PLK4 promotes centriole duplication by phosphorylating STIL to link the procentriole cartwheel to the microtubule wall.

Tyler Chistopher Moyer,Andrew Jon Holland

doi:10.7554/elife.46054

Tyler Chistopher Moyer, Andrew Jon Holland

Open Access

https://doi.org/10.7554/elife.46054

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Journal: eLife	Publication Date: May 22, 2019
Citations: 60	License type: CC BY 4.0

Affiliation: Johns Hopkins Medicine, Johns Hopkins University

Abstract

Centrioles play critical roles in organizing the assembly of the mitotic spindle and templating the formation of primary cilia. Centriole duplication occurs once per cell cycle and is regulated by Polo-like kinase 4 (PLK4). Although significant progress has been made in understanding centriole composition, we have limited knowledge of how PLK4 activity controls specific steps in centriole formation. Here, we show that PLK4 phosphorylates its centriole substrate STIL on a conserved site, S428, to promote STIL binding to CPAP. This phospho-dependent binding interaction is conserved in Drosophila and facilitates the stable incorporation of both STIL and CPAP into the centriole. We propose that procentriole assembly requires PLK4 to phosphorylate STIL in two different regions: phosphorylation of residues in the STAN motif allow STIL to bind SAS6 and initiate cartwheel assembly, while phosphorylation of S428 promotes the binding of STIL to CPAP, linking the cartwheel to microtubules of the centriole wall.

Highlights

Centrioles are microtubule-based structures that recruit a surrounding pericentriolar material (PCM) to form the centrosome (Nigg and Holland, 2018; Gonczy, 2012)
To determine whether phosphorylation of STIL by Polo-like kinase 4 (PLK4) might affect the interaction of STIL with other components of the centriole duplication machinery, we tested the ability of Myc-GFP-STIL to interact with its known centriolar binding partners in the presence of kinase active (PLK4WT) or kinase dead (PLK4KD) PLK4
Of the 84 in vitro phosphorylation sites we identified on STIL, S428 was of particular interest as it is highly conserved, matches the PLK4 consensus phosphorylation sequence and is positioned close to the known CPAP binding region on STIL (Figure 2A, Figure 2—figure supplement 1) (Cottee et al, 2013; Kettenbach et al, 2012; Johnson et al, 2007; Hatzopoulos et al, 2013)

Summary

Introduction

Centrioles are microtubule-based structures that recruit a surrounding pericentriolar material (PCM) to form the centrosome (Nigg and Holland, 2018; Gonczy, 2012). Centrosomes nucleate the formation of the microtubule cytoskeleton in interphase cells and form the poles of the mitotic spindle during cell division. In late G2 phase, the two centriole pairs separate and increase PCM recruitment to promote the formation of the mitotic spindle. Defects in centriole biogenesis can result in the formation of supernumerary centrosomes which promote mitotic errors that can contribute to tumorigenesis (Levine et al, 2017; Levine and Holland, 2018; Basto et al, 2008; Sercin et al, 2016; Coelho et al, 2015). Mutations in centriole proteins are linked to growth retardation syndromes and autosomal recessive primary microcephaly (MPCH) in human patients (Nigg and Raff, 2009; Chavali et al, 2014)

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