologue TbAUK1 inhibited spindle formation, mitosis, cytokinesis, and organelle replication without altering cell morphology. In the present study, an RNA interference knockdown of TbAUK1 or overexpression of inactive mutant TbAUK1-K58R in the bloodstream form also resulted in defects in spindle formation, chromosome segregation, and cytokinesis but allowed multiple rounds of nuclear DNA synthesis, nucleolus multiplication, and continuous replication of kinetoplast, basal body, and flagellum. The typical trypanosome morphology was lost to an enlarged round shape filled with microtubules. It is thus apparent that there are distinctive mechanisms of action of TbAUK1 in regulating cell division between the two developmental stages of trypanosome. While it exerts a tight control on mitosis, organelle replication, and cytokinesis in the procyclic form, it regulates cytokinesis without rigid control over either nuclear DNA synthesis or organelle replication in the bloodstream form. The molecular basis underlining these discrepancies remains to be explored.Trypanosoma brucei is an ancient unicellular eukaryote whose propagation involves a cyclic transmission between the mammalian host and the insect vector tsetse fly. A trypanosome cell contains a single flagellum, basal body, nucleus, mitochondrion, and the mitochondrial DNA complex known as the kinetoplast. Each of these organelles/structures is replicated and segregated into the two daughter cells in a wellsynchronized manner during cell division, implying a delicate coordination among these events (25). Other than the usual sequential G 1 , S, G 2 , and M phases, there is also an S phase for the kinetoplast completed prior to the nuclear S phase (45). The kinetoplast is replicated prior to mitosis and closely associated with the flagellum basal body, with its segregation dependent on the separation of replicated basal bodies, leading the cell through cytokinesis and eventual cell division in good coordination with nuclear division (30).Treatment of the procyclic form of T. brucei cells with okadaic acid prevented kinetoplast/basal body segregation and resulted in multinucleated cells, suggesting a normal progression of mitosis in cells defective in cytokinesis, which is controlled largely by the kinetoplast cycle (7). Treatment with aphidicolin or rhizoxin (29) or RNA interference (RNAi) silencing of the G 1 or mitotic cyclins (15, 24) or the cdc2-related kinases (CRKs) (37) in the procyclic-form cells resulted in inhibition of nuclear division but not kinetoplast/basal body segregation and subsequent cytokinesis, resulting in anucleate daughter cells designated the zoids. These results suggest that the checkpoint linking mitosis with cytokinesis may be weak or absent in the procyclic form of T. brucei and that kinetoplast/ basal body segregation alone is sufficient to drive some of t...

Tousled-like kinase in a microbial eukaryote regulates spindle assembly and S-phase progression by interacting with Aurora kinase and chromatin assembly factors

“…However, as TLK1 regulates chromosome segregation in cooperation with AUK1 (Fig. 5), which is also required for spindle assembly in T. brucei (Tu et al, 2006;Li and Wang, 2006), TLK1 might regulate spindle assembly by being a substrate activator of AUK1, as was observed in C. elegans (Han et al, 2005).…”

“…The 1N2K cells were apparently defective in chromosome segregation, as monitored with the L1C6 antibody (Fig. 1D, left panel), which stains the nucleolus and serves as a cellular marker for chromosome segregation in T. brucei (Ogbadoyi et al, 2000;Durand-Dubief and Bastin, 2003;Li and Wang, 2006). The control 1N2K cell has a segregating nucleolus, whereas the nucleolus merely becomes slightly enlarged in the TLK1-deficient cell (Fig.…”

“…In summary, TLK1 cooperates with Aurora kinase to regulate spindle assembly and chromosome segregation, and it performs a role in DNA replication probably by regulating histone modification in trypanosomes. (Hammarton et al, 2003;Tu and Wang, 2004;Li and Wang, 2006). …”

“…An Aurora kinase homolog, AUK1, is essential for spindle formation, chromosome segregation and cytokinesis in both bloodstream and procyclic forms of T. brucei (Tu et al, 2006;Li and Wang, 2006), suggesting that, like the Aurora kinase Ark1 in Schizosaccharomyces pombe, AUK1 has the functions of both the Aurora A and Aurora B kinases possessed by the higher eukaryotes (Petersen et al, 2001). AUK1 displays a subcellular localization pattern typical of a chromosomal passenger -that is, being concentrated in the nucleus during prophase but translocated to the spindle midzone in late anaphase (Tu et al, 2006;Li and Wang, 2006).…”

“…AUK1 displays a subcellular localization pattern typical of a chromosomal passenger -that is, being concentrated in the nucleus during prophase but translocated to the spindle midzone in late anaphase (Tu et al, 2006;Li and Wang, 2006). This localization is mediated by the formation of a chromosomal passenger complex between Aurora B kinase and several other proteins such as Survivin, Borealin/Dasra and INCENP, the substrate activator of Aurora B kinase, in metazoa (Ducat and Zheng, 2004).…”