Regulation of microtubule-based microtubule nucleation by mammalian polo-like kinase 1

Abstract

Bipolar spindle formation is pivotal for accurate segregation of mitotic chromosomes during cell division. A growing body of evidence suggests that, in addition to centrosome-and chromatinbased microtubule (MT) nucleation, MT-based MT nucleation plays an important role for proper bipolar spindle formation in various eukaryotic organisms. Although a recently discovered Augmin complex appears to play a central role in this event, how Augmin is regulated remains unknown. Here we provide evidence that a mammalian polo-like kinase 1 (Plk1) localizes to mitotic spindles and promotes MT-based MT nucleation by directly regulating Augmin. Mechanistically, we demonstrated that Cdc2-dependent phosphorylation on a γ-tubulin ring complex (γ-TuRC) recruitment protein, Nedd1/GCP-WD, at the previously uncharacterized S460 residue induces the Nedd1-Plk1 interaction. This step appeared to be critical to allow Plk1 to phosphorylate the Hice1 subunit of the Augmin complex to promote the Augmin-MT interaction and MT-based MT nucleation from within the spindle. Loss of either the Nedd1 S460 function or the Plk1-dependent Hice1 phosphorylation impaired both the Augmin-MT interaction and γ-tubulin recruitment to the spindles, thus resulting in improper bipolar spindle formation that ultimately leads to mitotic arrest and apoptotic cell death. Thus, via the formation of the Nedd1-Plk1 complex and subsequent Augmin phosphorylation, Plk1 regulates spindle MT-based MT nucleation to accomplish normal bipolar spindle formation and mitotic progression. mitosis | spindle assembly | polo-box domain | distributive phosphorylation | Ser/Thr protein kinase

The where, when and how of microtubule nucleation – one ring to rule them all

“…Tethers cTuRCs to unattached kinetochores to support nucleation of kinetochore microtubules (Mishra et al, 2010) Plk1 Binds and phosphorylates GCP-WD subunit in mitosis; might contribute to cTuRC recruitment to the centrosome (Haren et al, 2009;Johmura et al, 2011;Zhang et al, 2009 Associate with cTuRCs and phosphorylate cTuRC-associated proteins to promote microtubule nucleation (Dráberová et al, 1999;Kukharskyy et al, 2004;Macurek et al, 2008;Sulimenko et al, 2006) BRCA1 E3 ligase activity ubiquitylates c-tubulin to inhibit centrosomal nucleation activity (Sankaran et al, 2005;Starita et al, 2004) …”

Microtubule assembly during mitosis – from distinct origins to distinct functions?

“…The mitotic kinases cyclin-dependent kinase 1 (CDK1), polo-like kinase 1 (PLK1) and Aurora A have been shown to be involved in c-tubulin recruitment to the centrosome (Blagden and Glover, 2003). PLK1 and Aurora A activities are involved in c-TuRC recruitment by the Augmin complex (Johmura et al, 2011;Tsai et al, 2011) (Tables 2, 3). Although the mechanism of c-tubulin targeting in the chromosomal pathway remains poorly understood, Aurora A kinase is also required for MT nucleation and stabilization around chromatin (Bayliss et al, 2003;Sardon et al, 2008) (Table 3).…”

“…The targeting of c-tubulin to each MT nucleation site is, at least in part, directed by the c-TuRCassociated protein NEDD1 (neural precursor cell expressed, developmentally down-regulated 1, also known as GCP7) (Lüders et al, 2006;Zhu et al, 2008) (Table 2). Interestingly, NEDD1 is hyperphosphorylated in mitosis, and its activity is differentially regulated by mitotic kinases at the centrosomes and in the acentrosomal MT amplification pathway (Johmura et al, 2011;Zhang et al, 2009).…”

“…This is fundamental for our understanding of how, in only a few minutes, the cell is able to build a machinery as complex as the mitotic spindle. (Gomez-Ferreria et al, 2007;Raynaud-Messina and Merdes, 2007) Augmin complex, FAM29A (Bucciarelli et al, 2009) TPX2, RHAMM (also known as HMMR) (Groen et al, 2004) Targeting regulators PLK1, Aurora A, CDK1 (Barr and Gergely, 2007) PLK1, Aurora A (Johmura et al, 2011;Tsai et al, 2011) Ran-GTP, Aurora A Tsai et al, 2003) In the upper part of the table, the proteins involved specifically in c-tubulin targeting to the MT nucleation sites are indicated for each of the three pathways. In the lower part of the table, the main regulators of c-tubulin recruitment for each MT assembly pathway are shown.…”

TONNEAU2/FASS Regulates the Geometry of Microtubule Nucleation and Cortical Array Organization in Interphase Arabidopsis Cells

“…Studies on spindle formation in animal cells have shown that targeting of g-TURCs to the centrosome and spindle microtubules is regulated during the cell cycle. The cascade of phosphorylation events triggered by cyclin-dependent kinase 1 (CDK1) and of polo-like kinase 1 (PLK1) has been demonstrated to promote binding of the g-TURC targeting factor GCP-WD to spindle microtubules, facilitating the acentrosomal nucleation important during spindle formation (Lü ders et al, 2006;Zhang et al, 2009;Johmura et al, 2011). In addition to CDK1 and PLK1, several other kinases and phosphatases were found to be involved in the regulation of microtubule nucleation at centrosomes (Fry et al, 1998;Horn et al, 2007;Kim et al, 2007;Sardon et al, 2008), highlighting the important role of protein phosphorylation in microtubule organization.…”