Spatiotemporal organization of Aurora-B by APC/CCdh1 after mitosis coordinates cell spreading via FHOD1

Abstract

Spatiotemporal regulation of mitotic kinase activity underlies the extensive rearrangement of cellular components required for cell division. One highly dynamic mitotic kinase is Aurora kinase B (AurB), which has multiple roles defined by the changing localization of the chromosome passenger complex (CPC) as cells progress through mitosis, including regulation of cytokinesis and abscission. Like other mitotic kinases, AurB is a target of the anaphase-promoting complex (APC/C) ubiquitin ligase during mitotic exit, but it is not known if APC/C-mediated destruction plays any specific role in controling AurB activity. Here we have examined the contribution of APC/CCdh1 to organization of AurB activity as cells exit mitosis and re-enter interphase. We report that APC/CCdh1-dependent proteolysis restricts a cell cortex-associated pool of active AurB in space and time. In early G1 phase this pool of AurB is found at protrusions associated with cell spreading. AurB retention at the cortex depends on a formin, FHOD1, critically required to organize the cytoskeleton after division. We identify AurB phosphorylation sites in FHOD1 and show that phosphomutant FHOD1 is impaired in post-mitotic assembly of oriented actin cables. We propose that Cdh1 contributes to spatiotemporal organization of AurB activity and that organization of FHOD1 activity by AurB contributes to daughter cell spreading after mitosis.

VRK1 and AURKB form a complex that cross inhibit their kinase activity and the phosphorylation of histone H3 in the progression of mitosis

“…AURKB is degraded at the end of mitosis by ubiquitin-mediated degradation in the proteasome [24, 37, 38]. Therefore, it was tested whether depletion of VRK1 might affect the stability of AURKB.…”

Cytokinetic abscission is an acute G1 event

“…In support of this model, it was recently shown that the APC/C Cdh1 -mediated degradation of AuroraB controls cell adhesion and spreading occurring »45 min after anaphase onset. 24 Interference with this process causes severe abscission delays. 25 It is, therefore, possible that continuous AuroraB degradation in G1 is critical for abscission onset through the regulation of cell spreading.…”

Using in Vivo Biotinylated Ubiquitin to Describe a Mitotic Exit Ubiquitome from Human Cells

“…U2OS-bio Ub cells and U2OS-BirA cells were obtained via clonal selection with 170 g/ml Hygromycin B (Calbiochem) after transfection of U2OS tet-OFF cells with pTREbio Ub 6 -BirA or pTRE-BirA in a ratio of 10:1 with pCMVhygro and were supplied with 5 M biotin in culture. The hTERT-RPE1 (RPE) cell line stably expressing ␣-actinin-Venus was described previously (33). Cells were synchronized to prometaphase by sequential blocks with 25 mM thymidine and 10 M S-trityl-L-cysteine (STLC, Tocris Bioscience, Bristol, UK) for 20 h and 16 h, respectively, and collected via mitotic shake-off.…”

“…We then compared cell spreading in cells expressing wildtype and non-ubiquitinable versions of mCherry-RacGAP1 using hTERT-RPE ␣-actinin-Venus cells in which cell spreading can readily be visualized and quantified (33). Cells electroporated with RacGAP1 constructs were recorded through mitosis by means of fluorescence time-lapse imaging, and the areas of daughter cells, normalized for the area at metaphase, were plotted against time.…”