Exploring the Functional Interactions between Aurora B, INCENP, and Survivin in Mitosis

Abstract

The function of the Aurora B kinase at centromeres and the central spindle is crucial for chromosome segregation and cytokinesis, respectively. Herein, we have investigated the regulation of human Aurora B by its complex partners inner centromere protein (INCENP) and survivin. We found that overexpression of a catalytically inactive, dominant-negative mutant of Aurora B impaired the localization of the entire Aurora B/INCENP/survivin complex to centromeres and the central spindle and severely disturbed mitotic progression. Similar results were also observed after depletion, by RNA interference, of either Aurora B, INCENP, or survivin. These data suggest that Aurora B kinase activity and the formation of the Aurora B/INCENP/survivin complex both contribute to its proper localization. Using recombinant proteins, we found that Aurora B kinase activity was stimulated by INCENP and that the C-terminal region of INCENP was sufficient for activation. Under identical assay conditions, survivin did not detectably influence kinase activity. Human INCENP was a substrate of Aurora B and mass spectrometry identified three consecutive residues (threonine 893, serine 894, and serine 895) containing at least two phosphorylation sites. A nonphosphorylatable mutant (TSS893-895AAA) was a poor activator of Aurora B, demonstrating that INCENP phosphorylation is important for kinase activation.

Subcellular localization and nucleocytoplasmic transport of the chromosomal passenger proteins before nuclear envelope breakdown

“…Aurora B undergoes CRM1-mediated nuclear export The nucleocytoplasmic distribution of Aurora B fused to a MYC tag, similar in size to the VSV and HA epitopes used here, has been previously shown to be predominantly nuclear in U2OS cells (Honda et al, 2003), and both nuclear and cytoplasmic in COS-7 cells (Kawajiri et al, 2003;Yasui et al, 2004). These previous observations, and the heterogeneous distribution we found in MCF-7 cells, suggested that the nucleocytoplasmic distribution of Aurora B before NEB could be a dynamic process.…”

“…INCENP is a large protein that has been proposed to act as a scaffold for the binding of the other passenger proteins (Bolton et al, 2002). Aurora B phosphorylates INCENP, which, in turn, activates the kinase activity of Aurora B in a positive feedback loop (Bishop and Schumacher, 2002;Honda et al, 2003). Survivin is a member of the inhibitor of apoptosis protein family that plays a dual role as regulator of both apoptosis and cell division (reviewed by Altieri, 2003a).…”

Chromosomes with delayed replication timing lead to checkpoint activation, delayed recruitment of Aurora B and chromosome instability

“…Three members of the passenger complex are: INCENP, Aurora B kinase and survivin. INCENP, a microtubule-binding protein is responsible for binding Aurora B (Honda et al, 2003). Survivin has an essential role in chromosome alignment (Lens et al, 2003) and is important during mitosis for proper localization of INCENP and Aurora B (reviewed by Carmena and Earnshaw, 2003).…”

Expression of cytoplasmic and nuclear Survivin in primary and secondary human glioblastoma

“…Since Survivin has both nuclear and cytoplasmic targets, which appear to serve different functions (Mahotka et al, 2002), two different commercially available Survivin antibodies, which recognise predominantly nuclear and cytoplasmic forms of Survivin, were used (Altura et al, 2003). Nuclear Survivin has been suggested to play an important role in chromosomal segregation during mitosis Honda et al, 2003), while cytoplasmic Survivin has been characterised as antiapoptotic (O'Connor et al, 2002). In addition, the TUNEL assay was used to detect the apoptotic status of the GBMs and apoptotic status was correlated with the expression of different Survivin forms.…”